Aging-like Changes in the Transcriptome of Irradiated Microglia

PubMed Central

Li, Matthew D.; Burns, Terry C.; Kumar, Sunny; Morgan, Alexander A.; Sloan, Steven A.; Palmer, Theo D.

2014-01-01

Whole brain irradiation remains important in the management of brain tumors. Although necessary for improving survival outcomes, cranial irradiation also results in cognitive decline in long-term survivors. A chronic inflammatory state characterized by microglial activation has been implicated in radiation-induced brain injury. We here provide the first comprehensive transcriptional profile of irradiated microglia. Fluorescence-activated cell sorting (FACS) was used to isolate CD11b+ microglia from the hippocampi of C57BL/6 and Balb/c mice 1 month after 10Gy cranial irradiation. Affymetrix gene expression profiles were evaluated using linear modeling, rank product analyses. One month after irradiation, a conserved irradiation signature across strains was identified, comprising 448 and 85 differentially up- and down-regulated genes, respectively. Gene set enrichment analysis (GSEA) demonstrated enrichment for inflammation, including M1 macrophage-associated genes, but also an unexpected enrichment for extracellular matrix and blood coagulation-related gene sets, in contrast previously described microglial states. Weighted gene co-expression network analysis (WGCNA) confirmed these findings and further revealed alterations in mitochondrial function. The RNA-seq transcriptome of microglia 24h post-radiation proved similar to the 1-month transcriptome, but additionally featured alterations in apoptotic and lysosomal gene expression. Re-analysis of published aging mouse microglia transcriptome data demonstrated striking similarity to the 1 month irradiated microglia transcriptome, suggesting that shared mechanisms may underlie aging and chronic irradiation-induced cognitive decline. PMID:25690519

Expression signature as a biomarker for prenatal diagnosis of trisomy 21.

PubMed

Volk, Marija; Maver, Aleš; Lovrečić, Luca; Juvan, Peter; Peterlin, Borut

2013-01-01

A universal biomarker panel with the potential to predict high-risk pregnancies or adverse pregnancy outcome does not exist. Transcriptome analysis is a powerful tool to capture differentially expressed genes (DEG), which can be used as biomarker-diagnostic-predictive tool for various conditions in prenatal setting. In search of biomarker set for predicting high-risk pregnancies, we performed global expression profiling to find DEG in Ts21. Subsequently, we performed targeted validation and diagnostic performance evaluation on a larger group of case and control samples. Initially, transcriptomic profiles of 10 cultivated amniocyte samples with Ts21 and 9 with normal euploid constitution were determined using expression microarrays. Datasets from Ts21 transcriptomic studies from GEO repository were incorporated. DEG were discovered using linear regression modelling and validated using RT-PCR quantification on an independent sample of 16 cases with Ts21 and 32 controls. The classification performance of Ts21 status based on expression profiling was performed using supervised machine learning algorithm and evaluated using a leave-one-out cross validation approach. Global gene expression profiling has revealed significant expression changes between normal and Ts21 samples, which in combination with data from previously performed Ts21 transcriptomic studies, were used to generate a multi-gene biomarker for Ts21, comprising of 9 gene expression profiles. In addition to biomarker's high performance in discriminating samples from global expression profiling, we were also able to show its discriminatory performance on a larger sample set 2, validated using RT-PCR experiment (AUC=0.97), while its performance on data from previously published studies reached discriminatory AUC values of 1.00. Our results show that transcriptomic changes might potentially be used to discriminate trisomy of chromosome 21 in the prenatal setting. As expressional alterations reflect both, causal and reactive cellular mechanisms, transcriptomic changes may thus have future potential in the diagnosis of a wide array of heterogeneous diseases that result from genetic disturbances.

Deep sequencing-based transcriptome profiling analysis of bacteria-challenged Lateolabrax japonicus reveals insight into the immune-relevant genes in marine fish

PubMed Central

2010-01-01

Background Systematic research on fish immunogenetics is indispensable in understanding the origin and evolution of immune systems. This has long been a challenging task because of the limited number of deep sequencing technologies and genome backgrounds of non-model fish available. The newly developed Solexa/Illumina RNA-seq and Digital gene expression (DGE) are high-throughput sequencing approaches and are powerful tools for genomic studies at the transcriptome level. This study reports the transcriptome profiling analysis of bacteria-challenged Lateolabrax japonicus using RNA-seq and DGE in an attempt to gain insights into the immunogenetics of marine fish. Results RNA-seq analysis generated 169,950 non-redundant consensus sequences, among which 48,987 functional transcripts with complete or various length encoding regions were identified. More than 52% of these transcripts are possibly involved in approximately 219 known metabolic or signalling pathways, while 2,673 transcripts were associated with immune-relevant genes. In addition, approximately 8% of the transcripts appeared to be fish-specific genes that have never been described before. DGE analysis revealed that the host transcriptome profile of Vibrio harveyi-challenged L. japonicus is considerably altered, as indicated by the significant up- or down-regulation of 1,224 strong infection-responsive transcripts. Results indicated an overall conservation of the components and transcriptome alterations underlying innate and adaptive immunity in fish and other vertebrate models. Analysis suggested the acquisition of numerous fish-specific immune system components during early vertebrate evolution. Conclusion This study provided a global survey of host defence gene activities against bacterial challenge in a non-model marine fish. Results can contribute to the in-depth study of candidate genes in marine fish immunity, and help improve current understanding of host-pathogen interactions and evolutionary history of immunogenetics from fish to mammals. PMID:20707909

Global Analysis of Transcriptome Responses and Gene Expression Profiles to Cold Stress of Jatropha curcas L.

PubMed Central

Wang, Haibo; Zou, Zhurong; Wang, Shasha; Gong, Ming

2013-01-01

Background Jatropha curcas L., also called the Physic nut, is an oil-rich shrub with multiple uses, including biodiesel production, and is currently exploited as a renewable energy resource in many countries. Nevertheless, because of its origin from the tropical MidAmerican zone, J. curcas confers an inherent but undesirable characteristic (low cold resistance) that may seriously restrict its large-scale popularization. This adaptive flaw can be genetically improved by elucidating the mechanisms underlying plant tolerance to cold temperatures. The newly developed Illumina Hiseq™ 2000 RNA-seq and Digital Gene Expression (DGE) are deep high-throughput approaches for gene expression analysis at the transcriptome level, using which we carefully investigated the gene expression profiles in response to cold stress to gain insight into the molecular mechanisms of cold response in J. curcas. Results In total, 45,251 unigenes were obtained by assembly of clean data generated by RNA-seq analysis of the J. curcas transcriptome. A total of 33,363 and 912 complete or partial coding sequences (CDSs) were determined by protein database alignments and ESTScan prediction, respectively. Among these unigenes, more than 41.52% were involved in approximately 128 known metabolic or signaling pathways, and 4,185 were possibly associated with cold resistance. DGE analysis was used to assess the changes in gene expression when exposed to cold condition (12°C) for 12, 24, and 48 h. The results showed that 3,178 genes were significantly upregulated and 1,244 were downregulated under cold stress. These genes were then functionally annotated based on the transcriptome data from RNA-seq analysis. Conclusions This study provides a global view of transcriptome response and gene expression profiling of J. curcas in response to cold stress. The results can help improve our current understanding of the mechanisms underlying plant cold resistance and favor the screening of crucial genes for genetically enhancing cold resistance in J. curcas. PMID:24349370

Global analysis of transcriptome responses and gene expression profiles to cold stress of Jatropha curcas L.

PubMed

Wang, Haibo; Zou, Zhurong; Wang, Shasha; Gong, Ming

2013-01-01

Jatropha curcas L., also called the Physic nut, is an oil-rich shrub with multiple uses, including biodiesel production, and is currently exploited as a renewable energy resource in many countries. Nevertheless, because of its origin from the tropical MidAmerican zone, J. curcas confers an inherent but undesirable characteristic (low cold resistance) that may seriously restrict its large-scale popularization. This adaptive flaw can be genetically improved by elucidating the mechanisms underlying plant tolerance to cold temperatures. The newly developed Illumina Hiseq™ 2000 RNA-seq and Digital Gene Expression (DGE) are deep high-throughput approaches for gene expression analysis at the transcriptome level, using which we carefully investigated the gene expression profiles in response to cold stress to gain insight into the molecular mechanisms of cold response in J. curcas. In total, 45,251 unigenes were obtained by assembly of clean data generated by RNA-seq analysis of the J. curcas transcriptome. A total of 33,363 and 912 complete or partial coding sequences (CDSs) were determined by protein database alignments and ESTScan prediction, respectively. Among these unigenes, more than 41.52% were involved in approximately 128 known metabolic or signaling pathways, and 4,185 were possibly associated with cold resistance. DGE analysis was used to assess the changes in gene expression when exposed to cold condition (12°C) for 12, 24, and 48 h. The results showed that 3,178 genes were significantly upregulated and 1,244 were downregulated under cold stress. These genes were then functionally annotated based on the transcriptome data from RNA-seq analysis. This study provides a global view of transcriptome response and gene expression profiling of J. curcas in response to cold stress. The results can help improve our current understanding of the mechanisms underlying plant cold resistance and favor the screening of crucial genes for genetically enhancing cold resistance in J. curcas.

Next-generation sequencing facilitates quantitative analysis of wild-type and Nrl−/− retinal transcriptomes

PubMed Central

Brooks, Matthew J.; Rajasimha, Harsha K.; Roger, Jerome E.

2011-01-01

Purpose Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis. Methods Retinal mRNA profiles of 21-day-old wild-type (WT) and neural retina leucine zipper knockout (Nrl−/−) mice were generated by deep sequencing, in triplicate, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays. Results Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the retinas of WT and Nrl−/− mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRT–PCR. RNA-seq data had a linear relationship with qRT–PCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and Nrl−/− retina, with a fold change ≥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling. Conclusions Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions. PMID:22162623

A Transcriptome Meta-Analysis Proposes Novel Biological Roles for the Antifungal Protein AnAFP in Aspergillus niger

PubMed Central

Schäpe, Paul; Müller-Hagen, Dirk; Ouedraogo, Jean-Paul; Heiderich, Caroline; Jedamzick, Johanna; van den Hondel, Cees A.; Ram, Arthur F.; Meyer, Vera

2016-01-01

Understanding the genetic, molecular and evolutionary basis of cysteine-stabilized antifungal proteins (AFPs) from fungi is important for understanding whether their function is mainly defensive or associated with fungal growth and development. In the current study, a transcriptome meta-analysis of the Aspergillus niger γ-core protein AnAFP was performed to explore co-expressed genes and pathways, based on independent expression profiling microarrays covering 155 distinct cultivation conditions. This analysis uncovered that anafp displays a highly coordinated temporal and spatial transcriptional profile which is concomitant with key nutritional and developmental processes. Its expression profile coincides with early starvation response and parallels with genes involved in nutrient mobilization and autophagy. Using fluorescence- and luciferase reporter strains we demonstrated that the anafp promoter is active in highly vacuolated compartments and foraging hyphal cells during carbon starvation with CreA and FlbA, but not BrlA, as most likely regulators of anafp. A co-expression network analysis supported by luciferase-based reporter assays uncovered that anafp expression is embedded in several cellular processes including allorecognition, osmotic and oxidative stress survival, development, secondary metabolism and autophagy, and predicted StuA and VelC as additional regulators. The transcriptomic resources available for A. niger provide unparalleled resources to investigate the function of proteins. Our work illustrates how transcriptomic meta-analyses can lead to hypotheses regarding protein function and predict a role for AnAFP during slow growth, allorecognition, asexual development and nutrient recycling of A. niger and propose that it interacts with the autophagic machinery to enable these processes. PMID:27835655

A Transcriptome Meta-Analysis Proposes Novel Biological Roles for the Antifungal Protein AnAFP in Aspergillus niger.

PubMed

Paege, Norman; Jung, Sascha; Schäpe, Paul; Müller-Hagen, Dirk; Ouedraogo, Jean-Paul; Heiderich, Caroline; Jedamzick, Johanna; Nitsche, Benjamin M; van den Hondel, Cees A; Ram, Arthur F; Meyer, Vera

2016-01-01

Understanding the genetic, molecular and evolutionary basis of cysteine-stabilized antifungal proteins (AFPs) from fungi is important for understanding whether their function is mainly defensive or associated with fungal growth and development. In the current study, a transcriptome meta-analysis of the Aspergillus niger γ-core protein AnAFP was performed to explore co-expressed genes and pathways, based on independent expression profiling microarrays covering 155 distinct cultivation conditions. This analysis uncovered that anafp displays a highly coordinated temporal and spatial transcriptional profile which is concomitant with key nutritional and developmental processes. Its expression profile coincides with early starvation response and parallels with genes involved in nutrient mobilization and autophagy. Using fluorescence- and luciferase reporter strains we demonstrated that the anafp promoter is active in highly vacuolated compartments and foraging hyphal cells during carbon starvation with CreA and FlbA, but not BrlA, as most likely regulators of anafp. A co-expression network analysis supported by luciferase-based reporter assays uncovered that anafp expression is embedded in several cellular processes including allorecognition, osmotic and oxidative stress survival, development, secondary metabolism and autophagy, and predicted StuA and VelC as additional regulators. The transcriptomic resources available for A. niger provide unparalleled resources to investigate the function of proteins. Our work illustrates how transcriptomic meta-analyses can lead to hypotheses regarding protein function and predict a role for AnAFP during slow growth, allorecognition, asexual development and nutrient recycling of A. niger and propose that it interacts with the autophagic machinery to enable these processes.

The vagal ganglia transcriptome identifies candidate therapeutics for airway hyperreactivity.

PubMed

Reznikov, Leah R; Meyerholz, David K; Abou Alaiwa, Mahmoud H; Kuan, Shin-Ping; Liao, Yan-Shin J; Bormann, Nicholas L; Bair, Thomas B; Price, Margaret; Stoltz, David A; Welsh, Michael J

2018-04-05

Mainstay therapeutics are ineffective in some people with asthma, suggesting a need for additional agents. In the current study, we used vagal ganglia transcriptome profiling and connectivity mapping to identify compounds beneficial for alleviating airway hyperreactivity. As a comparison, we also utilized previously published transcriptome data from sensitized mouse lungs and human asthmatic endobronchial biopsies. All transcriptomes revealed agents beneficial for mitigating airway hyperreactivity; however, only the vagal ganglia transcriptome identified agents used clinically to treat asthma (flunisolide, isoetarine). We also tested one compound identified by vagal ganglia transcriptome profiling that had not previously been linked to asthma and found that it had bronchodilator effects in both mouse and pig airways. These data suggest that transcriptome profiling of the vagal ganglia might be a novel strategy to identify potential asthma therapeutics.

Transcriptome Analysis of Chlorantraniliprole Resistance Development in the Diamondback Moth Plutella xylostella

PubMed Central

Hu, Zhendi; Chen, Huanyu; Yin, Fei; Li, Zhenyu; Dong, Xiaolin; Zhang, Deyong; Ren, Shunxiang; Feng, Xia

2013-01-01

Background The diamondback moth Plutella xyllostella has developed a high level of resistance to the latest insecticide chlorantraniliprole. A better understanding of P. xylostella’s resistance mechanism to chlorantraniliprole is needed to develop effective approaches for insecticide resistance management. Principal Findings To provide a comprehensive insight into the resistance mechanisms of P. xylostella to chlorantraniliprole, transcriptome assembly and tag-based digital gene expression (DGE) system were performed using Illumina HiSeq™ 2000. The transcriptome analysis of the susceptible strain (SS) provided 45,231 unigenes (with the size ranging from 200 bp to 13,799 bp), which would be efficient for analyzing the differences in different chlorantraniliprole-resistant P. xylostella stains. DGE analysis indicated that a total of 1215 genes (189 up-regulated and 1026 down-regulated) were gradient differentially expressed among the susceptible strain (SS) and different chlorantraniliprole-resistant P. xylostella strains, including low-level resistance (GXA), moderate resistance (LZA) and high resistance strains (HZA). A detailed analysis of gradient differentially expressed genes elucidated the existence of a phase-dependent divergence of biological investment at the molecular level. The genes related to insecticide resistance, such as P450, GST, the ryanodine receptor, and connectin, had different expression profiles in the different chlorantraniliprole-resistant DGE libraries, suggesting that the genes related to insecticide resistance are involved in P. xylostella resistance development against chlorantraniliprole. To confirm the results from the DGE, the expressional profiles of 4 genes related to insecticide resistance were further validated by qRT-PCR analysis. Conclusions The obtained transcriptome information provides large gene resources available for further studying the resistance development of P. xylostella to pesticides. The DGE data provide comprehensive insights into the gene expression profiles of the different chlorantraniliprole-resistant stains. These genes are specifically related to insecticide resistance, with different expressional profiles facilitating the study of the role of each gene in chlorantraniliprole resistance development. PMID:23977278

Gene Expression Analysis of Copper Tolerance and Wood Decay in the Brown Rot Fungus Fibroporia radiculosa

Treesearch

J. D. Tang; L. A. Parker; A. D. Perkins; T. S. Sonstegard; S. G. Schroeder; D. D. Nicholas; S. V. Diehl

2013-01-01

High-throughput transcriptomics was used to identify Fibroporia radiculosa genes that were differentially regulated during colonization of wood treated with a copper-based preservative. The transcriptome was profiled at two time points while the fungus was growing on wood treated with micronized copper quat (MCQ). A total of 917 transcripts were...

Developmental Transcriptome for a Facultatively Eusocial Bee, Megalopta genalis

PubMed Central

Jones, Beryl M.; Wcislo, William T.; Robinson, Gene E.

2015-01-01

Transcriptomes provide excellent foundational resources for mechanistic and evolutionary analyses of complex traits. We present a developmental transcriptome for the facultatively eusocial bee Megalopta genalis, which represents a potential transition point in the evolution of eusociality. A de novo transcriptome assembly of Megalopta genalis was generated using paired-end Illumina sequencing and the Trinity assembler. Males and females of all life stages were aligned to this transcriptome for analysis of gene expression profiles throughout development. Gene Ontology analysis indicates that stage-specific genes are involved in ion transport, cell–cell signaling, and metabolism. A number of distinct biological processes are upregulated in each life stage, and transitions between life stages involve shifts in dominant functional processes, including shifts from transcriptional regulation in embryos to metabolism in larvae, and increased lipid metabolism in adults. We expect that this transcriptome will provide a useful resource for future analyses to better understand the molecular basis of the evolution of eusociality and, more generally, phenotypic plasticity. PMID:26276382

Developmental Transcriptome for a Facultatively Eusocial Bee, Megalopta genalis.

PubMed

Jones, Beryl M; Wcislo, William T; Robinson, Gene E

2015-08-14

Transcriptomes provide excellent foundational resources for mechanistic and evolutionary analyses of complex traits. We present a developmental transcriptome for the facultatively eusocial bee Megalopta genalis, which represents a potential transition point in the evolution of eusociality. A de novo transcriptome assembly of Megalopta genalis was generated using paired-end Illumina sequencing and the Trinity assembler. Males and females of all life stages were aligned to this transcriptome for analysis of gene expression profiles throughout development. Gene Ontology analysis indicates that stage-specific genes are involved in ion transport, cell-cell signaling, and metabolism. A number of distinct biological processes are upregulated in each life stage, and transitions between life stages involve shifts in dominant functional processes, including shifts from transcriptional regulation in embryos to metabolism in larvae, and increased lipid metabolism in adults. We expect that this transcriptome will provide a useful resource for future analyses to better understand the molecular basis of the evolution of eusociality and, more generally, phenotypic plasticity. Copyright © 2015 Jones et al.

Transcriptome In Vivo Analysis (TIVA) of spatially defined single cells in intact live mouse and human brain tissue

PubMed Central

Lovatt, Ditte; Ruble, Brittani K.; Lee, Jaehee; Dueck, Hannah; Kim, Tae Kyung; Fisher, Stephen; Francis, Chantal; Spaethling, Jennifer M.; Wolf, John A.; Grady, M. Sean; Ulyanova, Alexandra V.; Yeldell, Sean B.; Griepenburg, Julianne C.; Buckley, Peter T.; Kim, Junhyong; Sul, Jai-Yoon; Dmochowski, Ivan J.; Eberwine, James

2014-01-01

Transcriptome profiling is an indispensable tool in advancing the understanding of single cell biology, but depends upon methods capable of isolating mRNA at the spatial resolution of a single cell. Current capture methods lack sufficient spatial resolution to isolate mRNA from individual in vivo resident cells without damaging adjacent tissue. Because of this limitation, it has been difficult to assess the influence of the microenvironment on the transcriptome of individual neurons. Here, we engineered a Transcriptome In Vivo Analysis (TIVA)-tag, which upon photoactivation enables mRNA capture from single cells in live tissue. Using the TIVA-tag in combination with RNA-seq to analyze transcriptome variance among single dispersed cells and in vivo resident mouse and human neurons, we show that the tissue microenvironment shapes the transcriptomic landscape of individual cells. The TIVA methodology provides the first noninvasive approach for capturing mRNA from single cells in their natural microenvironment. PMID:24412976

Safety assessment of plant varieties using transcriptomics profiling and a one-class classifier.

PubMed

van Dijk, Jeroen P; de Mello, Carla Souza; Voorhuijzen, Marleen M; Hutten, Ronald C B; Arisi, Ana Carolina Maisonnave; Jansen, Jeroen J; Buydens, Lutgarde M C; van der Voet, Hilko; Kok, Esther J

2014-10-01

An important part of the current hazard identification of novel plant varieties is comparative targeted analysis of the novel and reference varieties. Comparative analysis will become much more informative with unbiased analytical approaches, e.g. omics profiling. Data analysis estimating the similarity of new varieties to a reference baseline class of known safe varieties would subsequently greatly facilitate hazard identification. Further biological and eventually toxicological analysis would then only be necessary for varieties that fall outside this reference class. For this purpose, a one-class classifier tool was explored to assess and classify transcriptome profiles of potato (Solanum tuberosum) varieties in a model study. Profiles of six different varieties, two locations of growth, two year of harvest and including biological and technical replication were used to build the model. Two scenarios were applied representing evaluation of a 'different' variety and a 'similar' variety. Within the model higher class distances resulted for the 'different' test set compared with the 'similar' test set. The present study may contribute to a more global hazard identification of novel plant varieties. Copyright © 2014 Elsevier Inc. All rights reserved.

Transcriptome profile and unique genetic evolution of positively selected genes in yak lungs.

PubMed

Lan, DaoLiang; Xiong, XianRong; Ji, WenHui; Li, Jian; Mipam, Tserang-Donko; Ai, Yi; Chai, ZhiXin

2018-04-01

The yak (Bos grunniens), which is a unique bovine breed that is distributed mainly in the Qinghai-Tibetan Plateau, is considered a good model for studying plateau adaptability in mammals. The lungs are important functional organs that enable animals to adapt to their external environment. However, the genetic mechanism underlying the adaptability of yak lungs to harsh plateau environments remains unknown. To explore the unique evolutionary process and genetic mechanism of yak adaptation to plateau environments, we performed transcriptome sequencing of yak and cattle (Bos taurus) lungs using RNA-Seq technology and a subsequent comparison analysis to identify the positively selected genes in the yak. After deep sequencing, a normal transcriptome profile of yak lung that containing a total of 16,815 expressed genes was obtained, and the characteristics of yak lungs transcriptome was described by functional analysis. Furthermore, Ka/Ks comparison statistics result showed that 39 strong positively selected genes are identified from yak lungs. Further GO and KEGG analysis was conducted for the functional annotation of these genes. The results of this study provide valuable data for further explorations of the unique evolutionary process of high-altitude hypoxia adaptation in yaks in the Tibetan Plateau and the genetic mechanism at the molecular level.

Transcriptome Analysis at the Single-Cell Level Using SMART Technology.

PubMed

Fish, Rachel N; Bostick, Magnolia; Lehman, Alisa; Farmer, Andrew

2016-10-10

RNA sequencing (RNA-seq) is a powerful method for analyzing cell state, with minimal bias, and has broad applications within the biological sciences. However, transcriptome analysis of seemingly homogenous cell populations may in fact overlook significant heterogeneity that can be uncovered at the single-cell level. The ultra-low amount of RNA contained in a single cell requires extraordinarily sensitive and reproducible transcriptome analysis methods. As next-generation sequencing (NGS) technologies mature, transcriptome profiling by RNA-seq is increasingly being used to decipher the molecular signature of individual cells. This unit describes an ultra-sensitive and reproducible protocol to generate cDNA and sequencing libraries directly from single cells or RNA inputs ranging from 10 pg to 10 ng. Important considerations for working with minute RNA inputs are given. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.

RNA-Stabilized Whole Blood Samples but Not Peripheral Blood Mononuclear Cells Can Be Stored for Prolonged Time Periods Prior to Transcriptome Analysis

PubMed Central

Debey-Pascher, Svenja; Hofmann, Andrea; Kreusch, Fatima; Schuler, Gerold; Schuler-Thurner, Beatrice; Schultze, Joachim L.; Staratschek-Jox, Andrea

2011-01-01

Microarray-based transcriptome analysis of peripheral blood as surrogate tissue has become an important approach in clinical implementations. However, application of gene expression profiling in routine clinical settings requires careful consideration of the influence of sample handling and RNA isolation methods on gene expression profile outcome. We evaluated the effect of different sample preservation strategies (eg, cryopreservation of peripheral blood mononuclear cells or freezing of PAXgene-stabilized whole blood samples) on gene expression profiles. Expression profiles obtained from cryopreserved peripheral blood mononuclear cells differed substantially from those of their nonfrozen counterpart samples. Furthermore, expression profiles in cryopreserved peripheral blood mononuclear cell samples were found to undergo significant alterations with increasing storage period, whereas long-term freezing of PAXgene RNA stabilized whole blood samples did not significantly affect stability of gene expression profiles. This report describes important technical aspects contributing toward the establishment of robust and reliable guidance for gene expression studies using peripheral blood and provides a promising strategy for reliable implementation in routine handling for diagnostic purposes. PMID:21704280

Defining the transcriptome assembly and its use for genome dynamics and transcriptome profiling studies in pigeonpea (Cajanus cajan L.)

USDA-ARS?s Scientific Manuscript database

This study reports generation of large-scale genomic resources for pigeonpea, a so-called ‘orphan crop species’ of the semi-arid tropic regions. Roche FLX/454 sequencing was carried out on a normalized cDNA pool prepared from 31 tissues produced 494,353 short transcript reads (STRs). Cluster analysi...

RNA-Seq-based transcriptome analysis of dormant flower buds of Chinese cherry (Prunus pseudocerasus).

PubMed

Zhu, Youyin; Li, Yongqiang; Xin, Dedong; Chen, Wenrong; Shao, Xu; Wang, Yue; Guo, Weidong

2015-01-25

Bud dormancy is a critical biological process allowing Chinese cherry (Prunus pseudocerasus) to survive in winter. Due to the lake of genomic information, molecular mechanisms triggering endodormancy release in flower buds have remained unclear. Hence, we used Illumina RNA-Seq technology to carry out de novo transcriptome assembly and digital gene expression profiling of flower buds. Approximately 47million clean reads were assembled into 50,604 sequences with an average length of 837bp. A total of 37,650 unigene sequences were successfully annotated. 128 pathways were annotated by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and metabolic, biosynthesis of second metabolite and plant hormone signal transduction accounted for higher percentage in flower bud. In critical period of endodormancy release, 1644, significantly differentially expressed genes (DEGs) were identified from expression profile. DEGs related to oxidoreductase activity were especially abundant in Gene Ontology (GO) molecular function category. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis demonstrated that DEGs were involved in various metabolic processes, including phytohormone metabolism. Quantitative real-time PCR (qRT-PCR) analysis indicated that levels of DEGs for abscisic acid and gibberellin biosynthesis decreased while the abundance of DEGs encoding their degradation enzymes increased and GID1 was down-regulated. Concomitant with endodormancy release, MADS-box transcription factors including P. pseudocerasus dormancy-associated MADS-box (PpcDAM), Agamous-like2, and APETALA3-like genes, shown remarkably epigenetic roles. The newly generated transcriptome and gene expression profiling data provide valuable genetic information for revealing transcriptomic variation during bud dormancy in Chinese cherry. The uncovered data should be useful for future studies of bud dormancy in Prunus fruit trees lacking genomic information. Copyright © 2014 Elsevier B.V. All rights reserved.

The Human Pancreas Proteome Defined by Transcriptomics and Antibody-Based Profiling

PubMed Central

Fagerberg, Linn; Hallström, Björn M.; Schwenk, Jochen M.; Uhlén, Mathias; Korsgren, Olle; Lindskog, Cecilia

2014-01-01

The pancreas is composed of both exocrine glands and intermingled endocrine cells to execute its diverse functions, including enzyme production for digestion of nutrients and hormone secretion for regulation of blood glucose levels. To define the molecular constituents with elevated expression in the human pancreas, we employed a genome-wide RNA sequencing analysis of the human transcriptome to identify genes with elevated expression in the human pancreas. This quantitative transcriptomics data was combined with immunohistochemistry-based protein profiling to allow mapping of the corresponding proteins to different compartments and specific cell types within the pancreas down to the single cell level. Analysis of whole pancreas identified 146 genes with elevated expression levels, of which 47 revealed a particular higher expression as compared to the other analyzed tissue types, thus termed pancreas enriched. Extended analysis of in vitro isolated endocrine islets identified an additional set of 42 genes with elevated expression in these specialized cells. Although only 0.7% of all genes showed an elevated expression level in the pancreas, this fraction of transcripts, in most cases encoding secreted proteins, constituted 68% of the total mRNA in pancreas. This demonstrates the extreme specialization of the pancreas for production of secreted proteins. Among the elevated expression profiles, several previously not described proteins were identified, both in endocrine cells (CFC1, FAM159B, RBPJL and RGS9) and exocrine glandular cells (AQP12A, DPEP1, GATM and ERP27). In summary, we provide a global analysis of the pancreas transcriptome and proteome with a comprehensive list of genes and proteins with elevated expression in pancreas. This list represents an important starting point for further studies of the molecular repertoire of pancreatic cells and their relation to disease states or treatment effects. PMID:25546435

Integrated analysis of transcriptome and lipid profiling reveals the co-influences of inositol-choline and Snf1 in controlling lipid biosynthesis in yeast.

PubMed

Chumnanpuen, Pramote; Zhang, Jie; Nookaew, Intawat; Nielsen, Jens

2012-07-01

In the yeast Saccharomyces cerevisiae many genes involved in lipid biosynthesis are transcriptionally controlled by inositol-choline and the protein kinase Snf1. Here we undertook a global study on how inositol-choline and Snf1 interact in controlling lipid metabolism in yeast. Using both a reference strain (CEN.PK113-7D) and a snf1Δ strain cultured at different nutrient limitations (carbon and nitrogen), at a fixed specific growth rate of 0.1 h(-1), and at different inositol choline concentrations, we quantified the expression of genes involved in lipid biosynthesis and the fluxes towards the different lipid components. Through integrated analysis of the transcriptome, the lipid profiling and the fluxome, it was possible to obtain a high quality, large-scale dataset that could be used to identify correlations and associations between the different components. At the transcription level, Snf1 and inositol-choline interact either directly through the main phospholipid-involving transcription factors (i.e. Ino2, Ino4, and Opi1) or through other transcription factors e.g. Gis1, Mga2, and Hac1. However, there seems to be flux regulation at the enzyme levels of several lipid involving enzymes. The analysis showed the strength of using both transcriptome and lipid profiling analysis for mapping the co-influence of inositol-choline and Snf1 on phospholipid metabolism.

Transcriptomics in cancer diagnostics: developments in technology, clinical research and commercialization.

PubMed

Sager, Monica; Yeat, Nai Chien; Pajaro-Van der Stadt, Stefan; Lin, Charlotte; Ren, Qiuyin; Lin, Jimmy

2015-01-01

Transcriptomic technologies are evolving to diagnose cancer earlier and more accurately to provide greater predictive and prognostic utility to oncologists and patients. Digital techniques such as RNA sequencing are replacing still-imaging techniques to provide more detailed analysis of the transcriptome and aberrant expression that causes oncogenesis, while companion diagnostics are developing to determine the likely effectiveness of targeted treatments. This article examines recent advancements in molecular profiling research and technology as applied to cancer diagnosis, clinical applications and predictions for the future of personalized medicine in oncology.

Use of homologous and heterologous gene expression profiling tools to characterize transcription dynamics during apple fruit maturation and ripening.

PubMed

Costa, Fabrizio; Alba, Rob; Schouten, Henk; Soglio, Valeria; Gianfranceschi, Luca; Serra, Sara; Musacchi, Stefano; Sansavini, Silviero; Costa, Guglielmo; Fei, Zhangjun; Giovannoni, James

2010-10-25

Fruit development, maturation and ripening consists of a complex series of biochemical and physiological changes that in climacteric fruits, including apple and tomato, are coordinated by the gaseous hormone ethylene. These changes lead to final fruit quality and understanding of the functional machinery underlying these processes is of both biological and practical importance. To date many reports have been made on the analysis of gene expression in apple. In this study we focused our investigation on the role of ethylene during apple maturation, specifically comparing transcriptomics of normal ripening with changes resulting from application of the hormone receptor competitor 1-methylcyclopropene. To gain insight into the molecular process regulating ripening in apple, and to compare to tomato (model species for ripening studies), we utilized both homologous and heterologous (tomato) microarray to profile transcriptome dynamics of genes involved in fruit development and ripening, emphasizing those which are ethylene regulated.The use of both types of microarrays facilitated transcriptome comparison between apple and tomato (for the later using data previously published and available at the TED: tomato expression database) and highlighted genes conserved during ripening of both species, which in turn represent a foundation for further comparative genomic studies. The cross-species analysis had the secondary aim of examining the efficiency of heterologous (specifically tomato) microarray hybridization for candidate gene identification as related to the ripening process. The resulting transcriptomics data revealed coordinated gene expression during fruit ripening of a subset of ripening-related and ethylene responsive genes, further facilitating the analysis of ethylene response during fruit maturation and ripening. Our combined strategy based on microarray hybridization enabled transcriptome characterization during normal climacteric apple ripening, as well as definition of ethylene-dependent transcriptome changes. Comparison with tomato fruit maturation and ethylene responsive transcriptome activity facilitated identification of putative conserved orthologous ripening-related genes, which serve as an initial set of candidates for assessing conservation of gene activity across genomes of fruit bearing plant species.

Isolation of Polysomal RNA for Analyzing Stress-Responsive Genes Regulated at the Translational Level in Plants.

PubMed

Li, Yong-Fang; Mahalingam, Ramamurthy; Sunkar, Ramanjulu

2017-01-01

Alteration of gene expression is an essential mechanism, which allows plants to respond and adapt to adverse environmental conditions. Transcriptome and proteome analyses in plants exposed to abiotic stresses revealed that protein levels are not correlated with the changes in corresponding mRNAs, indicating regulation at translational level is another major regulator for gene expression. Analysis of translatome, which refers to all mRNAs associated with ribosomes, thus has the potential to bridge the gap between transcriptome and proteome. Polysomal RNA profiling and recently developed ribosome profiling (Ribo-seq) are two main methods for translatome analysis at global level. Here, we describe the classical procedure for polysomal RNA isolation by sucrose gradient ultracentrifugation followed by highthroughput RNA-seq to identify genes regulated at translational level. Polysomal RNA can be further used for a variety of downstream applications including Northern blot analysis, qRT-PCR, RNase protection assay, and microarray-based gene expression profiling.

Linking gene regulation and the exo-metabolome: A comparative transcriptomics approach to identify genes that impact on the production of volatile aroma compounds in yeast

PubMed Central

Rossouw, Debra; Næs, Tormod; Bauer, Florian F

2008-01-01

Background 'Omics' tools provide novel opportunities for system-wide analysis of complex cellular functions. Secondary metabolism is an example of a complex network of biochemical pathways, which, although well mapped from a biochemical point of view, is not well understood with regards to its physiological roles and genetic and biochemical regulation. Many of the metabolites produced by this network such as higher alcohols and esters are significant aroma impact compounds in fermentation products, and different yeast strains are known to produce highly divergent aroma profiles. Here, we investigated whether we can predict the impact of specific genes of known or unknown function on this metabolic network by combining whole transcriptome and partial exo-metabolome analysis. Results For this purpose, the gene expression levels of five different industrial wine yeast strains that produce divergent aroma profiles were established at three different time points of alcoholic fermentation in synthetic wine must. A matrix of gene expression data was generated and integrated with the concentrations of volatile aroma compounds measured at the same time points. This relatively unbiased approach to the study of volatile aroma compounds enabled us to identify candidate genes for aroma profile modification. Five of these genes, namely YMR210W, BAT1, AAD10, AAD14 and ACS1 were selected for overexpression in commercial wine yeast, VIN13. Analysis of the data show a statistically significant correlation between the changes in the exo-metabome of the overexpressing strains and the changes that were predicted based on the unbiased alignment of transcriptomic and exo-metabolomic data. Conclusion The data suggest that a comparative transcriptomics and metabolomics approach can be used to identify the metabolic impacts of the expression of individual genes in complex systems, and the amenability of transcriptomic data to direct applications of biotechnological relevance. PMID:18990252

Transcriptome Profiling of the Abdominal Skin of Larimichthys crocea in Light Stress

NASA Astrophysics Data System (ADS)

Han, Zhaofang; Lv, Changhuan; Xiao, Shijun; Ye, Kun; Zhang, Dongling; Tsai, Huai Jen; Wang, Zhiyong

2018-04-01

Large yellow croaker ( Larimichthys crocea), one of the most important marine fish species in China, can change its abdominal skin color when it is shifted from light to dark or from dark to light, providing us an opportunity of investigating the molecular responding mechanism of teleost in light stress. The gene expression profile of fish under light stress is rarely documented. In this research, the transcriptome profiles of the abdominal skin of L. crocea exposed to light or dark for 0 h, 0.5 h and 2 h were produced by next-generation sequencing (NGS). The cluster results demonstrated that stress period, rather than light intensity ( e.g., light or dark), is the major influencing factor. Differently expressed genes (DEGs) were identified between 0 h and 0.5 h groups, between 0 h and 2 h groups, between 0.5 h light and 0.5 h dark, and between 2 h light and 2 h dark, respectively. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation revealed that the genes relating to immunity, energy metabolism, and cytoskeletal protein binding were significantly enriched. The detailed analysis of transcriptome profiles also revealed regular gene expression trends, indicating that the elaborate gene regulation networks underlined the molecular responses of the fish to light stress. This transcriptome analysis suggested that systematic and complicated regulatory cascades were functionally activated in response to external stress, and coloration change caused by light stress was mainly attributed to the change in the density of chromatophores for L. crocea. This study also provided valuable information for skin coloration or light stress research on other marine fish species.

Transcriptome analysis of Brassica napus pod using RNA-Seq and identification of lipid-related candidate genes.

PubMed

Xu, Hai-Ming; Kong, Xiang-Dong; Chen, Fei; Huang, Ji-Xiang; Lou, Xiang-Yang; Zhao, Jian-Yi

2015-10-24

Brassica napus is an important oilseed crop. Dissection of the genetic architecture underlying oil-related biological processes will greatly facilitates the genetic improvement of rapeseed. The differential gene expression during pod development offers a snapshot on the genes responsible for oil accumulation in. To identify candidate genes in the linkage peaks reported previously, we used RNA sequencing (RNA-Seq) technology to analyze the pod transcriptomes of German cultivar Sollux and Chinese inbred line Gaoyou. The RNA samples were collected for RNA-Seq at 5-7, 15-17 and 25-27 days after flowering (DAF). Bioinformatics analysis was performed to investigate differentially expressed genes (DEGs). Gene annotation analysis was integrated with QTL mapping and Brassica napus pod transcriptome profiling to detect potential candidate genes in oilseed. Four hundred sixty five and two thousand, one hundred fourteen candidate DEGs were identified, respectively, between two varieties at the same stages and across different periods of each variety. Then, 33 DEGs between Sollux and Gaoyou were identified as the candidate genes affecting seed oil content by combining those DEGs with the quantitative trait locus (QTL) mapping results, of which, one was found to be homologous to Arabidopsis thaliana lipid-related genes. Intervarietal DEGs of lipid pathways in QTL regions represent important candidate genes for oil-related traits. Integrated analysis of transcriptome profiling, QTL mapping and comparative genomics with other relative species leads to efficient identification of most plausible functional genes underlying oil-content related characters, offering valuable resources for bettering breeding program of Brassica napus. This study provided a comprehensive overview on the pod transcriptomes of two varieties with different oil-contents at the three developmental stages.

Gene expression analysis of induced pluripotent stem cells from aneuploid chromosomal syndromes

PubMed Central

2013-01-01

Background Human aneuploidy is the leading cause of early pregnancy loss, mental retardation, and multiple congenital anomalies. Due to the high mortality associated with aneuploidy, the pathophysiological mechanisms of aneuploidy syndrome remain largely unknown. Previous studies focused mostly on whether dosage compensation occurs, and the next generation transcriptomics sequencing technology RNA-seq is expected to eventually uncover the mechanisms of gene expression regulation and the related pathological phenotypes in human aneuploidy. Results Using next generation transcriptomics sequencing technology RNA-seq, we profiled the transcriptomes of four human aneuploid induced pluripotent stem cell (iPSC) lines generated from monosomy × (Turner syndrome), trisomy 8 (Warkany syndrome 2), trisomy 13 (Patau syndrome), and partial trisomy 11:22 (Emanuel syndrome) as well as two umbilical cord matrix iPSC lines as euploid controls to examine how phenotypic abnormalities develop with aberrant karyotype. A total of 466 M (50-bp) reads were obtained from the six iPSC lines, and over 13,000 mRNAs were identified by gene annotation. Global analysis of gene expression profiles and functional analysis of differentially expressed (DE) genes were implemented. Over 5000 DE genes are determined between aneuploidy and euploid iPSCs respectively while 9 KEGG pathways are overlapped enriched in four aneuploidy samples. Conclusions Our results demonstrate that the extra or missing chromosome has extensive effects on the whole transcriptome. Functional analysis of differentially expressed genes reveals that the genes most affected in aneuploid individuals are related to central nervous system development and tumorigenesis. PMID:24564826

Dose-Response Analysis of RNA-Seq Profiles in Archival Formalin-Fixed Paraffin-Embedded (FFPE) Samples.

EPA Science Inventory

Use of archival resources has been limited to date by inconsistent methods for genomic profiling of degraded RNA from formalin-fixed paraffin-embedded (FFPE) samples. RNA-sequencing offers a promising way to address this problem. Here we evaluated transcriptomic dose responses us...

Defining the Human Macula Transcriptome and Candidate Retinal Disease Genes UsingEyeSAGE

PubMed Central

Rickman, Catherine Bowes; Ebright, Jessica N.; Zavodni, Zachary J.; Yu, Ling; Wang, Tianyuan; Daiger, Stephen P.; Wistow, Graeme; Boon, Kathy; Hauser, Michael A.

2009-01-01

Purpose To develop large-scale, high-throughput annotation of the human macula transcriptome and to identify and prioritize candidate genes for inherited retinal dystrophies, based on ocular-expression profiles using serial analysis of gene expression (SAGE). Methods Two human retina and two retinal pigment epithelium (RPE)/choroid SAGE libraries made from matched macula or midperipheral retina and adjacent RPE/choroid of morphologically normal 28- to 66-year-old donors and a human central retina longSAGE library made from 41- to 66-year-old donors were generated. Their transcription profiles were entered into a relational database, EyeSAGE, including microarray expression profiles of retina and publicly available normal human tissue SAGE libraries. EyeSAGE was used to identify retina- and RPE-specific and -associated genes, and candidate genes for retina and RPE disease loci. Differential and/or cell-type specific expression was validated by quantitative and single-cell RT-PCR. Results Cone photoreceptor-associated gene expression was elevated in the macula transcription profiles. Analysis of the longSAGE retina tags enhanced tag-to-gene mapping and revealed alternatively spliced genes. Analysis of candidate gene expression tables for the identified Bardet-Biedl syndrome disease gene (BBS5) in the BBS5 disease region table yielded BBS5 as the top candidate. Compelling candidates for inherited retina diseases were identified. Conclusions The EyeSAGE database, combining three different gene-profiling platforms including the authors’ multidonor-derived retina/RPE SAGE libraries and existing single-donor retina/RPE libraries, is a powerful resource for definition of the retina and RPE transcriptomes. It can be used to identify retina-specific genes, including alternatively spliced transcripts and to prioritize candidate genes within mapped retinal disease regions. PMID:16723438

Defining the human macula transcriptome and candidate retinal disease genes using EyeSAGE.

PubMed

Bowes Rickman, Catherine; Ebright, Jessica N; Zavodni, Zachary J; Yu, Ling; Wang, Tianyuan; Daiger, Stephen P; Wistow, Graeme; Boon, Kathy; Hauser, Michael A

2006-06-01

To develop large-scale, high-throughput annotation of the human macula transcriptome and to identify and prioritize candidate genes for inherited retinal dystrophies, based on ocular-expression profiles using serial analysis of gene expression (SAGE). Two human retina and two retinal pigment epithelium (RPE)/choroid SAGE libraries made from matched macula or midperipheral retina and adjacent RPE/choroid of morphologically normal 28- to 66-year-old donors and a human central retina longSAGE library made from 41- to 66-year-old donors were generated. Their transcription profiles were entered into a relational database, EyeSAGE, including microarray expression profiles of retina and publicly available normal human tissue SAGE libraries. EyeSAGE was used to identify retina- and RPE-specific and -associated genes, and candidate genes for retina and RPE disease loci. Differential and/or cell-type specific expression was validated by quantitative and single-cell RT-PCR. Cone photoreceptor-associated gene expression was elevated in the macula transcription profiles. Analysis of the longSAGE retina tags enhanced tag-to-gene mapping and revealed alternatively spliced genes. Analysis of candidate gene expression tables for the identified Bardet-Biedl syndrome disease gene (BBS5) in the BBS5 disease region table yielded BBS5 as the top candidate. Compelling candidates for inherited retina diseases were identified. The EyeSAGE database, combining three different gene-profiling platforms including the authors' multidonor-derived retina/RPE SAGE libraries and existing single-donor retina/RPE libraries, is a powerful resource for definition of the retina and RPE transcriptomes. It can be used to identify retina-specific genes, including alternatively spliced transcripts and to prioritize candidate genes within mapped retinal disease regions.

RNA-seq Transcriptome Analysis of Panax japonicus, and Its Comparison with Other Panax Species to Identify Potential Genes Involved in the Saponins Biosynthesis

PubMed Central

Rai, Amit; Yamazaki, Mami; Takahashi, Hiroki; Nakamura, Michimi; Kojoma, Mareshige; Suzuki, Hideyuki; Saito, Kazuki

2016-01-01

The Panax genus has been a source of natural medicine, benefitting human health over the ages, among which the Panax japonicus represents an important species. Our understanding of several key pathways and enzymes involved in the biosynthesis of ginsenosides, a pharmacologically active class of metabolites and a major chemical constituents of the rhizome extracts from the Panax species, are limited. Limited genomic information, and lack of studies on comparative transcriptomics across the Panax species have restricted our understanding of the biosynthetic mechanisms of these and many other important classes of phytochemicals. Herein, we describe Illumina based RNA sequencing analysis to characterize the transcriptome and expression profiles of genes expressed in the five tissues of P. japonicus, and its comparison with other Panax species. RNA sequencing and de novo transcriptome assembly for P. japonicus resulted in a total of 135,235 unigenes with 78,794 (58.24%) unigenes being annotated using NCBI-nr database. Transcriptome profiling, and gene ontology enrichment analysis for five tissues of P. japonicus showed that although overall processes were evenly conserved across all tissues. However, each tissue was characterized by several unique unigenes with the leaves showing the most unique unigenes among the tissues studied. A comparative analysis of the P. japonicus transcriptome assembly with publically available transcripts from other Panax species, namely, P. ginseng, P. notoginseng, and P. quinquefolius also displayed high sequence similarity across all Panax species, with P. japonicus showing highest similarity with P. ginseng. Annotation of P. japonicus transcriptome resulted in the identification of putative genes encoding all enzymes from the triterpene backbone biosynthetic pathways, and identified 24 and 48 unigenes annotated as cytochrome P450 (CYP) and glycosyltransferases (GT), respectively. These CYPs and GTs annotated unigenes were conserved across all Panax species and co-expressed with other the transcripts involved in the triterpenoid backbone biosynthesis pathways. Unigenes identified in this study represent strong candidates for being involved in the triterpenoid saponins biosynthesis, and can serve as a basis for future validation studies. PMID:27148308

clusterProfiler: an R package for comparing biological themes among gene clusters.

PubMed

Yu, Guangchuang; Wang, Li-Gen; Han, Yanyan; He, Qing-Yu

2012-05-01

Increasing quantitative data generated from transcriptomics and proteomics require integrative strategies for analysis. Here, we present an R package, clusterProfiler that automates the process of biological-term classification and the enrichment analysis of gene clusters. The analysis module and visualization module were combined into a reusable workflow. Currently, clusterProfiler supports three species, including humans, mice, and yeast. Methods provided in this package can be easily extended to other species and ontologies. The clusterProfiler package is released under Artistic-2.0 License within Bioconductor project. The source code and vignette are freely available at http://bioconductor.org/packages/release/bioc/html/clusterProfiler.html.

Global transcriptomic profiling using small volumes of whole blood: a cost-effective method for translational genomic biomarker identification in small animals.

PubMed

Fricano, Meagan M; Ditewig, Amy C; Jung, Paul M; Liguori, Michael J; Blomme, Eric A G; Yang, Yi

2011-01-01

Blood is an ideal tissue for the identification of novel genomic biomarkers for toxicity or efficacy. However, using blood for transcriptomic profiling presents significant technical challenges due to the transcriptomic changes induced by ex vivo handling and the interference of highly abundant globin mRNA. Most whole blood RNA stabilization and isolation methods also require significant volumes of blood, limiting their effective use in small animal species, such as rodents. To overcome these challenges, a QIAzol-based RNA stabilization and isolation method (QSI) was developed to isolate sufficient amounts of high quality total RNA from 25 to 500 μL of rat whole blood. The method was compared to the standard PAXgene Blood RNA System using blood collected from rats exposed to saline or lipopolysaccharide (LPS). The QSI method yielded an average of 54 ng total RNA per μL of rat whole blood with an average RNA Integrity Number (RIN) of 9, a performance comparable with the standard PAXgene method. Total RNA samples were further processed using the NuGEN Ovation Whole Blood Solution system and cDNA was hybridized to Affymetrix Rat Genome 230 2.0 Arrays. The microarray QC parameters using RNA isolated with the QSI method were within the acceptable range for microarray analysis. The transcriptomic profiles were highly correlated with those using RNA isolated with the PAXgene method and were consistent with expected LPS-induced inflammatory responses. The present study demonstrated that the QSI method coupled with NuGEN Ovation Whole Blood Solution system is cost-effective and particularly suitable for transcriptomic profiling of minimal volumes of whole blood, typical of those obtained with small animal species.

Transcriptome analysis in non-model species: a new method for the analysis of heterologous hybridization on microarrays

PubMed Central

2010-01-01

Background Recent developments in high-throughput methods of analyzing transcriptomic profiles are promising for many areas of biology, including ecophysiology. However, although commercial microarrays are available for most common laboratory models, transcriptome analysis in non-traditional model species still remains a challenge. Indeed, the signal resulting from heterologous hybridization is low and difficult to interpret because of the weak complementarity between probe and target sequences, especially when no microarray dedicated to a genetically close species is available. Results We show here that transcriptome analysis in a species genetically distant from laboratory models is made possible by using MAXRS, a new method of analyzing heterologous hybridization on microarrays. This method takes advantage of the design of several commercial microarrays, with different probes targeting the same transcript. To illustrate and test this method, we analyzed the transcriptome of king penguin pectoralis muscle hybridized to Affymetrix chicken microarrays, two organisms separated by an evolutionary distance of approximately 100 million years. The differential gene expression observed between different physiological situations computed by MAXRS was confirmed by real-time PCR on 10 genes out of 11 tested. Conclusions MAXRS appears to be an appropriate method for gene expression analysis under heterologous hybridization conditions. PMID:20509979

Analysis of the Human Prostate-Specific Proteome Defined by Transcriptomics and Antibody-Based Profiling Identifies TMEM79 and ACOXL as Two Putative, Diagnostic Markers in Prostate Cancer

PubMed Central

O'Hurley, Gillian; Busch, Christer; Fagerberg, Linn; Hallström, Björn M.; Stadler, Charlotte; Tolf, Anna; Lundberg, Emma; Schwenk, Jochen M.; Jirström, Karin; Bjartell, Anders; Gallagher, William M.; Uhlén, Mathias; Pontén, Fredrik

2015-01-01

To better understand prostate function and disease, it is important to define and explore the molecular constituents that signify the prostate gland. The aim of this study was to define the prostate specific transcriptome and proteome, in comparison to 26 other human tissues. Deep sequencing of mRNA (RNA-seq) and immunohistochemistry-based protein profiling were combined to identify prostate specific gene expression patterns and to explore tissue biomarkers for potential clinical use in prostate cancer diagnostics. We identified 203 genes with elevated expression in the prostate, 22 of which showed more than five-fold higher expression levels compared to all other tissue types. In addition to previously well-known proteins we identified two poorly characterized proteins, TMEM79 and ACOXL, with potential to differentiate between benign and cancerous prostatic glands in tissue biopsies. In conclusion, we have applied a genome-wide analysis to identify the prostate specific proteome using transcriptomics and antibody-based protein profiling to identify genes with elevated expression in the prostate. Our data provides a starting point for further functional studies to explore the molecular repertoire of normal and diseased prostate including potential prostate cancer markers such as TMEM79 and ACOXL. PMID:26237329

Single-cell analysis of the transcriptome and its application in the characterization of stem cells and early embryos.

PubMed

Liu, Na; Liu, Lin; Pan, Xinghua

2014-07-01

Cellular heterogeneity within a cell population is a common phenomenon in multicellular organisms, tissues, cultured cells, and even FACS-sorted subpopulations. Important information may be masked if the cells are studied as a mass. Transcriptome profiling is a parameter that has been intensively studied, and relatively easier to address than protein composition. To understand the basis and importance of heterogeneity and stochastic aspects of the cell function and its mechanisms, it is essential to examine transcriptomes of a panel of single cells. High-throughput technologies, starting from microarrays and now RNA-seq, provide a full view of the expression of transcriptomes but are limited by the amount of RNA for analysis. Recently, several new approaches for amplification and sequencing the transcriptome of single cells or a limited low number of cells have been developed and applied. In this review, we summarize these major strategies, such as PCR-based methods, IVT-based methods, phi29-DNA polymerase-based methods, and several other methods, including their principles, characteristics, advantages, and limitations, with representative applications in cancer stem cells, early development, and embryonic stem cells. The prospects for development of future technology and application of transcriptome analysis in a single cell are also discussed.

Gene expression profiles in rainbow trout, Onchorynchus mykiss, exposed to a simple chemical mixture.

PubMed

Hook, Sharon E; Skillman, Ann D; Gopalan, Banu; Small, Jack A; Schultz, Irvin R

2008-03-01

Among proposed uses for microarrays in environmental toxiciology is the identification of key contributors to toxicity within a mixture. However, it remains uncertain whether the transcriptomic profiles resulting from exposure to a mixture have patterns of altered gene expression that contain identifiable contributions from each toxicant component. We exposed isogenic rainbow trout Onchorynchus mykiss, to sublethal levels of ethynylestradiol, 2,2,4,4-tetrabromodiphenyl ether, and chromium VI or to a mixture of all three toxicants Fluorescently labeled complementary DNA (cDNA) were generated and hybridized against a commercially available Salmonid array spotted with 16,000 cDNAs. Data were analyzed using analysis of variance (p<0.05) with a Benjamani-Hochberg multiple test correction (Genespring [Agilent] software package) to identify up and downregulated genes. Gene clustering patterns that can be used as "expression signatures" were determined using hierarchical cluster analysis. The gene ontology terms associated with significantly altered genes were also used to identify functional groups that were associated with toxicant exposure. Cross-ontological analytics approach was used to assign functional annotations to genes with "unknown" function. Our analysis indicates that transcriptomic profiles resulting from the mixture exposure resemble those of the individual contaminant exposures, but are not a simple additive list. However, patterns of altered genes representative of each component of the mixture are clearly discernible, and the functional classes of genes altered represent the individual components of the mixture. These findings indicate that the use of microarrays to identify transcriptomic profiles may aid in the identification of key stressors within a chemical mixture, ultimately improving environmental assessment.

Stem cells isolated from adipose tissue of obese patients show changes in their transcriptomic profile that indicate loss in stemcellness and increased commitment to an adipocyte-like phenotype

PubMed Central

2013-01-01

Background The adipose tissue is an endocrine regulator and a risk factor for atherosclerosis and cardiovascular disease when by excessive accumulation induces obesity. Although the adipose tissue is also a reservoir for stem cells (ASC) their function and “stemcellness” has been questioned. Our aim was to investigate the mechanisms by which obesity affects subcutaneous white adipose tissue (WAT) stem cells. Results Transcriptomics, in silico analysis, real-time polymerase chain reaction (PCR) and western blots were performed on isolated stem cells from subcutaneous abdominal WAT of morbidly obese patients (ASCmo) and of non-obese individuals (ASCn). ASCmo and ASCn gene expression clustered separately from each other. ASCmo showed downregulation of “stemness” genes and upregulation of adipogenic and inflammatory genes with respect to ASCn. Moreover, the application of bioinformatics and Ingenuity Pathway Analysis (IPA) showed that the transcription factor Smad3 was tentatively affected in obese ASCmo. Validation of this target confirmed a significantly reduced Smad3 nuclear translocation in the isolated ASCmo. Conclusions The transcriptomic profile of the stem cells reservoir in obese subcutaneous WAT is highly modified with significant changes in genes regulating stemcellness, lineage commitment and inflammation. In addition to body mass index, cardiovascular risk factor clustering further affect the ASC transcriptomic profile inducing loss of multipotency and, hence, capacity for tissue repair. In summary, the stem cells in the subcutaneous WAT niche of obese patients are already committed to adipocyte differentiation and show an upregulated inflammatory gene expression associated to their loss of stemcellness. PMID:24040759

Mining genes involved in insecticide resistance of Liposcelis bostrychophila Badonnel by transcriptome and expression profile analysis.

PubMed

Dou, Wei; Shen, Guang-Mao; Niu, Jin-Zhi; Ding, Tian-Bo; Wei, Dan-Dan; Wang, Jin-Jun

2013-01-01

Recent studies indicate that infestations of psocids pose a new risk for global food security. Among the psocids species, Liposcelis bostrychophila Badonnel has gained recognition in importance because of its parthenogenic reproduction, rapid adaptation, and increased worldwide distribution. To date, the molecular data available for L. bostrychophila is largely limited to genes identified through homology. Also, no transcriptome data relevant to psocids infection is available. In this study, we generated de novo assembly of L. bostrychophila transcriptome performed through the short read sequencing technology (Illumina). In a single run, we obtained more than 51 million sequencing reads that were assembled into 60,012 unigenes (mean size = 711 bp) by Trinity. The transcriptome sequences from different developmental stages of L. bostrychophila including egg, nymph and adult were annotated with non-redundant (Nr) protein database, gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG orthology (KO). The analysis revealed three major enzyme families involved in insecticide metabolism as differentially expressed in the L. bostrychophila transcriptome. A total of 49 P450-, 31 GST- and 21 CES-specific genes representing the three enzyme families were identified. Besides, 16 transcripts were identified to contain target site sequences of resistance genes. Furthermore, we profiled gene expression patterns upon insecticide (malathion and deltamethrin) exposure using the tag-based digital gene expression (DGE) method. The L. bostrychophila transcriptome and DGE data provide gene expression data that would further our understanding of molecular mechanisms in psocids. In particular, the findings of this investigation will facilitate identification of genes involved in insecticide resistance and designing of new compounds for control of psocids.

Mining Genes Involved in Insecticide Resistance of Liposcelis bostrychophila Badonnel by Transcriptome and Expression Profile Analysis

PubMed Central

Dou, Wei; Shen, Guang-Mao; Niu, Jin-Zhi; Ding, Tian-Bo; Wei, Dan-Dan; Wang, Jin-Jun

2013-01-01

Background Recent studies indicate that infestations of psocids pose a new risk for global food security. Among the psocids species, Liposcelis bostrychophila Badonnel has gained recognition in importance because of its parthenogenic reproduction, rapid adaptation, and increased worldwide distribution. To date, the molecular data available for L. bostrychophila is largely limited to genes identified through homology. Also, no transcriptome data relevant to psocids infection is available. Methodology and Principal Findings In this study, we generated de novo assembly of L. bostrychophila transcriptome performed through the short read sequencing technology (Illumina). In a single run, we obtained more than 51 million sequencing reads that were assembled into 60,012 unigenes (mean size = 711 bp) by Trinity. The transcriptome sequences from different developmental stages of L. bostrychophila including egg, nymph and adult were annotated with non-redundant (Nr) protein database, gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG orthology (KO). The analysis revealed three major enzyme families involved in insecticide metabolism as differentially expressed in the L. bostrychophila transcriptome. A total of 49 P450-, 31 GST- and 21 CES-specific genes representing the three enzyme families were identified. Besides, 16 transcripts were identified to contain target site sequences of resistance genes. Furthermore, we profiled gene expression patterns upon insecticide (malathion and deltamethrin) exposure using the tag-based digital gene expression (DGE) method. Conclusion The L. bostrychophila transcriptome and DGE data provide gene expression data that would further our understanding of molecular mechanisms in psocids. In particular, the findings of this investigation will facilitate identification of genes involved in insecticide resistance and designing of new compounds for control of psocids. PMID:24278202

Dynamic transcriptomic analysis in hircine longissimus dorsi muscle from fetal to neonatal development stages.

PubMed

Zhan, Siyuan; Zhao, Wei; Song, Tianzeng; Dong, Yao; Guo, Jiazhong; Cao, Jiaxue; Zhong, Tao; Wang, Linjie; Li, Li; Zhang, Hongping

2018-01-01

Muscle growth and development from fetal to neonatal stages consist of a series of delicately regulated and orchestrated changes in expression of genes. In this study, we performed whole transcriptome profiling based on RNA-Seq of caprine longissimus dorsi muscle tissue obtained from prenatal stages (days 45, 60, and 105 of gestation) and neonatal stage (the 3-day-old newborn) to identify genes that are differentially expressed and investigate their temporal expression profiles. A total of 3276 differentially expressed genes (DEGs) were identified (Q value < 0.01). Time-series expression profile clustering analysis indicated that DEGs were significantly clustered into eight clusters which can be divided into two classes (Q value < 0.05), class I profiles with downregulated patterns and class II profiles with upregulated patterns. Based on cluster analysis, GO enrichment analysis found that 75, 25, and 8 terms to be significantly enriched in biological process (BP), cellular component (CC), and molecular function (MF) categories in class I profiles, while 35, 21, and 8 terms to be significantly enriched in BP, CC, and MF in class II profiles. KEGG pathway analysis revealed that DEGs from class I profiles were significantly enriched in 22 pathways and the most enriched pathway was Rap1 signaling pathway. DEGs from class II profiles were significantly enriched in 17 pathways and the mainly enriched pathway was AMPK signaling pathway. Finally, six selected DEGs from our sequencing results were confirmed by qPCR. Our study provides a comprehensive understanding of the molecular mechanisms during goat skeletal muscle development from fetal to neonatal stages and valuable information for future studies of muscle development in goats.

Comparative transcriptome profiling of chilling stress responsiveness in grafted watermelon seedlings.

PubMed

Xu, Jinhua; Zhang, Man; Liu, Guang; Yang, Xingping; Hou, Xilin

2016-12-01

Rootstock grafting may improve the resistance of watermelon plants to low temperatures. However, information regarding the molecular responses of rootstock grafted plants to chilling stress is limited. To elucidate the molecular mechanisms of chilling tolerance in grafted plants, the transcriptomic responses of grafted watermelon under chilling stress were analyzed using RNA-seq analysis. Sequencing data were used for digital gene expression (DGE) analysis to characterize the transcriptomic responses in grafted watermelon seedlings. A total of 702 differentially-expressed genes (DEGs) were found in rootstock grafted (RG) watermelon relative to self-grafted (SG) watermelon; among these genes, 522 genes were up-regulated and 180 were down-regulated. Additionally, 164 and 953 genes were found to specifically expressed in RG and SG seedlings under chilling stress, respectively. Functional annotations revealed that up-regulated DEGs are involved in protein processing, plant-pathogen interaction and the spliceosome, whereas down-regulated DEGs are associated with photosynthesis. Moreover, 13 DEGs were randomly selected for quantitative real time PCR (qRT-PCR) analysis. The expression profiles of these 13 DEGs were consistent with those detected by the DGE analysis, supporting the reliability of the DGE data. This work provides additional insight into the molecular basis of grafted watermelon responses to chilling stress. Copyright © 2016. Published by Elsevier Masson SAS.

Clinical Correlations of Transcriptional Profile in Patients Infected with Avian Influenza H7N9 Virus.

PubMed

Guan, Wenda; Wu, Nicholas C; Lee, Horace H Y; Li, Yimin; Jiang, Wenxin; Shen, Lihan; Wu, Douglas C; Chen, Rongchang; Zhong, Nanshan; Wilson, Ian A; Peiris, Malik; Yang, Zifeng; Mok, Chris K P

2018-05-28

Avian influenza A (H7N9) viruses emerged in China in 2013 and caused zoonotic disease associated with a case-fatality ratio of over 30%. Transcriptional profiles in peripheral blood reflect host responses and can help to elucidate disease pathogenesis. We correlated serial blood transcriptomic profiles of patients with avian influenza A (H7N9) virus infection and determined the biological significances from the analysis. We found that specific gene expression profiles in the blood were strongly correlated with the PaO2/FiO2 ratio and viral load in the lower respiratory tract (LRT). Cell cycle and leukocyte-related immunity were activated at the acute stage of the infection while T cell functions and various metabolic processes were associated with the recovery phase of the illness. A transition from systemic innate to adaptive immunity was found. We developed a novel approach for transcriptomic analysis to identify key host responses that were strongly correlated with specific clinical and virologic parameters in patients with H7N9 infection.

Transcriptomic responses to wounding: meta-analysis of gene expression microarray data.

PubMed

Sass, Piotr Andrzej; Dąbrowski, Michał; Charzyńska, Agata; Sachadyn, Paweł

2017-11-07

A vast amount of microarray data on transcriptomic response to injury has been collected so far. We designed the analysis in order to identify the genes displaying significant changes in expression after wounding in different organisms and tissues. This meta-analysis is the first study to compare gene expression profiles in response to wounding in as different tissues as heart, liver, skin, bones, and spinal cord, and species, including rat, mouse and human. We collected available microarray transcriptomic profiles obtained from different tissue injury experiments and selected the genes showing a minimum twofold change in expression in response to wounding in prevailing number of experiments for each of five wound healing stages we distinguished: haemostasis & early inflammation, inflammation, early repair, late repair and remodelling. During the initial phases after wounding, haemostasis & early inflammation and inflammation, the transcriptomic responses showed little consistency between different tissues and experiments. For the later phases, wound repair and remodelling, we identified a number of genes displaying similar transcriptional responses in all examined tissues. As revealed by ontological analyses, activation of certain pathways was rather specific for selected phases of wound healing, such as e.g. responses to vitamin D pronounced during inflammation. Conversely, we observed induction of genes encoding inflammatory agents and extracellular matrix proteins in all wound healing phases. Further, we selected several genes differentially upregulated throughout different stages of wound response, including established factors of wound healing in addition to those previously unreported  in this context such as PTPRC and AQP4. We found that transcriptomic responses to wounding showed similar traits in a diverse selection of tissues including skin, muscles, internal organs and nervous system. Notably, we distinguished transcriptional induction of inflammatory genes not only in the initial response to wounding, but also later, during wound repair and tissue remodelling.

Transcriptomic immune response of Tenebrio molitor pupae to parasitization by Scleroderma guani.

PubMed

Zhu, Jia-Ying; Yang, Pu; Zhang, Zhong; Wu, Guo-Xing; Yang, Bin

2013-01-01

Host and parasitoid interaction is one of the most fascinating relationships of insects, which is currently receiving an increasing interest. Understanding the mechanisms evolved by the parasitoids to evade or suppress the host immune system is important for dissecting this interaction, while it was still poorly known. In order to gain insight into the immune response of Tenebrio molitor to parasitization by Scleroderma guani, the transcriptome of T. molitor pupae was sequenced with focus on immune-related gene, and the non-parasitized and parasitized T. molitor pupae were analyzed by digital gene expression (DGE) analysis with special emphasis on parasitoid-induced immune-related genes using Illumina sequencing. In a single run, 264,698 raw reads were obtained. De novo assembly generated 71,514 unigenes with mean length of 424 bp. Of those unigenes, 37,373 (52.26%) showed similarity to the known proteins in the NCBI nr database. Via analysis of the transcriptome data in depth, 430 unigenes related to immunity were identified. DGE analysis revealed that parasitization by S. guani had considerable impacts on the transcriptome profile of T. molitor pupae, as indicated by the significant up- or down-regulation of 3,431 parasitism-responsive transcripts. The expression of a total of 74 unigenes involved in immune response of T. molitor was significantly altered after parasitization. obtained T. molitor transcriptome, in addition to establishing a fundamental resource for further research on functional genomics, has allowed the discovery of a large group of immune genes that might provide a meaningful framework to better understand the immune response in this species and other beetles. The DGE profiling data provides comprehensive T. molitor immune gene expression information at the transcriptional level following parasitization, and sheds valuable light on the molecular understanding of the host-parasitoid interaction.

Arabidopsis Gene Family Profiler (aGFP)--user-oriented transcriptomic database with easy-to-use graphic interface.

PubMed

Dupl'áková, Nikoleta; Renák, David; Hovanec, Patrik; Honysová, Barbora; Twell, David; Honys, David

2007-07-23

Microarray technologies now belong to the standard functional genomics toolbox and have undergone massive development leading to increased genome coverage, accuracy and reliability. The number of experiments exploiting microarray technology has markedly increased in recent years. In parallel with the rapid accumulation of transcriptomic data, on-line analysis tools are being introduced to simplify their use. Global statistical data analysis methods contribute to the development of overall concepts about gene expression patterns and to query and compose working hypotheses. More recently, these applications are being supplemented with more specialized products offering visualization and specific data mining tools. We present a curated gene family-oriented gene expression database, Arabidopsis Gene Family Profiler (aGFP; http://agfp.ueb.cas.cz), which gives the user access to a large collection of normalised Affymetrix ATH1 microarray datasets. The database currently contains NASC Array and AtGenExpress transcriptomic datasets for various tissues at different developmental stages of wild type plants gathered from nearly 350 gene chips. The Arabidopsis GFP database has been designed as an easy-to-use tool for users needing an easily accessible resource for expression data of single genes, pre-defined gene families or custom gene sets, with the further possibility of keyword search. Arabidopsis Gene Family Profiler presents a user-friendly web interface using both graphic and text output. Data are stored at the MySQL server and individual queries are created in PHP script. The most distinguishable features of Arabidopsis Gene Family Profiler database are: 1) the presentation of normalized datasets (Affymetrix MAS algorithm and calculation of model-based gene-expression values based on the Perfect Match-only model); 2) the choice between two different normalization algorithms (Affymetrix MAS4 or MAS5 algorithms); 3) an intuitive interface; 4) an interactive "virtual plant" visualizing the spatial and developmental expression profiles of both gene families and individual genes. Arabidopsis GFP gives users the possibility to analyze current Arabidopsis developmental transcriptomic data starting with simple global queries that can be expanded and further refined to visualize comparative and highly selective gene expression profiles.

Transcriptome analysis of genes involved in defense against alkaline stress in roots of wild jujube (Ziziphus acidojujuba)

PubMed Central

Tian, Shan; Wang, Bei; Zhao, Xusheng

2017-01-01

Wild jujube (Ziziphus acidojujuba Mill.) is highly tolerant to alkaline, saline and drought stress; however, no studies have performed transcriptome profiling to study the response of wild jujube to these and other abiotic stresses. In this study, we examined the tolerance of wild jujube to NaHCO3-NaOH solution and analyzed gene expression profiles in response to alkaline stress. Physiological experiments revealed that H2O2 content in leaves increased significantly and root activity decreased quickly during alkaline of pH 9.5 treatment. For transcriptome analysis, wild jujube plants grown hydroponically were treated with NaHCO3-NaOH solution for 0, 1, and 12 h and six transcriptomes from roots were built. In total, 32,758 genes were generated, and 3,604 differentially expressed genes (DEGs) were identified. After 1 h, 853 genes showed significantly different expression between control and treated plants; after 12 h, expression of 2,856 genes was significantly different. The expression pattern of nine genes was validated by quantitative real-time PCR. After gene annotation and gene ontology enrichment analysis, the genes encoding transcriptional factors, serine/threonine-protein kinases, heat shock proteins, cysteine-like kinases, calmodulin-like proteins, and reactive oxygen species (ROS) scavengers were found to be closely involved in alkaline stress response. These results will provide useful insights for elucidating the mechanisms underlying alkaline tolerance in wild jujube. PMID:28976994

Staying alive in adversity: transcriptome dynamics in the stress-resistant dauer larva.

PubMed

Holt, Suzan J

2006-10-01

In response to food depletion and overcrowding, the soil nematode Caenorhabditis elegans can arrest development and form an alternate third larval stage called the dauer. Though nonfeeding, the dauer larva is long lived and stress resistant. Metabolic and transcription rates are lowered but the transcriptome of the dauer is complex. In this study, distribution analysis of transcript profiles generated by Serial Analysis of Gene Expression (SAGE) in dauer larvae and in mixed developmental stages is presented. An inverse relationship was observed between frequency and abundance/copy number of SAGE tag types (transcripts) in both profiles. In the dauer profile, a relatively greater proportion of highly abundant transcripts was counterbalanced by a smaller fraction of low to moderately abundant transcripts. Comparisons of abundant tag counts between the two profiles revealed relative enrichment in the dauer profile of transcripts with predicted or known involvement in ribosome biogenesis and protein synthesis, membrane transport, and immune responses. Translation-coupled mRNA decay is proposed as part of an immune-like stress response in the dauer larva. An influence of genomic region on transcript level may reflect the coordination of transcription and mRNA turnover.

Strand-specific transcriptome profiling with directly labeled RNA on genomic tiling microarrays

PubMed Central

2011-01-01

Background With lower manufacturing cost, high spot density, and flexible probe design, genomic tiling microarrays are ideal for comprehensive transcriptome studies. Typically, transcriptome profiling using microarrays involves reverse transcription, which converts RNA to cDNA. The cDNA is then labeled and hybridized to the probes on the arrays, thus the RNA signals are detected indirectly. Reverse transcription is known to generate artifactual cDNA, in particular the synthesis of second-strand cDNA, leading to false discovery of antisense RNA. To address this issue, we have developed an effective method using RNA that is directly labeled, thus by-passing the cDNA generation. This paper describes this method and its application to the mapping of transcriptome profiles. Results RNA extracted from laboratory cultures of Porphyromonas gingivalis was fluorescently labeled with an alkylation reagent and hybridized directly to probes on genomic tiling microarrays specifically designed for this periodontal pathogen. The generated transcriptome profile was strand-specific and produced signals close to background level in most antisense regions of the genome. In contrast, high levels of signal were detected in the antisense regions when the hybridization was done with cDNA. Five antisense areas were tested with independent strand-specific RT-PCR and none to negligible amplification was detected, indicating that the strong antisense cDNA signals were experimental artifacts. Conclusions An efficient method was developed for mapping transcriptome profiles specific to both coding strands of a bacterial genome. This method chemically labels and uses extracted RNA directly in microarray hybridization. The generated transcriptome profile was free of cDNA artifactual signals. In addition, this method requires fewer processing steps and is potentially more sensitive in detecting small amount of RNA compared to conventional end-labeling methods due to the incorporation of more fluorescent molecules per RNA fragment. PMID:21235785

Transcriptomic Studies of the Effect of nod Gene-Inducing Molecules in Rhizobia: Different Weapons, One Purpose

PubMed Central

Jiménez-Guerrero, Irene; Acosta-Jurado, Sebastián; Navarro-Gómez, Pilar; López-Baena, Francisco Javier; Ollero, Francisco Javier

2017-01-01

Simultaneous quantification of transcripts of the whole bacterial genome allows the analysis of the global transcriptional response under changing conditions. RNA-seq and microarrays are the most used techniques to measure these transcriptomic changes, and both complement each other in transcriptome profiling. In this review, we exhaustively compiled the symbiosis-related transcriptomic reports (microarrays and RNA sequencing) carried out hitherto in rhizobia. This review is specially focused on transcriptomic changes that takes place when five rhizobial species, Bradyrhizobium japonicum (=diazoefficiens) USDA 110, Rhizobium leguminosarum biovar viciae 3841, Rhizobium tropici CIAT 899, Sinorhizobium (=Ensifer) meliloti 1021 and S. fredii HH103, recognize inducing flavonoids, plant-exuded phenolic compounds that activate the biosynthesis and export of Nod factors (NF) in all analysed rhizobia. Interestingly, our global transcriptomic comparison also indicates that each rhizobial species possesses its own arsenal of molecular weapons accompanying the set of NF in order to establish a successful interaction with host legumes. PMID:29267254

Venom gland transcriptomics for identifying, cataloging, and characterizing venom proteins in snakes.

PubMed

Brahma, Rajeev Kungur; McCleary, Ryan J R; Kini, R Manjunatha; Doley, Robin

2015-01-01

Snake venoms are cocktails of protein toxins that play important roles in capture and digestion of prey. Significant qualitative and quantitative variation in snake venom composition has been observed among and within species. Understanding these variations in protein components is instrumental in interpreting clinical symptoms during human envenomation and in searching for novel venom proteins with potential therapeutic applications. In the last decade, transcriptomic analyses of venom glands have helped in understanding the composition of various snake venoms in great detail. Here we review transcriptomic analysis as a powerful tool for understanding venom profile, variation and evolution. Copyright © 2014 Elsevier Ltd. All rights reserved.

Use of homologous and heterologous gene expression profiling tools to characterize transcription dynamics during apple fruit maturation and ripening

PubMed Central

2010-01-01

Background Fruit development, maturation and ripening consists of a complex series of biochemical and physiological changes that in climacteric fruits, including apple and tomato, are coordinated by the gaseous hormone ethylene. These changes lead to final fruit quality and understanding of the functional machinery underlying these processes is of both biological and practical importance. To date many reports have been made on the analysis of gene expression in apple. In this study we focused our investigation on the role of ethylene during apple maturation, specifically comparing transcriptomics of normal ripening with changes resulting from application of the hormone receptor competitor 1-Methylcyclopropene. Results To gain insight into the molecular process regulating ripening in apple, and to compare to tomato (model species for ripening studies), we utilized both homologous and heterologous (tomato) microarray to profile transcriptome dynamics of genes involved in fruit development and ripening, emphasizing those which are ethylene regulated. The use of both types of microarrays facilitated transcriptome comparison between apple and tomato (for the later using data previously published and available at the TED: tomato expression database) and highlighted genes conserved during ripening of both species, which in turn represent a foundation for further comparative genomic studies. The cross-species analysis had the secondary aim of examining the efficiency of heterologous (specifically tomato) microarray hybridization for candidate gene identification as related to the ripening process. The resulting transcriptomics data revealed coordinated gene expression during fruit ripening of a subset of ripening-related and ethylene responsive genes, further facilitating the analysis of ethylene response during fruit maturation and ripening. Conclusion Our combined strategy based on microarray hybridization enabled transcriptome characterization during normal climacteric apple ripening, as well as definition of ethylene-dependent transcriptome changes. Comparison with tomato fruit maturation and ethylene responsive transcriptome activity facilitated identification of putative conserved orthologous ripening-related genes, which serve as an initial set of candidates for assessing conservation of gene activity across genomes of fruit bearing plant species. PMID:20973957

Application of D-Crustacean Hyperglycemic Hormone Induces Peptidases Transcription and Suppresses Glycolysis-Related Transcripts in the Hepatopancreas of the Crayfish Pontastacus leptodactylus — Results of a Transcriptomic Study

PubMed Central

De Moro, Gianluca; Gerdol, Marco; Guarnaccia, Corrado; Mosco, Alessandro; Pallavicini, Alberto; Giulianini, Piero Giulio

2013-01-01

The crustacean Hyperglycemic Hormone (cHH) is a neuropeptide present in many decapods. Two different chiral isomers are simultaneously present in Astacid crayfish and their specific biological functions are still poorly understood. The present study is aimed at better understanding the potentially different effect of each of the isomers on the hepatopancreatic gene expression profile in the crayfish Pontastacus leptodactylus, in the context of short term hyperglycemia. Hence, two different chemically synthesized cHH enantiomers, containing either L- or D-Phe3, were injected to the circulation of intermolt females following removal of their X organ-Sinus gland complex. The effects triggered by the injection of the two alternate isomers were detected after one hour through measurement of circulating glucose levels. Triggered changes of the transcriptome expression profile in the hepatopancreas were analyzed by RNA-seq. A whole transcriptome shotgun sequence assembly provided the assumedly complete transcriptome of P. leptodactylus hepatopancreas, followed by RNA-seq analysis of changes in the expression level of many genes caused by the application of each of the hormone isomers. Circulating glucose levels were much higher in response to the D-isoform than to the L-isoform injection, one hour from injection. Similarly, the RNA-seq analysis confirmed a stronger effect on gene expression following the administration of D-cHH, while just limited alterations were caused by the L-isomer. These findings demonstrated a more prominent short term effect of the D-cHH on the transcription profile and shed light on the effect of the D-isomer on specific functional gene groups. Another contribution of the study is the construction of a de novo assembly of the hepatopancreas transcriptome, consisting of 39,935 contigs, that dramatically increases the molecular information available for this species and for crustaceans in general, providing an efficient tool for studying gene expression patterns in this organ. PMID:23840318

Human and feline adipose-derived mesenchymal stem cells have comparable phenotype, immunomodulatory functions, and transcriptome.

PubMed

Clark, Kaitlin C; Fierro, Fernando A; Ko, Emily Mills; Walker, Naomi J; Arzi, Boaz; Tepper, Clifford G; Dahlenburg, Heather; Cicchetto, Andrew; Kol, Amir; Marsh, Lyndsey; Murphy, William J; Fazel, Nasim; Borjesson, Dori L

2017-03-20

Adipose-derived mesenchymal stem cells (ASCs) are a promising cell therapy to treat inflammatory and immune-mediated diseases. Development of appropriate pre-clinical animal models is critical to determine safety and attain early efficacy data for the most promising therapeutic candidates. Naturally occurring diseases in cats already serve as valuable models to inform human clinical trials in oncologic, cardiovascular, and genetic diseases. The objective of this study was to complete a comprehensive side-by-side comparison of human and feline ASCs, with an emphasis on their immunomodulatory capacity and transcriptome. Human and feline ASCs were evaluated for phenotype, immunomodulatory profile, and transcriptome. Additionally, transwells were used to determine the role of cell-cell contact in ASC-mediated inhibition of lymphocyte proliferation in both humans and cats. Similar to human ASCs, feline ASCs were highly proliferative at low passages and fit the minimal criteria of multipotent stem cells including a compatible surface protein phenotype, osteogenic capacity, and normal karyotype. Like ASCs from all species, feline ASCs inhibited mitogen-activated lymphocyte proliferation in vitro, with or without direct ASC-lymphocyte contact. Feline ASCs mimic human ASCs in their mediator secretion pattern, including prostaglandin E2, indoleamine 2,3 dioxygenase, transforming growth factor beta, and interleukin-6, all augmented by interferon gamma secretion by lymphocytes. The transcriptome of three unactivated feline ASC lines were highly similar. Functional analysis of the most highly expressed genes highlighted processes including: 1) the regulation of apoptosis; 2) cell adhesion; 3) response to oxidative stress; and 4) regulation of cell differentiation. Finally, feline ASCs had a similar gene expression profile to noninduced human ASCs. Findings suggest that feline ASCs modulate lymphocyte proliferation using soluble mediators that mirror the human ASC secretion pattern. Uninduced feline ASCs have similar gene expression profiles to uninduced human ASCs, as revealed by transcriptome analysis. These data will help inform clinical trials using cats with naturally occurring diseases as surrogate models for human clinical trials in the regenerative medicine arena.

Differential transcriptome analysis reveals genes related to cold tolerance in seabuckthorn carpenter moth, Eogystia hippophaecolus

PubMed Central

Hu, Ping; Wang, Tao; Tao, Jing; Zong, Shixiang

2017-01-01

Seabuckthorn carpenter moth, Eogystia hippophaecolus (Lepidoptera: Cossidae), is an important pest of sea buckthorn (Hippophae rhamnoides), which is a shrub that has significant ecological and economic value in China. E. hippophaecolus is highly cold tolerant, but limited studies have been conducted to elucidate the molecular mechanisms underlying its cold resistance. Here we sequenced the E. hippophaecolus transcriptome using RNA-Seq technology and performed de novo assembly from the short paired-end reads. We investigated the larval response to cold stress by comparing gene expression profiles between treatments. We obtained 118,034 unigenes, of which 22,161 were annotated with gene descriptions, conserved domains, gene ontology terms, and metabolic pathways. These resulted in 57 GO terms and 193 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. By comparing transcriptome profiles for differential gene expression, we identified many differentially expressed proteins and genes, including heat shock proteins and cuticular proteins which have previously been reported to be involved in cold resistance of insects. This study provides a global transcriptome analysis and an assessment of differential gene expression in E. hippophaecolus under cold stress. We found seven differential expressed genes in common between developmental stages, which were verified with qPCR. Our findings facilitate future genomic studies aimed at improving our understanding of the molecular mechanisms underlying the response of insects to low temperatures. PMID:29131867

Differential transcriptome analysis reveals genes related to cold tolerance in seabuckthorn carpenter moth, Eogystia hippophaecolus.

PubMed

Cui, Mingming; Hu, Ping; Wang, Tao; Tao, Jing; Zong, Shixiang

2017-01-01

Seabuckthorn carpenter moth, Eogystia hippophaecolus (Lepidoptera: Cossidae), is an important pest of sea buckthorn (Hippophae rhamnoides), which is a shrub that has significant ecological and economic value in China. E. hippophaecolus is highly cold tolerant, but limited studies have been conducted to elucidate the molecular mechanisms underlying its cold resistance. Here we sequenced the E. hippophaecolus transcriptome using RNA-Seq technology and performed de novo assembly from the short paired-end reads. We investigated the larval response to cold stress by comparing gene expression profiles between treatments. We obtained 118,034 unigenes, of which 22,161 were annotated with gene descriptions, conserved domains, gene ontology terms, and metabolic pathways. These resulted in 57 GO terms and 193 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. By comparing transcriptome profiles for differential gene expression, we identified many differentially expressed proteins and genes, including heat shock proteins and cuticular proteins which have previously been reported to be involved in cold resistance of insects. This study provides a global transcriptome analysis and an assessment of differential gene expression in E. hippophaecolus under cold stress. We found seven differential expressed genes in common between developmental stages, which were verified with qPCR. Our findings facilitate future genomic studies aimed at improving our understanding of the molecular mechanisms underlying the response of insects to low temperatures.

Transcriptomic and metabolomic profiles of Chinese citrus fly, Bactrocera minax (Diptera: Tephritidae), along with pupal development provide insight into diapause program

PubMed Central

Fan, Huan; Xiong, Ke-Cai; Liu, Ying-Hong

2017-01-01

The Chinese citrus fly, Bactrocera minax (Enderlein), is a devastating citrus pest in Asia. This univoltine insect enters obligatory pupal diapause in each generation, while little is known about the course and the molecular mechanisms of diapause. In this study, the course of diapause was determined by measuring the respiratory rate throughout the pupal stage. In addition, the variation of transcriptomic and metabolomic profiles of pupae at five developmental stages (pre-, early-, middle-, late-, and post-diapause) were evaluated by next-generation sequencing technology and 1H nuclear magnetic resonance spectroscopy (NMR), respectively. A total of 4,808 genes were significantly altered in ten pairwise comparisons, representing major shifts in metabolism and signal transduction as well as endocrine system and digestive system. Gene expression profiles were validated by qRT-PCR analysis. In addition, 48 metabolites were identified and quantified by 1H NMR. Nine of which significantly contributed to the variation in the metabolomic profiles, especially proline and trehalose. Moreover, the samples collected within diapause maintenance (early-, middle-, and late-diapause) only exhibited marginal transcriptomic and metabolomic variation with each other. These findings greatly improve our understanding of B. minax diapause and lay the foundation for further pertinent studies. PMID:28704500

Bioorthogonal Metabolic Labeling of Nascent RNA in Neurons Improves the Sensitivity of Transcriptome-Wide Profiling.

PubMed

Zajaczkowski, Esmi L; Zhao, Qiong-Yi; Zhang, Zong Hong; Li, Xiang; Wei, Wei; Marshall, Paul R; Leighton, Laura J; Nainar, Sarah; Feng, Chao; Spitale, Robert C; Bredy, Timothy W

2018-06-15

Transcriptome-wide expression profiling of neurons has provided important insights into the underlying molecular mechanisms and gene expression patterns that transpire during learning and memory formation. However, there is a paucity of tools for profiling stimulus-induced RNA within specific neuronal cell populations. A bioorthogonal method to chemically label nascent (i.e., newly transcribed) RNA in a cell-type-specific and temporally controlled manner, which is also amenable to bioconjugation via click chemistry, was recently developed and optimized within conventional immortalized cell lines. However, its value within a more fragile and complicated cellular system such as neurons, as well as for transcriptome-wide expression profiling, has yet to be demonstrated. Here, we report the visualization and sequencing of activity-dependent nascent RNA derived from neurons using this labeling method. This work has important implications for improving transcriptome-wide expression profiling and visualization of nascent RNA in neurons, which has the potential to provide valuable insights into the mechanisms underlying neural plasticity, learning, and memory.

The resemblance and disparity of gene expression in dormant and non-dormant seeds and crown buds of leafy spurge (Euphorbia esula)

USDA-ARS?s Scientific Manuscript database

Overlaps in transcriptome profiles between different phases of bud and seed dormancy have not been determined. Thus, we compared various phases of dormancy between seeds and buds to identify common genes and molecular processes. Cluster analysis of expression profiles for 201 selected genes indicate...

GENE EXPRESSION CHANGES IN ARABIDOPSIS THALIANA SEEDLING ROOTS EXPOSED TO THE MUNITION HEXAHYDRO-1,3,5-TRINITRO-1,3,5-TRIAZINE

EPA Science Inventory

Arabidopsis thaliana root transcriptome responses to the munition, hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), were assessed using serial analysis of gene expression (SAGE). Comparison of the transcriptional profile for the RDX response to a profile previously described for Ar...

Comparative transcriptome analysis by RNAseq of necrotic enteritis Clostridium perfringens during in vivo colonization and in vitro conditions.

PubMed

Parreira, Valeria R; Russell, Kay; Athanasiadou, Spiridoula; Prescott, John F

2016-08-12

Necrotic enteritis (NE) caused by netB-positive type A Clostridium perfringens is an important bacterial disease of poultry. Through its complex regulatory system, C. perfringens orchestrates the expression of a collection of toxins and extracellular enzymes that are crucial for the development of the disease; environmental conditions play an important role in their regulation. In this study, and for the first time, global transcriptomic analysis was performed on ligated intestinal loops in chickens colonized with a netB-positive C. perfringens strain, as well as the same strain propagated in vitro under various nutritional and environmental conditions. Analysis of the respective pathogen transcriptomes revealed up to 673 genes that were significantly expressed in vivo. Gene expression profiles in vivo were most similar to those of C. perfringens grown in nutritionally-deprived conditions. Taken together, our results suggest a bacterial transcriptome responses to the early stages of adaptation, and colonization of, the chicken intestine. Our work also reveals how netB-positive C. perfringens reacts to different environmental conditions including those in the chicken intestine.

Assessing mechanisms of toxicant response in the amphipod Melita plumulosa through transcriptomic profiling.

PubMed

Hook, Sharon E; Osborn, Hannah L; Spadaro, David A; Simpson, Stuart L

2014-01-01

This study describes the function of transcripts with altered abundance in the epibenthic amphipod, Melita plumulosa, following whole-sediment exposure to a series of common environmental contaminants. M. plumulosa were exposed for 48 h to sediments spiked and equilibrated with the following contaminants at concentrations predicted to cause sublethal effects to reproduction: porewater ammonia 30 mg L(-1); bifenthrin at 100 μg kg(-1); fipronil at 50 μg kg(-1); 0.6% diesel; 0.3% crude oil; 250 mg Cu kg(-1); 400 mg Ni kg(-1); and 400 mg Zn kg(-1). RNA was extracted and hybridized against a custom Agilent microarray developed for this species. Although the microarray represented a partial transcriptome and not all features on the array could be annotated, unique transcriptomic profiles were generated for each of the contaminant exposures. Hierarchical clustering grouped the expression profiles together by contaminant class, with copper and zinc, the petroleum products and nickel, and the pesticides each forming a distinct cluster. Many of the transcriptional changes observed were consistent with patterns previously described in other crustaceans. The changes in the transcriptome demonstrated that contaminant exposure caused changes in digestive function, growth and moulting, and the cytoskeleton following metal exposure, whereas exposure to petroleum products caused changes in carbohydrate metabolism, xenobiotic metabolism and hormone cycling. Functional analysis of these gene expression profiles can provide a better understanding of modes of toxic action and permits the prediction of mixture effects within contaminated ecosystems. Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.

De novo transcriptome analysis of rose-scented geranium provides insights into the metabolic specificity of terpene and tartaric acid biosynthesis.

PubMed

Narnoliya, Lokesh K; Kaushal, Girija; Singh, Sudhir P; Sangwan, Rajender S

2017-01-13

Rose-scented geranium (Pelargonium sp.) is a perennial herb that produces a high value essential oil of fragrant significance due to the characteristic compositional blend of rose-oxide and acyclic monoterpenoids in foliage. Recently, the plant has also been shown to produce tartaric acid in leaf tissues. Rose-scented geranium represents top-tier cash crop in terms of economic returns and significance of the plant and plant products. However, there has hardly been any study on its metabolism and functional genomics, nor any genomic expression dataset resource is available in public domain. Therefore, to begin the gains in molecular understanding of specialized metabolic pathways of the plant, de novo sequencing of rose-scented geranium leaf transcriptome, transcript assembly, annotation, expression profiling as well as their validation were carried out. De novo transcriptome analysis resulted a total of 78,943 unique contigs (average length: 623 bp, and N50 length: 752 bp) from 15.44 million high quality raw reads. In silico functional annotation led to the identification of several putative genes representing terpene, ascorbic acid and tartaric acid biosynthetic pathways, hormone metabolism, and transcription factors. Additionally, a total of 6,040 simple sequence repeat (SSR) motifs were identified in 6.8% of the expressed transcripts. The highest frequency of SSR was of tri-nucleotides (50%). Further, transcriptome assembly was validated for randomly selected putative genes by standard PCR-based approach. In silico expression profile of assembled contigs were validated by real-time PCR analysis of selected transcripts. Being the first report on transcriptome analysis of rose-scented geranium the data sets and the leads and directions reflected in this investigation will serve as a foundation for pursuing and understanding molecular aspects of its biology, and specialized metabolic pathways, metabolic engineering, genetic diversity as well as molecular breeding.

Transcriptomic signatures in seeds of apple (Malus domestica L. Borkh) during fruitlet abscission.

PubMed

Ferrero, Sergio; Carretero-Paulet, Lorenzo; Mendes, Marta Adelina; Botton, Alessandro; Eccher, Giulia; Masiero, Simona; Colombo, Lucia

2015-01-01

Abscission is the regulated process of detachment of an organ from a plant. In apple the abscission of fruits occurs during their early development to control the fruit load depending on the nutritional state of the plant. In order to control production and obtain fruits with optimal market qualities, the horticultural procedure of thinning is performed to further reduce the number of fruitlets. In this study we have conducted a transcriptomic profiling of seeds from two different types of fruitlets, according to size and position in the fruit cluster. Transcriptomic profiles of central and lateral fruit seeds were obtained by RNAseq. Comparative analysis was performed by the functional categorization of differentially expressed genes by means of Gene Ontology (GO) annotation of the apple genome. Our results revealed the overexpression of genes involved in responses to stress, hormone biosynthesis and also the response and/or transport of auxin and ethylene. A smaller set of genes, mainly related to ion transport and homeostasis, were found to be down-regulated. The transcriptome characterization described in this manuscript contributes to unravelling the molecular mechanisms and pathways involved in the physiological abscission of apple fruits and suggests a role for seeds in this process.

Transcriptomic Signatures in Seeds of Apple (Malus domestica L. Borkh) during Fruitlet Abscission

PubMed Central

Ferrero, Sergio; Carretero-Paulet, Lorenzo; Mendes, Marta Adelina; Botton, Alessandro; Eccher, Giulia; Masiero, Simona; Colombo, Lucia

2015-01-01

Abscission is the regulated process of detachment of an organ from a plant. In apple the abscission of fruits occurs during their early development to control the fruit load depending on the nutritional state of the plant. In order to control production and obtain fruits with optimal market qualities, the horticultural procedure of thinning is performed to further reduce the number of fruitlets. In this study we have conducted a transcriptomic profiling of seeds from two different types of fruitlets, according to size and position in the fruit cluster. Transcriptomic profiles of central and lateral fruit seeds were obtained by RNAseq. Comparative analysis was performed by the functional categorization of differentially expressed genes by means of Gene Ontology (GO) annotation of the apple genome. Our results revealed the overexpression of genes involved in responses to stress, hormone biosynthesis and also the response and/or transport of auxin and ethylene. A smaller set of genes, mainly related to ion transport and homeostasis, were found to be down-regulated. The transcriptome characterization described in this manuscript contributes to unravelling the molecular mechanisms and pathways involved in the physiological abscission of apple fruits and suggests a role for seeds in this process. PMID:25781174

Global transcriptome profiling reveals molecular mechanisms of metal tolerance in a chronically exposed wild population of brown trout.

PubMed

Uren Webster, T M; Bury, N; van Aerle, R; Santos, E M

2013-08-06

Worldwide, a number of viable populations of fish are found in environments heavily contaminated with metals, including brown trout (Salmo trutta) inhabiting the River Hayle in South-West of England. This population is chronically exposed to a water-borne mixture of metals, including copper and zinc, at concentrations lethal to naïve fish. We aimed to investigate the molecular mechanisms employed by the River Hayle brown trout to tolerate high metal concentrations. To achieve this, we combined tissue metal analysis with whole-transcriptome profiling using RNA-seq on an Illumina platform. Metal concentrations in the Hayle trout, compared to fish from a relatively unimpacted river, were significantly increased in the gills, liver and kidney (63-, 34- and 19-fold respectively), but not the gut. This confirms that these fish can tolerate considerable metal accumulation, highlighting the importance of these tissues in metal uptake (gill), storage and detoxification (liver, kidney). We sequenced, assembled and annotated the brown trout transcriptome using a de novo approach. Subsequent gene expression analysis identified 998 differentially expressed transcripts and functional analysis revealed that metal- and ion-homeostasis pathways are likely to be the most important mechanisms contributing to the metal tolerance exhibited by this population.

Global Transcriptome Profiling Reveals Molecular Mechanisms of Metal Tolerance in a Chronically Exposed Wild Population of Brown Trout

PubMed Central

2013-01-01

Worldwide, a number of viable populations of fish are found in environments heavily contaminated with metals, including brown trout (Salmo trutta) inhabiting the River Hayle in South-West of England. This population is chronically exposed to a water-borne mixture of metals, including copper and zinc, at concentrations lethal to naïve fish. We aimed to investigate the molecular mechanisms employed by the River Hayle brown trout to tolerate high metal concentrations. To achieve this, we combined tissue metal analysis with whole-transcriptome profiling using RNA-seq on an Illumina platform. Metal concentrations in the Hayle trout, compared to fish from a relatively unimpacted river, were significantly increased in the gills, liver and kidney (63-, 34- and 19-fold respectively), but not the gut. This confirms that these fish can tolerate considerable metal accumulation, highlighting the importance of these tissues in metal uptake (gill), storage and detoxification (liver, kidney). We sequenced, assembled and annotated the brown trout transcriptome using a de novo approach. Subsequent gene expression analysis identified 998 differentially expressed transcripts and functional analysis revealed that metal- and ion-homeostasis pathways are likely to be the most important mechanisms contributing to the metal tolerance exhibited by this population. PMID:23834071

SAGE ANALYSIS OF TRANSCRIPTOME RESPONSES IN ARABIDOPSIS ROOTS EXPOSED TO 2,4,6-TRINITROTOLUENE

EPA Science Inventory

Serial Analysis of Gene Expression (SAGE) was used to profile transcript levels in Arabidopsis thaliana roots and assess their responses to 2,4,6-trinitrotoluene (TNT) exposure. SAGE libraries representing control and TNT-exposed seedling root transcripts were constructed, and ea...

Comparative Transcriptomic Characterization of the Early Development in Pacific White Shrimp Litopenaeus vannamei

PubMed Central

Wei, Jiankai; Zhang, Xiaojun; Yu, Yang; Huang, Hao; Li, Fuhua; Xiang, Jianhai

2014-01-01

Penaeid shrimp has a distinctive metamorphosis stage during early development. Although morphological and biochemical studies about this ontogeny have been developed for decades, researches on gene expression level are still scarce. In this study, we have investigated the transcriptomes of five continuous developmental stages in Pacific white shrimp (Litopenaeus vannamei) with high throughput Illumina sequencing technology. The reads were assembled and clustered into 66,815 unigenes, of which 32,398 have putative homologues in nr database, 14,981 have been classified into diverse functional categories by Gene Ontology (GO) annotation and 26,257 have been associated with 255 pathways by KEGG pathway mapping. Meanwhile, the differentially expressed genes (DEGs) between adjacent developmental stages were identified and gene expression patterns were clustered. By GO term enrichment analysis, KEGG pathway enrichment analysis and functional gene profiling, the physiological changes during shrimp metamorphosis could be better understood, especially histogenesis, diet transition, muscle development and exoskeleton reconstruction. In conclusion, this is the first study that characterized the integrated transcriptomic profiles during early development of penaeid shrimp, and these findings will serve as significant references for shrimp developmental biology and aquaculture research. PMID:25197823

Transcriptional profiling: a potential anti-doping strategy.

PubMed

Rupert, J L

2009-12-01

Evolving challenges require evolving responses. The use of illicit performance enhancing drugs by athletes permeates the reality and the perception of elite sports. New drugs with ergogenic or masking potential are quickly adopted, driven by a desire to win and the necessity of avoiding detection. To counter this trend, anti-doping authorities are continually refining existing assays and developing new testing strategies. In the post-genome era, genetic- and molecular-based tests are being evaluated as potential approaches to detect new and sophisticated forms of doping. Transcriptome analysis, in which a tissue's complement of mRNA transcripts is characterized, is one such method. The quantity and composition of a tissue's transcriptome is highly reflective of milieu and metabolic activity. There is much interest in transcriptional profiling in medical diagnostics and, as transcriptional information can be obtained from a variety of easily accessed tissues, similar approaches could be used in doping control. This article briefly reviews current understanding of the transcriptome, common methods of global analysis of gene expression and non-invasive sample sources. While the focus of this article is on anti-doping, the principles and methodology described could be applied to any research in which non-invasive, yet biologically informative sampling is desired.

The study of transcriptome profiles in Holstein cows with miscarriage during peri-implantation.

PubMed

Zhao, Guoli; Li, Yanyan; Kang, Xiaolong; Huang, Liang; Li, Peng; Zhou, Jinghang; Shi, Yuangang

2018-05-31

In this study, the transcriptome profile of cows who experienced miscarriage during peri-implantation was investigated. The transcriptome was checked by RNA sequencing, and the analyzed by bioinformatics methods. The results suggested that serum progesterone levels were significantly decreased in the cows who miscarried compared with the pregnant cows at 18 d, 21d, 33 d, 39 d and 51 d after artificial insemination. The RNA sequencing results suggested that 32, 176, 5, 10 and 2 differentially expressed genes (DEGs) were identified in the pregnant cows and the cows who miscarried at 18, 21, 33, 39 and 51 d after artificial insemination. Furthermore, the DEGs were analysed with hierarchical clustering and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and 15, 101, 1, 2 and 2 DEGs were upregulated, and 17, 74, 4, 8 and 0 DEGs were downregulated in the cows in the pregnant and miscarriage groups, respectively at 18, 21 33, 39 and 51 d after artificial insemination. These DEGs were distributed to 13, 20, 3, 6 and 20 pathways. This analysis has identified genes and pathways crucial for pregnancy and miscarriage in cows.

RAS oncogene-mediated deregulation of the transcriptome: from molecular signature to function.

PubMed

Schäfer, Reinhold; Sers, Christine

2011-01-01

Transcriptome analysis of cancer cells has developed into a standard procedure to elucidate multiple features of the malignant process and to link gene expression to clinical properties. Gene expression profiling based on microarrays provides essentially correlative information and needs to be transferred to the functional level in order to understand the activity and contribution of individual genes or sets of genes as elements of the gene signature. To date, there exist significant gaps in the functional understanding of gene expression profiles. Moreover, the processes that drive the profound transcriptional alterations that characterize cancer cells remain mainly elusive. We have used pathway-restricted gene expression profiles derived from RAS oncogene-transformed cells and from RAS-expressing cancer cells to identify regulators downstream of the MAPK pathway.We describe the role of epigenetic regulation exemplified by the control of several immune genes in generic cell lines and colorectal cancer cells, particularly the functional interaction between signaling and DNA methylation. Moreover, we assess the role of the architectural transcription factor high mobility AT-hook 2 (HMGA2) as a regulator of the RAS-responsive transcriptome in ovarian epithelial cells. Finally, we describe an integrated approach combining pathway interference in colorectal cancer cells, gene expression profiling and computational analysis of regulatory elements of deregulated target genes. This strategy resulted in the identification of Y-box binding protein 1 (YBX1) as a regulator of MAPK-dependent proliferation and gene expression. The implications for a therapeutic application of HMGA2 gene silencing and the role of YBX1 as a prognostic factor are discussed.

Comparative transcriptomics between Synechococcus PCC 7942 and Synechocystis PCC 6803 provide insights into mechanisms of adaptation to stress.

DOE PAGES

Konstantinos, Billis; Billini, Maria; Tripp, Harry J.; ...

2014-09-23

Background: Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 are model cyanobacteria from which the metabolism and adaptive responses of other cyanobacteria are inferred. Here we report the gene expression response of these two strains to a variety of nutrient and environmental stresses of varying duration, using transcriptomics. Our data comprise both stranded and 5' enriched libraries in order to elucidate many aspects of the transcriptome. Results: Both organisms were exposed to stress conditions due to nutrient deficiency (inorganic carbon) or change of environmental conditions (salinity, temperature, pH, light) sampled at 1 and 24 hours after the application ofmore » stress. The transcriptome profile of each strain revealed similarities and differences in gene expression for photosynthetic and respiratory electron transport chains and carbon fixation. Transcriptome profiles also helped us improve the structural annotation of the genome and identify possible missed genes (including anti-sense) and determine transcriptional units (operons). Finally, we predicted association of proteins of unknown function biochemical pathways by associating them to well-characterized ones based on their transcript levels correlation. Conclusions: Overall, this study results an informative annotation of those species and the comparative analysis of the response of the two organisms revealed similarities but also significant changes in the way they respond to external stress and the duration of the response« less

Comparative transcriptomics between Synechococcus PCC 7942 and Synechocystis PCC 6803 provide insights into mechanisms of adaptation to stress.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Konstantinos, Billis; Billini, Maria; Tripp, Harry J.

Background: Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 are model cyanobacteria from which the metabolism and adaptive responses of other cyanobacteria are inferred. Here we report the gene expression response of these two strains to a variety of nutrient and environmental stresses of varying duration, using transcriptomics. Our data comprise both stranded and 5' enriched libraries in order to elucidate many aspects of the transcriptome. Results: Both organisms were exposed to stress conditions due to nutrient deficiency (inorganic carbon) or change of environmental conditions (salinity, temperature, pH, light) sampled at 1 and 24 hours after the application ofmore » stress. The transcriptome profile of each strain revealed similarities and differences in gene expression for photosynthetic and respiratory electron transport chains and carbon fixation. Transcriptome profiles also helped us improve the structural annotation of the genome and identify possible missed genes (including anti-sense) and determine transcriptional units (operons). Finally, we predicted association of proteins of unknown function biochemical pathways by associating them to well-characterized ones based on their transcript levels correlation. Conclusions: Overall, this study results an informative annotation of those species and the comparative analysis of the response of the two organisms revealed similarities but also significant changes in the way they respond to external stress and the duration of the response« less

Modular organization of the white spruce (Picea glauca) transcriptome reveals functional organization and evolutionary signatures.

PubMed

Raherison, Elie S M; Giguère, Isabelle; Caron, Sébastien; Lamara, Mebarek; MacKay, John J

2015-07-01

Transcript profiling has shown the molecular bases of several biological processes in plants but few studies have developed an understanding of overall transcriptome variation. We investigated transcriptome structure in white spruce (Picea glauca), aiming to delineate its modular organization and associated functional and evolutionary attributes. Microarray analyses were used to: identify and functionally characterize groups of co-expressed genes; investigate expressional and functional diversity of vascular tissue preferential genes which were conserved among Picea species, and identify expression networks underlying wood formation. We classified 22 857 genes as variable (79%; 22 coexpression groups) or invariant (21%) by profiling across several vegetative tissues. Modular organization and complex transcriptome restructuring among vascular tissue preferential genes was revealed by their assignment to coexpression groups with partially overlapping profiles and partially distinct functions. Integrated analyses of tissue-based and temporally variable profiles identified secondary xylem gene networks, showed their remodelling over a growing season and identified PgNAC-7 (no apical meristerm (NAM), Arabidopsis transcription activation factor (ATAF) and cup-shaped cotyledon (CUC) transcription factor 007 in Picea glauca) as a major hub gene specific to earlywood formation. Reference profiling identified comprehensive, statistically robust coexpressed groups, revealing that modular organization underpins the evolutionary conservation of the transcriptome structure. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Transcriptomic markers meet the real world: finding diagnostic signatures of corticosteroid treatment in commercial beef samples

PubMed Central

2012-01-01

Background The use of growth-promoters in beef cattle, despite the EU ban, remains a frequent practice. The use of transcriptomic markers has already proposed to identify indirect evidence of anabolic hormone treatment. So far, such approach has been tested in experimentally treated animals. Here, for the first time commercial samples were analyzed. Results Quantitative determination of Dexamethasone (DEX) residues in the urine collected at the slaughterhouse was performed by Liquid Chromatography-Mass Spectrometry (LC-MS). DNA-microarray technology was used to obtain transcriptomic profiles of skeletal muscle in commercial samples and negative controls. LC-MS confirmed the presence of low level of DEX residues in the urine of the commercial samples suspect for histological classification. Principal Component Analysis (PCA) on microarray data identified two clusters of samples. One cluster included negative controls and a subset of commercial samples, while a second cluster included part of the specimens collected at the slaughterhouse together with positives for corticosteroid treatment based on thymus histology and LC-MS. Functional analysis of the differentially expressed genes (3961) between the two groups provided further evidence that animals clustering with positive samples might have been treated with corticosteroids. These suspect samples could be reliably classified with a specific classification tool (Prediction Analysis of Microarray) using just two genes. Conclusions Despite broad variation observed in gene expression profiles, the present study showed that DNA-microarrays can be used to find transcriptomic signatures of putative anabolic treatments and that gene expression markers could represent a useful screening tool. PMID:23110699

Digital Marine Bioprospecting: Mining New Neurotoxin Drug Candidates from the Transcriptomes of Cold-Water Sea Anemones

PubMed Central

Urbarova, Ilona; Karlsen, Bård Ove; Okkenhaug, Siri; Seternes, Ole Morten; Johansen, Steinar D.; Emblem, Åse

2012-01-01

Marine bioprospecting is the search for new marine bioactive compounds and large-scale screening in extracts represents the traditional approach. Here, we report an alternative complementary protocol, called digital marine bioprospecting, based on deep sequencing of transcriptomes. We sequenced the transcriptomes from the adult polyp stage of two cold-water sea anemones, Bolocera tuediae and Hormathia digitata. We generated approximately 1.1 million quality-filtered sequencing reads by 454 pyrosequencing, which were assembled into approximately 120,000 contigs and 220,000 single reads. Based on annotation and gene ontology analysis we profiled the expressed mRNA transcripts according to known biological processes. As a proof-of-concept we identified polypeptide toxins with a potential blocking activity on sodium and potassium voltage-gated channels from digital transcriptome libraries. PMID:23170083

De novo assembly and characterization of Muscovy duck liver transcriptome and analysis of differentially regulated genes in response to heat stress.

PubMed

Zeng, Tao; Zhang, Liping; Li, Jinjun; Wang, Deqian; Tian, Yong; Lu, Lizhi

2015-05-01

High temperature is a major abiotic stress limiting animal growth and productivity worldwide. The Muscovy duck (Cairina moschata), sometimes called the Barbary drake, is a type of duck with a fairly unusual domestication history. In Southeast Asia, duck meat is one of the top meats consumed, and as such, the production of the meat is an important topic of research. The transcriptomic and genomic data presently available are insufficient to understanding the molecular mechanism underlying the heat tolerance of Muscovy ducks. Thus, transcriptome and expression profiling data for this species are required as important resource for identifying genes and developing molecular marker. In this study, de novo transcriptome assembly and gene expression analysis using Illumina sequencing technology were performed. More than 225 million clean reads were generated and assembled into 36,903 unique transcripts with an average length of 1,135 bp. A total of 21,221 (57.50 %) unigenes were annotated. Gene Ontology (GO) analysis of the annotated unigenes revealed that the majority of sequenced genes were associated with transcription, signal transduction, and apoptosis. We also performed gene expression profiling analysis upon heat treatment in Muscovy ducks and identified 470 heat-response unique transcripts. GO term enrichment showed that protein folding and chaperone binding were significant enrichment, whereas KEGG pathway analyses showed that Ras and MAPKs were activated after heat stress in Muscovy ducks. Our research enriched sequences information of Muscovy duck, provided novel insights into responses to heat stress in these ducks, and serve as candidate genes or markers that can be used to guide future efforts to breed heat-tolerant duck strains.

Stem cell and neurogenic gene-expression profiles link prostate basal cells to aggressive prostate cancer

PubMed Central

Zhang, Dingxiao; Park, Daechan; Zhong, Yi; Lu, Yue; Rycaj, Kiera; Gong, Shuai; Chen, Xin; Liu, Xin; Chao, Hsueh-Ping; Whitney, Pamela; Calhoun-Davis, Tammy; Takata, Yoko; Shen, Jianjun; Iyer, Vishwanath R.; Tang, Dean G.

2016-01-01

The prostate gland mainly contains basal and luminal cells constructed as a pseudostratified epithelium. Annotation of prostate epithelial transcriptomes provides a foundation for discoveries that can impact disease understanding and treatment. Here we describe a genome-wide transcriptome analysis of human benign prostatic basal and luminal epithelial populations using deep RNA sequencing. Through molecular and biological characterizations, we show that the differential gene-expression profiles account for their distinct functional properties. Strikingly, basal cells preferentially express gene categories associated with stem cells, neurogenesis and ribosomal RNA (rRNA) biogenesis. Consistent with this profile, basal cells functionally exhibit intrinsic stem-like and neurogenic properties with enhanced rRNA transcription activity. Of clinical relevance, the basal cell gene-expression profile is enriched in advanced, anaplastic, castration-resistant and metastatic prostate cancers. Therefore, we link the cell-type-specific gene signatures to aggressive subtypes of prostate cancer and identify gene signatures associated with adverse clinical features. PMID:26924072

Stem cell and neurogenic gene-expression profiles link prostate basal cells to aggressive prostate cancer.

PubMed

Zhang, Dingxiao; Park, Daechan; Zhong, Yi; Lu, Yue; Rycaj, Kiera; Gong, Shuai; Chen, Xin; Liu, Xin; Chao, Hsueh-Ping; Whitney, Pamela; Calhoun-Davis, Tammy; Takata, Yoko; Shen, Jianjun; Iyer, Vishwanath R; Tang, Dean G

2016-02-29

The prostate gland mainly contains basal and luminal cells constructed as a pseudostratified epithelium. Annotation of prostate epithelial transcriptomes provides a foundation for discoveries that can impact disease understanding and treatment. Here we describe a genome-wide transcriptome analysis of human benign prostatic basal and luminal epithelial populations using deep RNA sequencing. Through molecular and biological characterizations, we show that the differential gene-expression profiles account for their distinct functional properties. Strikingly, basal cells preferentially express gene categories associated with stem cells, neurogenesis and ribosomal RNA (rRNA) biogenesis. Consistent with this profile, basal cells functionally exhibit intrinsic stem-like and neurogenic properties with enhanced rRNA transcription activity. Of clinical relevance, the basal cell gene-expression profile is enriched in advanced, anaplastic, castration-resistant and metastatic prostate cancers. Therefore, we link the cell-type-specific gene signatures to aggressive subtypes of prostate cancer and identify gene signatures associated with adverse clinical features.

Quantifying whole transcriptome size, a prerequisite for understanding transcriptome evolution across species: an example from a plant allopolyploid.

PubMed

Coate, Jeremy E; Doyle, Jeff J

2010-01-01

Evolutionary biologists are increasingly comparing gene expression patterns across species. Due to the way in which expression assays are normalized, such studies provide no direct information about expression per gene copy (dosage responses) or per cell and can give a misleading picture of genes that are differentially expressed. We describe an assay for estimating relative expression per cell. When used in conjunction with transcript profiling data, it is possible to compare the sizes of whole transcriptomes, which in turn makes it possible to compare expression per cell for each gene in the transcript profiling data set. We applied this approach, using quantitative reverse transcriptase-polymerase chain reaction and high throughput RNA sequencing, to a recently formed allopolyploid and showed that its leaf transcriptome was approximately 1.4-fold larger than either progenitor transcriptome (70% of the sum of the progenitor transcriptomes). In contrast, the allopolyploid genome is 94.3% as large as the sum of its progenitor genomes and retains > or =93.5% of the sum of its progenitor gene complements. Thus, "transcriptome downsizing" is greater than genome downsizing. Using this transcriptome size estimate, we inferred dosage responses for several thousand genes and showed that the majority exhibit partial dosage compensation. Homoeologue silencing is nonrandomly distributed across dosage responses, with genes showing extreme responses in either direction significantly more likely to have a silent homoeologue. This experimental approach will add value to transcript profiling experiments involving interspecies and interploidy comparisons by converting expression per transcriptome to expression per genome, eliminating the need for assumptions about transcriptome size.

The Isolation of Pure Populations of Neurons by Laser Capture Microdissection: Methods and Application in Neuroscience.

PubMed

Morris, Renée; Mehta, Prachi

2018-01-01

In mammals, the central nervous system (CNS) is constituted of various cellular elements, posing a challenge to isolating specific cell types to investigate their expression profile. As a result, tissue homogenization is not amenable to analyses of motor neurons profiling as these represent less than 10% of the total spinal cord cell population. One way to tackle the problem of tissue heterogeneity and obtain meaningful genomic, proteomic, and transcriptomic profiling is to use laser capture microdissection technology (LCM). In this chapter, we describe protocols for the capture of isolated populations of motor neurons from spinal cord tissue sections and for downstream transcriptomic analysis of motor neurons with RT-PCR. We have also included a protocol for the immunological confirmation that the captured neurons are indeed motor neurons. Although focused on spinal cord motor neurons, these protocols can be easily optimized for the isolation of any CNS neurons.

Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

PubMed Central

Park, Paul J.; Fuchs, Robert; Wei, Lai; Jorgensen, Brian G.; Redelman, Doug; Ward, Sean M.; Sanders, Kenton M.

2017-01-01

Transcriptome-scale data can reveal essential clues into understanding the underlying molecular mechanisms behind specific cellular functions and biological processes. Transcriptomics is a continually growing field of research utilized in biomarker discovery. The transcriptomic profile of interstitial cells of Cajal (ICC), which serve as slow-wave electrical pacemakers for gastrointestinal (GI) smooth muscle, has yet to be uncovered. Using copGFP-labeled ICC mice and flow cytometry, we isolated ICC populations from the murine small intestine and colon and obtained their transcriptomes. In analyzing the transcriptome, we identified a unique set of ICC-restricted markers including transcription factors, epigenetic enzymes/regulators, growth factors, receptors, protein kinases/phosphatases, and ion channels/transporters. This analysis provides new and unique insights into the cellular and biological functions of ICC in GI physiology. Additionally, we constructed an interactive ICC genome browser (http://med.unr.edu/physio/transcriptome) based on the UCSC genome database. To our knowledge, this is the first online resource that provides a comprehensive library of all known genetic transcripts expressed in primary ICC. Our genome browser offers a new perspective into the alternative expression of genes in ICC and provides a valuable reference for future functional studies. PMID:28426719

The application of transcriptomic data in the authentication of beef derived from contrasting production systems.

PubMed

Sweeney, Torres; Lejeune, Alex; Moloney, Aidan P; Monahan, Frank J; Gettigan, Paul Mc; Downey, Gerard; Park, Stephen D E; Ryan, Marion T

2016-09-21

Differences between cattle production systems can influence the nutritional and sensory characteristics of beef, in particular its fatty acid (FA) composition. As beef products derived from pasture-based systems can demand a higher premium from consumers, there is a need to understand the biological characteristics of pasture produced meat and subsequently to develop methods of authentication for these products. Here, we describe an approach to authentication that focuses on differences in the transcriptomic profile of muscle from animals finished in different systems of production of practical relevance to the Irish beef industry. The objectives of this study were to identify a panel of differentially expressed (DE) genes/networks in the muscle of cattle raised outdoors on pasture compared to animals raised indoors on a concentrate based diet and to subsequently identify an optimum panel which can classify the meat based on a production system. A comparison of the muscle transcriptome of outdoor/pasture-fed and Indoor/concentrate-fed cattle resulted in the identification of 26 DE genes. Functional analysis of these genes identified two significant networks (1: Energy Production, Lipid Metabolism, Small Molecule Biochemistry; and 2: Lipid Metabolism, Molecular Transport, Small Molecule Biochemistry), both of which are involved in FA metabolism. The expression of selected up-regulated genes in the outdoor/pasture-fed animals correlated positively with the total n-3 FA content of the muscle. The pathway and network analysis of the DE genes indicate that peroxisome proliferator-activated receptor (PPAR) and FYN/AMPK could be implicit in the regulation of these alterations to the lipid profile. In terms of authentication, the expression profile of three DE genes (ALAD, EIF4EBP1 and NPNT) could almost completely separate the samples based on production system (95 % authentication for animals on pasture-based and 100 % for animals on concentrate- based diet) in this context. The majority of DE genes between muscle of the outdoor/pasture-fed and concentrate-fed cattle were related to lipid metabolism and in particular β-oxidation. In this experiment the combined expression profiles of ALAD, EIF4EBP1 and NPNT were optimal in classifying the muscle transcriptome based on production system. Given the overall lack of comparable studies and variable concordance with those that do exist, the use of transcriptomic data in authenticating production systems requires more exploration across a range of contexts and breeds.

Re-evaluating microglia expression profiles using RiboTag and cell isolation strategies.

PubMed

Haimon, Zhana; Volaski, Alon; Orthgiess, Johannes; Boura-Halfon, Sigalit; Varol, Diana; Shemer, Anat; Yona, Simon; Zuckerman, Binyamin; David, Eyal; Chappell-Maor, Louise; Bechmann, Ingo; Gericke, Martin; Ulitsky, Igor; Jung, Steffen

2018-06-01

Transcriptome profiling is widely used to infer functional states of specific cell types, as well as their responses to stimuli, to define contributions to physiology and pathophysiology. Focusing on microglia, the brain's macrophages, we report here a side-by-side comparison of classical cell-sorting-based transcriptome sequencing and the 'RiboTag' method, which avoids cell retrieval from tissue context and yields translatome sequencing information. Conventional whole-cell microglial transcriptomes were found to be significantly tainted by artifacts introduced by tissue dissociation, cargo contamination and transcripts sequestered from ribosomes. Conversely, our data highlight the added value of RiboTag profiling for assessing the lineage accuracy of Cre recombinase expression in transgenic mice. Collectively, this study indicates method-based biases, reveals observer effects and establishes RiboTag-based translatome profiling as a valuable complement to standard sorting-based profiling strategies.

Hematopoietic Lineage Transcriptome Stability and Representation in PAXgene™ Collected Peripheral Blood Utilising SPIA Single-Stranded cDNA Probes for Microarray

PubMed Central

Kennedy, Laura; Vass, J. Keith; Haggart, D. Ross; Moore, Steve; Burczynski, Michael E.; Crowther, Dan; Miele, Gino

2008-01-01

Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgene™ RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2™ enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgene™ blood samples also advocate a short, fixed room temperature storage time for all PAXgene™ blood samples collected for the purposes of global transcriptional profiling in clinical studies. PMID:19578521

Transcriptome Analysis of Barbarea vulgaris Infested with Diamondback Moth (Plutella xylostella) Larvae

PubMed Central

Shen, Di; Wang, Haiping; Wu, Qingjun; Lu, Peng; Qiu, Yang; Song, Jiangping; Zhang, Youjun; Li, Xixiang

2013-01-01

Background The diamondback moth (DBM, Plutella xylostella) is a crucifer-specific pest that causes significant crop losses worldwide. Barbarea vulgaris (Brassicaceae) can resist DBM and other herbivorous insects by producing feeding-deterrent triterpenoid saponins. Plant breeders have long aimed to transfer this insect resistance to other crops. However, a lack of knowledge on the biosynthetic pathways and regulatory networks of these insecticidal saponins has hindered their practical application. A pyrosequencing-based transcriptome analysis of B. vulgaris during DBM larval feeding was performed to identify genes and gene networks responsible for saponin biosynthesis and its regulation at the genome level. Principal Findings Approximately 1.22, 1.19, 1.16, 1.23, 1.16, 1.20, and 2.39 giga base pairs of clean nucleotides were generated from B. vulgaris transcriptomes sampled 1, 4, 8, 12, 24, and 48 h after onset of P. xylostella feeding and from non-inoculated controls, respectively. De novo assembly using all data of the seven transcriptomes generated 39,531 unigenes. A total of 37,780 (95.57%) unigenes were annotated, 14,399 of which were assigned to one or more gene ontology terms and 19,620 of which were assigned to 126 known pathways. Expression profiles revealed 2,016–4,685 up-regulated and 557–5188 down-regulated transcripts. Secondary metabolic pathways, such as those of terpenoids, glucosinolates, and phenylpropanoids, and its related regulators were elevated. Candidate genes for the triterpene saponin pathway were found in the transcriptome. Orthological analysis of the transcriptome with four other crucifer transcriptomes identified 592 B. vulgaris-specific gene families with a P-value cutoff of 1e−5. Conclusion This study presents the first comprehensive transcriptome analysis of B. vulgaris subjected to a series of DBM feedings. The biosynthetic and regulatory pathways of triterpenoid saponins and other DBM deterrent metabolites in this plant were classified. The results of this study will provide useful data for future investigations on pest-resistance phytochemistry and plant breeding. PMID:23696897

Transcriptomic Immune Response of Tenebrio molitor Pupae to Parasitization by Scleroderma guani

PubMed Central

Zhu, Jia-Ying; Yang, Pu; Zhang, Zhong; Wu, Guo-Xing; Yang, Bin

2013-01-01

Background Host and parasitoid interaction is one of the most fascinating relationships of insects, which is currently receiving an increasing interest. Understanding the mechanisms evolved by the parasitoids to evade or suppress the host immune system is important for dissecting this interaction, while it was still poorly known. In order to gain insight into the immune response of Tenebrio molitor to parasitization by Scleroderma guani, the transcriptome of T. molitor pupae was sequenced with focus on immune-related gene, and the non-parasitized and parasitized T. molitor pupae were analyzed by digital gene expression (DGE) analysis with special emphasis on parasitoid-induced immune-related genes using Illumina sequencing. Methodology/Principal Findings In a single run, 264,698 raw reads were obtained. De novo assembly generated 71,514 unigenes with mean length of 424 bp. Of those unigenes, 37,373 (52.26%) showed similarity to the known proteins in the NCBI nr database. Via analysis of the transcriptome data in depth, 430 unigenes related to immunity were identified. DGE analysis revealed that parasitization by S. guani had considerable impacts on the transcriptome profile of T. molitor pupae, as indicated by the significant up- or down-regulation of 3,431 parasitism-responsive transcripts. The expression of a total of 74 unigenes involved in immune response of T. molitor was significantly altered after parasitization. Conclusions/Significance obtained T. molitor transcriptome, in addition to establishing a fundamental resource for further research on functional genomics, has allowed the discovery of a large group of immune genes that might provide a meaningful framework to better understand the immune response in this species and other beetles. The DGE profiling data provides comprehensive T. molitor immune gene expression information at the transcriptional level following parasitization, and sheds valuable light on the molecular understanding of the host-parasitoid interaction. PMID:23342153

RNA sequencing-based longitudinal transcriptomic profiling gives novel insights into the disease mechanism of generalized pustular psoriasis.

PubMed

Wang, Lingyan; Yu, Xiaoling; Wu, Chao; Zhu, Teng; Wang, Wenming; Zheng, Xiaofeng; Jin, Hongzhong

2018-06-05

Generalized pustular psoriasis (GPP) is a rare, episodic, potentially life-threatening inflammatory disease. However, the pathogenesis of GPP, and universally accepted therapies for treating it, remain undefined. To better understand the disease mechanism of GPP, we performed a transcriptome analysis to profile the gene expression of peripheral blood mononuclear cells (PBMCs) from patients enrolled at the time of diagnosis and receiving follow-up treatment for up to 6 months. RNA sequencing data revealed that gene expression in five GPP patients' PBMCs was profoundly altered following acitretin treatment. Differentially expressed gene (DEG) analysis suggested that genes related to psoriatic inflammation, including CXCL1, CXCL8 (IL-8), S100A8, S100A9, S100A12 and LCN2, were significantly downregulated in patients in remission from GPP. Functional enrichment and annotation analysis unveiled a cluster of DEGs significantly associated with the function of leukocytes, particularly neutrophils. Pathway analysis suggested that a variety of pro-inflammatory pathways were inhibited in patients in remission. This analysis not only reaffirmed known signaling pathways in GPP pathogenesis, but also implicated novel factors and pathways, such as cell cycle regulation pathways. Furthermore, regulator network analysis provided bioinformatics-based support for upstream molecules as potential therapeutic targets such as oncostatin M. This longitudinal analysis of blood transcriptomes provides the first evidence that dysregulated gene expression in peripheral blood may significantly contribute to psoriatic inflammation in GPP patients. Novel canonical pathways and biomarkers identified in the current research may provide insights to help understand GPP pathobiology and advance novel therapeutics.

Multifaceted role of nitric oxide in an in vitro mouse neuronal injury model: transcriptomic profiling defines the temporal recruitment of death signalling cascades

PubMed Central

Peng, Zhao Feng; Chen, Minghui Jessica; Manikandan, Jayapal; Melendez, Alirio J; Shui, Guanghou; Russo-Marie, Françoise; Whiteman, Matthew; Beart, Philip M; Moore, Philip K; Cheung, Nam Sang

2012-01-01

Abstract Nitric oxide is implicated in the pathogenesis of various neuropathologies characterized by oxidative stress. Although nitric oxide has been reported to be involved in the exacerbation of oxidative stress observed in several neuropathologies, existent data fail to provide a holistic description of how nitrergic pathobiology elicits neuronal injury. Here we provide a comprehensive description of mechanisms contributing to nitric oxide induced neuronal injury by global transcriptomic profiling. Microarray analyses were undertaken on RNA from murine primary cortical neurons treated with the nitric oxide generator DETA-NONOate (NOC-18, 0.5 mM) for 8–24 hrs. Biological pathway analysis focused upon 3672 gene probes which demonstrated at least a ±1.5-fold expression in a minimum of one out of three time-points and passed statistical analysis (one-way anova, P < 0.05). Numerous enriched processes potentially determining nitric oxide mediated neuronal injury were identified from the transcriptomic profile: cell death, developmental growth and survival, cell cycle, calcium ion homeostasis, endoplasmic reticulum stress, oxidative stress, mitochondrial homeostasis, ubiquitin-mediated proteolysis, and GSH and nitric oxide metabolism. Our detailed time-course study of nitric oxide induced neuronal injury allowed us to provide the first time a holistic description of the temporal sequence of cellular events contributing to nitrergic injury. These data form a foundation for the development of screening platforms and define targets for intervention in nitric oxide neuropathologies where nitric oxide mediated injury is causative. PMID:21352476

Transcriptome profiling to identify ATRA-responsive genes in human iPSC-derived endoderm for high-throughput point of departure analysis (SOT Annual Meeting)

EPA Science Inventory

Toxicological tipping points occur at chemical concentrations that overwhelm a cell’s adaptive response leading to permanent effects. We focused on retinoid signaling in differentiating endoderm to identify developmental pathways for tipping point analysis. Human induced pluripot...

Transcriptome analysis and metabolic profiling of green and red kale (Brassica oleracea var. acephala) seedlings.

PubMed

Jeon, Jin; Kim, Jae Kwang; Kim, HyeRan; Kim, Yeon Jeong; Park, Yun Ji; Kim, Sun Ju; Kim, Changsoo; Park, Sang Un

2018-02-15

Kale (Brassica oleracea var. acephala) is a rich source of numerous health-benefiting compounds, including vitamins, glucosinolates, phenolic compounds, and carotenoids. However, the genetic resources for exploiting the phyto-nutritional traits of kales are limited. To acquire precise information on secondary metabolites in kales, we performed a comprehensive analysis of the transcriptome and metabolome of green and red kale seedlings. Kale transcriptome datasets revealed 37,149 annotated genes and several secondary metabolite biosynthetic genes. HPLC analysis revealed 14 glucosinolates, 20 anthocyanins, 3 phenylpropanoids, and 6 carotenoids in the kale seedlings that were examined. Red kale contained more glucosinolates, anthocyanins, and phenylpropanoids than green kale, whereas the carotenoid contents were much higher in green kale than in red kale. Ultimately, our data will be a valuable resource for future research on kale bio-engineering and will provide basic information to define gene-to-metabolite networks in kale. Copyright © 2017 Elsevier Ltd. All rights reserved.

Quantitative developmental transcriptomes of the Mediterranean sea urchin Paracentrotus lividus.

PubMed

Gildor, Tsvia; Malik, Assaf; Sher, Noa; Avraham, Linor; Ben-Tabou de-Leon, Smadar

2016-02-01

Embryonic development progresses through the timely activation of thousands of differentially activated genes. Quantitative developmental transcriptomes provide the means to relate global patterns of differentially expressed genes to the emerging body plans they generate. The sea urchin is one of the classic model systems for embryogenesis and the models of its developmental gene regulatory networks are of the most comprehensive of their kind. Thus, the sea urchin embryo is an excellent system for studies of its global developmental transcriptional profiles. Here we produced quantitative developmental transcriptomes of the sea urchin Paracentrotus lividus (P. lividus) at seven developmental stages from the fertilized egg to prism stage. We generated de-novo reference transcriptome and identified 29,817 genes that are expressed at this time period. We annotated and quantified gene expression at the different developmental stages and confirmed the reliability of the expression profiles by QPCR measurement of a subset of genes. The progression of embryo development is reflected in the observed global expression patterns and in our principle component analysis. Our study illuminates the rich patterns of gene expression that participate in sea urchin embryogenesis and provide an essential resource for further studies of the dynamic expression of P. lividus genes. Copyright © 2015 Elsevier B.V. All rights reserved.

Transcriptional profiling of CD31(+) cells isolated from murine embryonic stem cells.

PubMed

Mariappan, Devi; Winkler, Johannes; Chen, Shuhua; Schulz, Herbert; Hescheler, Jürgen; Sachinidis, Agapios

2009-02-01

Identification of genes involved in endothelial differentiation is of great interest for the understanding of the cellular and molecular mechanisms involved in the development of new blood vessels. Mouse embryonic stem (mES) cells serve as a potential source of endothelial cells for transcriptomic analysis. We isolated endothelial cells from 8-days old embryoid bodies by immuno-magnetic separation using platelet endothelial cell adhesion molecule-1 (also known as CD31) expressed on both early and mature endothelial cells. CD31(+) cells exhibit endothelial-like behavior by being able to incorporate DiI-labeled acetylated low-density lipoprotein as well as form tubular structures on matrigel. Quantitative and semi-quantitative PCR analysis further demonstrated the increased expression of endothelial transcripts. To ascertain the specific transcriptomic identity of the CD31(+) cells, large-scale microarray analysis was carried out. Comparative bioinformatic analysis reveals an enrichment of the gene ontology categories angiogenesis, blood vessel morphogenesis, vasculogenesis and blood coagulation in the CD31(+) cell population. Based on the transcriptomic signatures of the CD31(+) cells, we conclude that this ES cell-derived population contains endothelial-like cells expressing a mesodermal marker BMP2 and possess an angiogenic potential. The transcriptomic characterization of CD31(+) cells enables an in vitro functional genomic model to identify genes required for angiogenesis.

Metabolic engineering of Escherichia coli for the production of l-valine based on transcriptome analysis and in silico gene knockout simulation

PubMed Central

Park, Jin Hwan; Lee, Kwang Ho; Kim, Tae Yong; Lee, Sang Yup

2007-01-01

The l-valine production strain of Escherichia coli was constructed by rational metabolic engineering and stepwise improvement based on transcriptome analysis and gene knockout simulation of the in silico genome-scale metabolic network. Feedback inhibition of acetohydroxy acid synthase isoenzyme III by l-valine was removed by site-directed mutagenesis, and the native promoter containing the transcriptional attenuator leader regions of the ilvGMEDA and ilvBN operon was replaced with the tac promoter. The ilvA, leuA, and panB genes were deleted to make more precursors available for l-valine biosynthesis. This engineered Val strain harboring a plasmid overexpressing the ilvBN genes produced 1.31 g/liter l-valine. Comparative transcriptome profiling was performed during batch fermentation of the engineered and control strains. Among the down-regulated genes, the lrp and ygaZH genes, which encode a global regulator Lrp and l-valine exporter, respectively, were overexpressed. Amplification of the lrp, ygaZH, and lrp-ygaZH genes led to the enhanced production of l-valine by 21.6%, 47.1%, and 113%, respectively. Further improvement was achieved by using in silico gene knockout simulation, which identified the aceF, mdh, and pfkA genes as knockout targets. The VAMF strain (Val ΔaceF Δmdh ΔpfkA) overexpressing the ilvBN, ilvCED, ygaZH, and lrp genes was able to produce 7.55 g/liter l-valine from 20 g/liter glucose in batch culture, resulting in a high yield of 0.378 g of l-valine per gram of glucose. These results suggest that an industrially competitive strain can be efficiently developed by metabolic engineering based on combined rational modification, transcriptome profiling, and systems-level in silico analysis. PMID:17463081

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling.

PubMed

Puente-Marin, Sara; Nombela, Iván; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio; Ortega-Villaizan, María Del Mar

2018-04-09

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation.

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling

PubMed Central

Puente-Marin, Sara; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio

2018-01-01

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation. PMID:29642539

Transcriptomic analysis of grain amaranth (Amaranthus hypochondriacus) using 454 pyrosequencing: comparison with A. tuberculatus, expression profiling in stems and in response to biotic and abiotic stress

PubMed Central

2011-01-01

Background Amaranthus hypochondriacus, a grain amaranth, is a C4 plant noted by its ability to tolerate stressful conditions and produce highly nutritious seeds. These possess an optimal amino acid balance and constitute a rich source of health-promoting peptides. Although several recent studies, mostly involving subtractive hybridization strategies, have contributed to increase the relatively low number of grain amaranth expressed sequence tags (ESTs), transcriptomic information of this species remains limited, particularly regarding tissue-specific and biotic stress-related genes. Thus, a large scale transcriptome analysis was performed to generate stem- and (a)biotic stress-responsive gene expression profiles in grain amaranth. Results A total of 2,700,168 raw reads were obtained from six 454 pyrosequencing runs, which were assembled into 21,207 high quality sequences (20,408 isotigs + 799 contigs). The average sequence length was 1,064 bp and 930 bp for isotigs and contigs, respectively. Only 5,113 singletons were recovered after quality control. Contigs/isotigs were further incorporated into 15,667 isogroups. All unique sequences were queried against the nr, TAIR, UniRef100, UniRef50 and Amaranthaceae EST databases for annotation. Functional GO annotation was performed with all contigs/isotigs that produced significant hits with the TAIR database. Only 8,260 sequences were found to be homologous when the transcriptomes of A. tuberculatus and A. hypochondriacus were compared, most of which were associated with basic house-keeping processes. Digital expression analysis identified 1,971 differentially expressed genes in response to at least one of four stress treatments tested. These included several multiple-stress-inducible genes that could represent potential candidates for use in the engineering of stress-resistant plants. The transcriptomic data generated from pigmented stems shared similarity with findings reported in developing stems of Arabidopsis and black cottonwood (Populus trichocarpa). Conclusions This study represents the first large-scale transcriptomic analysis of A. hypochondriacus, considered to be a highly nutritious and stress-tolerant crop. Numerous genes were found to be induced in response to (a)biotic stress, many of which could further the understanding of the mechanisms that contribute to multiple stress-resistance in plants, a trait that has potential biotechnological applications in agriculture. PMID:21752295

Understanding developmental and adaptive cues in pine through metabolite profiling and co-expression network analysis

PubMed Central

Cañas, Rafael A.; Canales, Javier; Muñoz-Hernández, Carmen; Granados, Jose M.; Ávila, Concepción; García-Martín, María L.; Cánovas, Francisco M.

2015-01-01

Conifers include long-lived evergreen trees of great economic and ecological importance, including pines and spruces. During their long lives conifers must respond to seasonal environmental changes, adapt to unpredictable environmental stresses, and co-ordinate their adaptive adjustments with internal developmental programmes. To gain insights into these responses, we examined metabolite and transcriptomic profiles of needles from naturally growing 25-year-old maritime pine (Pinus pinaster L. Aiton) trees over a year. The effect of environmental parameters such as temperature and rain on needle development were studied. Our results show that seasonal changes in the metabolite profiles were mainly affected by the needles’ age and acclimation for winter, but changes in transcript profiles were mainly dependent on climatic factors. The relative abundance of most transcripts correlated well with temperature, particularly for genes involved in photosynthesis or winter acclimation. Gene network analysis revealed relationships between 14 co-expressed gene modules and development and adaptation to environmental stimuli. Novel Myb transcription factors were identified as candidate regulators during needle development. Our systems-based analysis provides integrated data of the seasonal regulation of maritime pine growth, opening new perspectives for understanding the complex regulatory mechanisms underlying conifers’ adaptive responses. Taken together, our results suggest that the environment regulates the transcriptome for fine tuning of the metabolome during development. PMID:25873654

Transcription Profiling Analysis of Mango–Fusarium mangiferae Interaction

PubMed Central

Liu, Feng; Wu, Jing-bo; Zhan, Ru-lin; Ou, Xiong-chang

2016-01-01

Malformation caused by Fusarium mangiferae is one of the most destructive mango diseases affecting the canopy and floral development, leading to dramatic reduction in fruit yield. To further understand the mechanism of interaction between mango and F. mangiferae, we monitored the transcriptome profiles of buds from susceptible mango plants, which were challenged with F. mangiferae. More than 99 million reads were deduced by RNA-sequencing and were assembled into 121,267 unigenes. Based on the sequence similarity searches, 61,706 unigenes were identified, of which 21,273 and 50,410 were assigned to gene ontology categories and clusters of orthologous groups, respectively, and 33,243 were mapped to 119 KEGG pathways. The differentially expressed genes of mango were detected, having 15,830, 26,061, and 20,146 DEGs respectively, after infection for 45, 75, and 120 days. The analysis of the comparative transcriptome suggests that basic defense mechanisms play important roles in disease resistance. The data also show the transcriptional responses of interactions between mango and the pathogen and more drastic changes in the host transcriptome in response to the pathogen. These results could be used to develop new methods to broaden the resistance of mango to malformation, including the over-expression of key mango genes. PMID:27683574

Whole transcriptome RNA-Seq analysis of breast cancer recurrence risk using formalin-fixed paraffin-embedded tumor tissue.

PubMed

Sinicropi, Dominick; Qu, Kunbin; Collin, Francois; Crager, Michael; Liu, Mei-Lan; Pelham, Robert J; Pho, Mylan; Dei Rossi, Andrew; Jeong, Jennie; Scott, Aaron; Ambannavar, Ranjana; Zheng, Christina; Mena, Raul; Esteban, Jose; Stephans, James; Morlan, John; Baker, Joffre

2012-01-01

RNA biomarkers discovered by RT-PCR-based gene expression profiling of archival formalin-fixed paraffin-embedded (FFPE) tissue form the basis for widely used clinical diagnostic tests; however, RT-PCR is practically constrained in the number of transcripts that can be interrogated. We have developed and optimized RNA-Seq library chemistry as well as bioinformatics and biostatistical methods for whole transcriptome profiling from FFPE tissue. The chemistry accommodates low RNA inputs and sample multiplexing. These methods both enable rediscovery of RNA biomarkers for disease recurrence risk that were previously identified by RT-PCR analysis of a cohort of 136 patients, and also identify a high percentage of recurrence risk markers that were previously discovered using DNA microarrays in a separate cohort of patients, evidence that this RNA-Seq technology has sufficient precision and sensitivity for biomarker discovery. More than two thousand RNAs are strongly associated with breast cancer recurrence risk in the 136 patient cohort (FDR <10%). Many of these are intronic RNAs for which corresponding exons are not also associated with disease recurrence. A number of the RNAs associated with recurrence risk belong to novel RNA networks. It will be important to test the validity of these novel associations in whole transcriptome RNA-Seq screens of other breast cancer cohorts.

Whole Transcriptome RNA-Seq Analysis of Breast Cancer Recurrence Risk Using Formalin-Fixed Paraffin-Embedded Tumor Tissue

PubMed Central

Sinicropi, Dominick; Qu, Kunbin; Collin, Francois; Crager, Michael; Liu, Mei-Lan; Pelham, Robert J.; Pho, Mylan; Rossi, Andrew Dei; Jeong, Jennie; Scott, Aaron; Ambannavar, Ranjana; Zheng, Christina; Mena, Raul; Esteban, Jose; Stephans, James; Morlan, John; Baker, Joffre

2012-01-01

RNA biomarkers discovered by RT-PCR-based gene expression profiling of archival formalin-fixed paraffin-embedded (FFPE) tissue form the basis for widely used clinical diagnostic tests; however, RT-PCR is practically constrained in the number of transcripts that can be interrogated. We have developed and optimized RNA-Seq library chemistry as well as bioinformatics and biostatistical methods for whole transcriptome profiling from FFPE tissue. The chemistry accommodates low RNA inputs and sample multiplexing. These methods both enable rediscovery of RNA biomarkers for disease recurrence risk that were previously identified by RT-PCR analysis of a cohort of 136 patients, and also identify a high percentage of recurrence risk markers that were previously discovered using DNA microarrays in a separate cohort of patients, evidence that this RNA-Seq technology has sufficient precision and sensitivity for biomarker discovery. More than two thousand RNAs are strongly associated with breast cancer recurrence risk in the 136 patient cohort (FDR <10%). Many of these are intronic RNAs for which corresponding exons are not also associated with disease recurrence. A number of the RNAs associated with recurrence risk belong to novel RNA networks. It will be important to test the validity of these novel associations in whole transcriptome RNA-Seq screens of other breast cancer cohorts. PMID:22808097

Transcriptome Profiling of Shewanella oneidensis Gene Expression following Exposure to Acidic and Alkaline pH†

PubMed Central

Leaphart, Adam B.; Thompson, Dorothea K.; Huang, Katherine; Alm, Eric; Wan, Xiu-Feng; Arkin, Adam; Brown, Steven D.; Wu, Liyou; Yan, Tingfen; Liu, Xueduan; Wickham, Gene S.; Zhou, Jizhong

2006-01-01

The molecular response of Shewanella oneidensis MR-1 to variations in extracellular pH was investigated based on genomewide gene expression profiling. Microarray analysis revealed that cells elicited both general and specific transcriptome responses when challenged with environmental acid (pH 4) or base (pH 10) conditions over a 60-min period. Global responses included the differential expression of genes functionally linked to amino acid metabolism, transcriptional regulation and signal transduction, transport, cell membrane structure, and oxidative stress protection. Response to acid stress included the elevated expression of genes encoding glycogen biosynthetic enzymes, phosphate transporters, and the RNA polymerase sigma-38 factor (rpoS), whereas the molecular response to alkaline pH was characterized by upregulation of nhaA and nhaR, which are predicted to encode an Na+/H+ antiporter and transcriptional activator, respectively, as well as sulfate transport and sulfur metabolism genes. Collectively, these results suggest that S. oneidensis modulates multiple transporters, cell envelope components, and pathways of amino acid consumption and central intermediary metabolism as part of its transcriptome response to changing external pH conditions. PMID:16452448

Impact of Transcriptomics on Our Understanding of Pulmonary Fibrosis

PubMed Central

Vukmirovic, Milica; Kaminski, Naftali

2018-01-01

Idiopathic pulmonary fibrosis (IPF) is a lethal fibrotic lung disease characterized by aberrant remodeling of the lung parenchyma with extensive changes to the phenotypes of all lung resident cells. The introduction of transcriptomics, genome scale profiling of thousands of RNA transcripts, caused a significant inversion in IPF research. Instead of generating hypotheses based on animal models of disease, or biological plausibility, with limited validation in humans, investigators were able to generate hypotheses based on unbiased molecular analysis of human samples and then use animal models of disease to test their hypotheses. In this review, we describe the insights made from transcriptomic analysis of human IPF samples. We describe how transcriptomic studies led to identification of novel genes and pathways involved in the human IPF lung such as: matrix metalloproteinases, WNT pathway, epithelial genes, role of microRNAs among others, as well as conceptual insights such as the involvement of developmental pathways and deep shifts in epithelial and fibroblast phenotypes. The impact of lung and transcriptomic studies on disease classification, endotype discovery, and reproducible biomarkers is also described in detail. Despite these impressive achievements, the impact of transcriptomic studies has been limited because they analyzed bulk tissue and did not address the cellular and spatial heterogeneity of the IPF lung. We discuss new emerging technologies and applications, such as single-cell RNAseq and microenvironment analysis that may address cellular and spatial heterogeneity. We end by making the point that most current tissue collections and resources are not amenable to analysis using the novel technologies. To take advantage of the new opportunities, we need new efforts of sample collections, this time focused on access to all the microenvironments and cells in the IPF lung. PMID:29670881

Unique Transcriptome Patterns of the White and Grey Matter Corroborate Structural and Functional Heterogeneity in the Human Frontal Lobe

PubMed Central

Mills, James D.; Kavanagh, Tomas; Kim, Woojin S.; Chen, Bei Jun; Kawahara, Yoshihiro; Halliday, Glenda M.; Janitz, Michael

2013-01-01

The human frontal lobe has undergone accelerated evolution, leading to the development of unique human features such as language and self-reflection. Cortical grey matter and underlying white matter reflect distinct cellular compositions in the frontal lobe. Surprisingly little is known about the transcriptomal landscape of these distinct regions. Here, for the first time, we report a detailed transcriptomal profile of the frontal grey (GM) and white matter (WM) with resolution to alternatively spliced isoforms obtained using the RNA-Seq approach. We observed more vigorous transcriptome activity in GM compared to WM, presumably because of the presence of cellular bodies of neurons in the GM and RNA associated with the nucleus and perinuclear space. Among the top differentially expressed genes, we also identified a number of long intergenic non-coding RNAs (lincRNAs), specifically expressed in white matter, such as LINC00162. Furthermore, along with confirmation of expression of known markers for neurons and oligodendrocytes, we identified a number of genes and splicing isoforms that are exclusively expressed in GM or WM with examples of GABRB2 and PAK2 transcripts, respectively. Pathway analysis identified distinct physiological and biochemical processes specific to grey and white matter samples with a prevalence of synaptic processes in GM and myelination regulation and axonogenesis in the WM. Our study also revealed that expression of many genes, for example, the GPR123, is characterized by isoform switching, depending in which structure the gene is expressed. Our report clearly shows that GM and WM have perhaps surprisingly divergent transcriptome profiles, reflecting distinct roles in brain physiology. Further, this study provides the first reference data set for a normal human frontal lobe, which will be useful in comparative transcriptome studies of cerebral disorders, in particular, neurodegenerative diseases. PMID:24194939

Deep RNA sequencing reveals dynamic regulation of myocardial noncoding RNAs in failing human heart and remodeling with mechanical circulatory support.

PubMed

Yang, Kai-Chien; Yamada, Kathryn A; Patel, Akshar Y; Topkara, Veli K; George, Isaac; Cheema, Faisal H; Ewald, Gregory A; Mann, Douglas L; Nerbonne, Jeanne M

2014-03-04

Microarrays have been used extensively to profile transcriptome remodeling in failing human heart, although the genomic coverage provided is limited and fails to provide a detailed picture of the myocardial transcriptome landscape. Here, we describe sequencing-based transcriptome profiling, providing comprehensive analysis of myocardial mRNA, microRNA (miRNA), and long noncoding RNA (lncRNA) expression in failing human heart before and after mechanical support with a left ventricular (LV) assist device (LVAD). Deep sequencing of RNA isolated from paired nonischemic (NICM; n=8) and ischemic (ICM; n=8) human failing LV samples collected before and after LVAD and from nonfailing human LV (n=8) was conducted. These analyses revealed high abundance of mRNA (37%) and lncRNA (71%) of mitochondrial origin. miRNASeq revealed 160 and 147 differentially expressed miRNAs in ICM and NICM, respectively, compared with nonfailing LV. Among these, only 2 (ICM) and 5 (NICM) miRNAs are normalized with LVAD. RNASeq detected 18 480, including 113 novel, lncRNAs in human LV. Among the 679 (ICM) and 570 (NICM) lncRNAs differentially expressed with heart failure, ≈10% are improved or normalized with LVAD. In addition, the expression signature of lncRNAs, but not miRNAs or mRNAs, distinguishes ICM from NICM. Further analysis suggests that cis-gene regulation represents a major mechanism of action of human cardiac lncRNAs. The myocardial transcriptome is dynamically regulated in advanced heart failure and after LVAD support. The expression profiles of lncRNAs, but not mRNAs or miRNAs, can discriminate failing hearts of different pathologies and are markedly altered in response to LVAD support. These results suggest an important role for lncRNAs in the pathogenesis of heart failure and in reverse remodeling observed with mechanical support.

Hepatic transcriptome profiles differ among maturing beef heifers supplemented with inorganic, organic, or mixed (50% inorganic:50% organic) forms of dietary selenium.

PubMed

Matthews, James C; Zhang, Zhi; Patterson, Jennifer D; Bridges, Phillip J; Stromberg, Arnold J; Boling, J A

2014-09-01

Selenium (Se) is an important trace mineral that, due to deficiencies in the soil in many parts of the USA, must be supplemented directly to the diet of foraging cattle. Both organic and inorganic forms of dietary Se supplements are available and commonly used, and it is known that Se form affects tissue assimilation, bioavailability, and physiological responses. However, little is known about the effects of form of dietary Se supplements on gene expression profiles, which ostensibly account for Se form-dependent physiological processes. To determine if hepatic transcriptomes of growing beef (Angus-cross) heifers (0.5 kg gain/day) was altered by form of dietary supplemental Se, none (Control), or 3 mg Se/day as inorganic Se (ISe, sodium selenite), organic (OSe, Sel-Plex®), or a blend of ISe and OSe (1.5 mg:1.5 mg, Mix) Se was fed for 168 days, and the RNA expression profiles from biopsied liver tissues was compared by microarray analysis. The relative abundance of 139 RNA transcripts was affected by Se treatment, with 86 of these with complete gene annotations. Statistical and bioinformatic analysis of the annotated RNA transcripts revealed clear differences among the four Se treatment groups in their hepatic expression profiles, including (1) solely and commonly affected transcripts; (2) Control and OSe profiles being more similar than Mix and ISe treatments; (3) distinct OSe-, Mix-, and ISe-Se treatment-induced "phenotypes" that possessed both common and unique predicted physiological capacities; and (4) expression of three microRNAs were uniquely sensitive to OSe, ISe, or Mix treatments, including increased capacity for redox potential induced by OSe and Mix Se treatments resulting from decreased expression of MiR2300b messenger RNA. These findings indicate that the form of supplemental dietary Se consumed by cattle will affect the composition of liver transcriptomes resulting, presumably, in different physiological capacities.

A regulation probability model-based meta-analysis of multiple transcriptomics data sets for cancer biomarker identification.

PubMed

Xie, Xin-Ping; Xie, Yu-Feng; Wang, Hong-Qiang

2017-08-23

Large-scale accumulation of omics data poses a pressing challenge of integrative analysis of multiple data sets in bioinformatics. An open question of such integrative analysis is how to pinpoint consistent but subtle gene activity patterns across studies. Study heterogeneity needs to be addressed carefully for this goal. This paper proposes a regulation probability model-based meta-analysis, jGRP, for identifying differentially expressed genes (DEGs). The method integrates multiple transcriptomics data sets in a gene regulatory space instead of in a gene expression space, which makes it easy to capture and manage data heterogeneity across studies from different laboratories or platforms. Specifically, we transform gene expression profiles into a united gene regulation profile across studies by mathematically defining two gene regulation events between two conditions and estimating their occurring probabilities in a sample. Finally, a novel differential expression statistic is established based on the gene regulation profiles, realizing accurate and flexible identification of DEGs in gene regulation space. We evaluated the proposed method on simulation data and real-world cancer datasets and showed the effectiveness and efficiency of jGRP in identifying DEGs identification in the context of meta-analysis. Data heterogeneity largely influences the performance of meta-analysis of DEGs identification. Existing different meta-analysis methods were revealed to exhibit very different degrees of sensitivity to study heterogeneity. The proposed method, jGRP, can be a standalone tool due to its united framework and controllable way to deal with study heterogeneity.

A high-throughput approach to profile RNA structure.

PubMed

Delli Ponti, Riccardo; Marti, Stefanie; Armaos, Alexandros; Tartaglia, Gian Gaetano

2017-03-17

Here we introduce the Computational Recognition of Secondary Structure (CROSS) method to calculate the structural profile of an RNA sequence (single- or double-stranded state) at single-nucleotide resolution and without sequence length restrictions. We trained CROSS using data from high-throughput experiments such as Selective 2΄-Hydroxyl Acylation analyzed by Primer Extension (SHAPE; Mouse and HIV transcriptomes) and Parallel Analysis of RNA Structure (PARS; Human and Yeast transcriptomes) as well as high-quality NMR/X-ray structures (PDB database). The algorithm uses primary structure information alone to predict experimental structural profiles with >80% accuracy, showing high performances on large RNAs such as Xist (17 900 nucleotides; Area Under the ROC Curve AUC of 0.75 on dimethyl sulfate (DMS) experiments). We integrated CROSS in thermodynamics-based methods to predict secondary structure and observed an increase in their predictive power by up to 30%. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Transcriptome profiling in engrailed-2 mutant mice reveals common molecular pathways associated with autism spectrum disorders.

PubMed

Sgadò, Paola; Provenzano, Giovanni; Dassi, Erik; Adami, Valentina; Zunino, Giulia; Genovesi, Sacha; Casarosa, Simona; Bozzi, Yuri

2013-12-19

Transcriptome analysis has been used in autism spectrum disorder (ASD) to unravel common pathogenic pathways based on the assumption that distinct rare genetic variants or epigenetic modifications affect common biological pathways. To unravel recurrent ASD-related neuropathological mechanisms, we took advantage of the En2-/- mouse model and performed transcriptome profiling on cerebellar and hippocampal adult tissues. Cerebellar and hippocampal tissue samples from three En2-/- and wild type (WT) littermate mice were assessed for differential gene expression using microarray hybridization followed by RankProd analysis. To identify functional categories overrepresented in the differentially expressed genes, we used integrated gene-network analysis, gene ontology enrichment and mouse phenotype ontology analysis. Furthermore, we performed direct enrichment analysis of ASD-associated genes from the SFARI repository in our differentially expressed genes. Given the limited number of animals used in the study, we used permissive criteria and identified 842 differentially expressed genes in En2-/- cerebellum and 862 in the En2-/- hippocampus. Our functional analysis revealed that the molecular signature of En2-/- cerebellum and hippocampus shares convergent pathological pathways with ASD, including abnormal synaptic transmission, altered developmental processes and increased immune response. Furthermore, when directly compared to the repository of the SFARI database, our differentially expressed genes in the hippocampus showed enrichment of ASD-associated genes significantly higher than previously reported. qPCR was performed for representative genes to confirm relative transcript levels compared to those detected in microarrays. Despite the limited number of animals used in the study, our bioinformatic analysis indicates the En2-/- mouse is a valuable tool for investigating molecular alterations related to ASD.

Transcriptome display during tilapia sex determination and differentiation as revealed by RNA-Seq analysis.

PubMed

Tao, Wenjing; Chen, Jinlin; Tan, Dejie; Yang, Jing; Sun, Lina; Wei, Jing; Conte, Matthew A; Kocher, Thomas D; Wang, Deshou

2018-05-15

The factors determining sex in teleosts are diverse. Great efforts have been made to characterize the underlying genetic network in various species. However, only seven master sex-determining genes have been identified in teleosts. While the function of a few genes involved in sex determination and differentiation has been studied, we are far from fully understanding how genes interact to coordinate in this process. To enable systematic insights into fish sexual differentiation, we generated a dynamic co-expression network from tilapia gonadal transcriptomes at 5, 20, 30, 40, 90, and 180 dah (days after hatching), plus 45 and 90 dat (days after treatment) and linked gene expression profiles to both development and sexual differentiation. Transcriptomic profiles of female and male gonads at 5 and 20 dah exhibited high similarities except for a small number of genes that were involved in sex determination, while drastic changes were observed from 90 to 180 dah, with a group of differently expressed genes which were involved in gonadal differentiation and gametogenesis. Weighted gene correlation network analysis identified changes in the expression of Borealin, Gtsf1, tesk1, Zar1, Cdn15, and Rpl that were correlated with the expression of genes previously known to be involved in sex differentiation, such as Foxl2, Cyp19a1a, Gsdf, Dmrt1, and Amh. Global gonadal gene expression kinetics during sex determination and differentiation have been extensively profiled in tilapia. These findings provide insights into the genetic framework underlying sex determination and sexual differentiation, and expand our current understanding of developmental pathways during teleost sex determination.

Transcriptional Profiling of Mycobacterium tuberculosis Exposed to In Vitro Lysosomal Stress

PubMed Central

Lin, Wenwei; de Sessions, Paola Florez; Teoh, Garrett Hor Keong; Mohamed, Ahmad Naim Nazri; Zhu, Yuan O.; Koh, Vanessa Hui Qi; Ang, Michelle Lay Teng; Dedon, Peter C.; Hibberd, Martin Lloyd

2016-01-01

Increasing experimental evidence supports the idea that Mycobacterium tuberculosis has evolved strategies to survive within lysosomes of activated macrophages. To further our knowledge of M. tuberculosis response to the hostile lysosomal environment, we profiled the global transcriptional activity of M. tuberculosis when exposed to the lysosomal soluble fraction (SF) prepared from activated macrophages. Transcriptome sequencing (RNA-seq) analysis was performed using various incubation conditions, ranging from noninhibitory to cidal based on the mycobacterial replication or killing profile. Under inhibitory conditions that led to the absence of apparent mycobacterial replication, M. tuberculosis expressed a unique transcriptome with modulation of genes involved in general stress response, metabolic reprogramming, respiration, oxidative stress, dormancy response, and virulence. The transcription pattern also indicates characteristic cell wall remodeling with the possible outcomes of increased infectivity, intrinsic resistance to antibiotics, and subversion of the host immune system. Among the lysosome-specific responses, we identified the glgE-mediated 1,4 α-glucan synthesis pathway and a defined group of VapBC toxin/anti-toxin systems, both of which represent toxicity mechanisms that potentially can be exploited for killing intracellular mycobacteria. A meta-analysis including previously reported transcriptomic studies in macrophage infection and in vitro stress models was conducted to identify overlapping and nonoverlapping pathways. Finally, the Tap efflux pump-encoding gene Rv1258c was selected for validation. An M. tuberculosis ΔRv1258c mutant was constructed and displayed increased susceptibility to killing by lysosomal SF and the antimicrobial peptide LL-37, as well as attenuated survival in primary murine macrophages and human macrophage cell line THP-1. PMID:27324481

A high carbohydrate diet coordinately alters transcriptomic profiles in the adipose tissue leading to enhanced lipid biosynthesis

USDA-ARS?s Scientific Manuscript database

To assess the role of dietary macronutrient composition on adipose gene expression we evaluated changes in transcriptomic profiles in the WAT of rats following high carbohydrate (HC) diets. Female Sprague-Dawley rats received liquid diets at 187 or 220 kcal/kg3/4/d via intragastric infusion. Diets w...

Transcriptome and Gene Expression Analysis of the Rice Leaf Folder, Cnaphalocrosis medinalis

PubMed Central

Li, Shang-Wei; Yang, Hong; Liu, Yue-Feng; Liao, Qi-Rong; Du, Juan; Jin, Dao-Chao

2012-01-01

Background The rice leaf folder (RLF), Cnaphalocrocis medinalis (Guenee) (Lepidoptera: Pyralidae), is one of the most destructive pests affecting rice in Asia. Although several studies have been performed on the ecological and physiological aspects of this species, the molecular mechanisms underlying its developmental regulation, behavior, and insecticide resistance remain largely unknown. Presently, there is a lack of genomic information for RLF; therefore, studies aimed at profiling the RLF transcriptome expression would provide a better understanding of its biological function at the molecular level. Principal Findings De novo assembly of the RLF transcriptome was performed via the short read sequencing technology (Illumina). In a single run, we produced more than 23 million sequencing reads that were assembled into 44,941 unigenes (mean size = 474 bp) by Trinity. Through a similarity search, 25,281 (56.82%) unigenes matched known proteins in the NCBI Nr protein database. The transcriptome sequences were annotated with gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG orthology (KO). Additionally, we profiled gene expression during RLF development using a tag-based digital gene expression (DGE) system. Five DGE libraries were constructed, and variations in gene expression were compared between collected samples: eggs vs. 3rd instar larvae, 3rd instar larvae vs. pupae, pupae vs. adults. The results demonstrated that thousands of genes were significantly differentially expressed during various developmental stages. A number of the differentially expressed genes were confirmed by quantitative real-time PCR (qRT-PCR). Conclusions The RLF transcriptome and DGE data provide a comprehensive and global gene expression profile that would further promote our understanding of the molecular mechanisms underlying various biological characteristics, including development, elevated fecundity, flight, sex differentiation, olfactory behavior, and insecticide resistance in RLF. Therefore, these findings could help elucidate the intrinsic factors involved in the RLF-mediated destruction of rice and offer sustainable insect pest management. PMID:23185238

Deep sequencing reveals cell-type-specific patterns of single-cell transcriptome variation.

PubMed

Dueck, Hannah; Khaladkar, Mugdha; Kim, Tae Kyung; Spaethling, Jennifer M; Francis, Chantal; Suresh, Sangita; Fisher, Stephen A; Seale, Patrick; Beck, Sheryl G; Bartfai, Tamas; Kuhn, Bernhard; Eberwine, James; Kim, Junhyong

2015-06-09

Differentiation of metazoan cells requires execution of different gene expression programs but recent single-cell transcriptome profiling has revealed considerable variation within cells of seeming identical phenotype. This brings into question the relationship between transcriptome states and cell phenotypes. Additionally, single-cell transcriptomics presents unique analysis challenges that need to be addressed to answer this question. We present high quality deep read-depth single-cell RNA sequencing for 91 cells from five mouse tissues and 18 cells from two rat tissues, along with 30 control samples of bulk RNA diluted to single-cell levels. We find that transcriptomes differ globally across tissues with regard to the number of genes expressed, the average expression patterns, and within-cell-type variation patterns. We develop methods to filter genes for reliable quantification and to calibrate biological variation. All cell types include genes with high variability in expression, in a tissue-specific manner. We also find evidence that single-cell variability of neuronal genes in mice is correlated with that in rats consistent with the hypothesis that levels of variation may be conserved. Single-cell RNA-sequencing data provide a unique view of transcriptome function; however, careful analysis is required in order to use single-cell RNA-sequencing measurements for this purpose. Technical variation must be considered in single-cell RNA-sequencing studies of expression variation. For a subset of genes, biological variability within each cell type appears to be regulated in order to perform dynamic functions, rather than solely molecular noise.

Multiple plant hormones and cell wall metabolism regulate apple fruit maturation patterns and texture attributes

USDA-ARS?s Scientific Manuscript database

Molecular events regulating apple fruit ripening and sensory quality are largely unknown. Such knowledge is essential for genomic-assisted apple breeding and postharvest quality management. In this study, a parallel transcriptome profile analysis, scanning electron microscopic (SEM) examination and...

Genome-wide analysis of miRNA and mRNA transcriptomes during amelogenesis.

PubMed

Yin, Kaifeng; Hacia, Joseph G; Zhong, Zhe; Paine, Michael L

2014-11-19

In the rodent incisor during amelogenesis, as ameloblast cells transition from secretory stage to maturation stage, their morphology and transcriptome profiles change dramatically. Prior whole genome transcriptome analysis has given a broad picture of the molecular activities dominating both stages of amelogenesis, but this type of analysis has not included miRNA transcript profiling. In this study, we set out to document which miRNAs and corresponding target genes change significantly as ameloblasts transition from secretory- to maturation-stage amelogenesis. Total RNA samples from both secretory- and maturation-stage rat enamel organs were subjected to genome-wide miRNA and mRNA transcript profiling. We identified 59 miRNAs that were differentially expressed at the maturation stage relative to the secretory stage of enamel development (False Discovery Rate (FDR)<0.05, fold change (FC)≥1.8). In parallel, transcriptome profiling experiments identified 1,729 mRNA transcripts that were differentially expressed in the maturation stage compared to the secretory stage (FDR<0.05, FC≥1.8). Based on bioinformatics analyses, 5.8% (629 total) of these differentially expressed genes (DEGS) were highlighted as being the potential targets of 59 miRNAs that were differentially expressed in the opposite direction, in the same tissue samples. Although the number of predicted target DEGs was not higher than baseline expectations generated by examination of stably expressed miRNAs, Gene Ontology (GO) analysis showed that these 629 DEGS were enriched for ion transport, pH regulation, calcium handling, endocytotic, and apoptotic activities. Seven differentially expressed miRNAs (miR-21, miR-31, miR-488, miR-153, miR-135b, miR-135a and miR298) in secretory- and/or maturation-stage enamel organs were confirmed by in situ hybridization. Further, we used luciferase reporter assays to provide evidence that two of these differentially expressed miRNAs, miR-153 and miR-31, are potential regulators for their predicated target mRNAs, Lamp1 (miR-153) and Tfrc (miR-31). In conclusion, these data indicate that miRNAs exhibit a dynamic expression pattern during the transition from secretory-stage to maturation-stage tooth enamel formation. Although they represent only one of numerous mechanisms influencing gene activities, miRNAs specific to the maturation stage could be involved in regulating several key processes of enamel maturation by influencing mRNA stability and translation.

20180311 - Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells (SOT)

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

Expression Profiling Smackdown: Human Transcriptome Array HTA 2.0 vs. RNA-Seq

PubMed Central

Palermo, Meghann; Driscoll, Heather; Tighe, Scott; Dragon, Julie; Bond, Jeff; Shukla, Arti; Vangala, Mahesh; Vincent, James; Hunter, Tim

2014-01-01

The advent of both microarray and massively parallel sequencing have revolutionized high-throughput analysis of the human transcriptome. Due to limitations in microarray technology, detecting and quantifying coding transcript isoforms, in addition to non-coding transcripts, has been challenging. As a result, RNA-Seq has been the preferred method for characterizing the full human transcriptome, until now. A new high-resolution array from Affymetrix, GeneChip Human Transcriptome Array 2.0 (HTA 2.0), has been designed to interrogate all transcript isoforms in the human transcriptome with >6 million probes targeting coding transcripts, exon-exon splice junctions, and non-coding transcripts. Here we compare expression results from GeneChip HTA 2.0 and RNA-Seq data using identical RNA extractions from three samples each of healthy human mesothelial cells in culture, LP9-C1, and healthy mesothelial cells treated with asbestos, LP9-A1. For GeneChip HTA 2.0 sample preparation, we chose to compare two target preparation methods, NuGEN Ovation Pico WTA V2 with the Encore Biotin Module versus Affymetrix's GeneChip WT PLUS with the WT Terminal Labeling Kit, on identical RNA extractions from both untreated and treated samples. These same RNA extractions were used for the RNA-Seq library preparation. All analyses were performed in Partek Genomics Suite 6.6. Expression profiles for control and asbestos-treated mesothelial cells prepared with NuGEN versus Affymetrix target preparation methods (GeneChip HTA 2.0) are compared to each other as well as to RNA-Seq results.

Effect of high night temperature on storage lipids and transcriptome changes in developing seeds of oilseed rape.

PubMed

Zhou, Longhua; Yan, Tao; Chen, Xin; Li, Zhilan; Wu, Dezhi; Hua, Shuijin; Jiang, Lixi

2018-03-24

Global warming causes a faster increase of night temperature than of day temperature in tropical and subtropical zones. Little is known about the effect of high night temperature on storage lipids and transcriptome changes in oilseed rape. This study compared the total fatty acids and fatty acid compositions in seeds of two oilseed rape cultivars between high and low night temperatures. Their transcriptome profiles were also analyzed. High night temperature significantly affected the total fatty acids and fatty acid compositions in seeds of both low and high oil content cultivars, namely Jiuer-13 and Zheyou-50, thereby resulting in 18.9% and 13.7% total fatty acid reductions, respectively. In particular, high night temperature decreased the relative proportions of C18:0 and C18:1 but increased the proportions of C18:2 and C18:3 in both cultivars. In-depth analysis of transcriptome profiles revealed that high night temperature up-regulated gibberellin signaling during the night-time. This up-regulation was associated with the active expression of genes involved in fatty acid catabolism, such as those in β-oxidation and glyoxylate metabolism pathways. Although the effect of temperature on plant lipids has been previously examined, the present study is the first to focus on night temperature and its effect on the fatty acid composition in seeds.

Transcriptome Analysis of Spartina pectinata in Response to Freezing Stress

PubMed Central

Nah, Gyoungju; Lee, Moonsub; Kim, Do-Soon; Rayburn, A. Lane; Voigt, Thomas; Lee, D. K.

2016-01-01

Prairie cordgrass (Spartina pectinata), a perennial C4 grass native to the North American prairie, has several distinctive characteristics that potentially make it a model crop for production in stressful environments. However, little is known about the transcriptome dynamics of prairie cordgrass despite its unique freezing stress tolerance. Therefore, the purpose of this work was to explore the transcriptome dynamics of prairie cordgrass in response to freezing stress at -5°C for 5 min and 30 min. We used a RNA-sequencing method to assemble the S. pectinata leaf transcriptome and performed gene-expression profiling of the transcripts under freezing treatment. Six differentially expressed gene (DEG) groups were categorized from the profiling. In addition, two major consecutive orders of gene expression were observed in response to freezing; the first being the acute up-regulation of genes involved in plasma membrane modification, calcium-mediated signaling, proteasome-related proteins, and transcription regulators (e.g., MYB and WRKY). The follow-up and second response was of genes involved in encoding the putative anti-freezing protein and the previously known DNA and cell-damage-repair proteins. Moreover, we identified the genes involved in epigenetic regulation and circadian-clock expression. Our results indicate that freezing response in S. pectinata reflects dynamic changes in rapid-time duration, as well as in metabolic, transcriptional, post-translational, and epigenetic regulation. PMID:27032112

The Urinary Bladder Transcriptome and Proteome Defined by Transcriptomics and Antibody-Based Profiling

PubMed Central

Habuka, Masato; Fagerberg, Linn; Hallström, Björn M.; Pontén, Fredrik; Yamamoto, Tadashi; Uhlen, Mathias

2015-01-01

To understand functions and diseases of urinary bladder, it is important to define its molecular constituents and their roles in urinary bladder biology. Here, we performed genome-wide deep RNA sequencing analysis of human urinary bladder samples and identified genes up-regulated in the urinary bladder by comparing the transcriptome data to those of all other major human tissue types. 90 protein-coding genes were elevated in the urinary bladder, either with enhanced expression uniquely in the urinary bladder or elevated expression together with at least one other tissue (group enriched). We further examined the localization of these proteins by immunohistochemistry and tissue microarrays and 20 of these 90 proteins were localized to the whole urothelium with a majority not yet described in the context of the urinary bladder. Four additional proteins were found specifically in the umbrella cells (Uroplakin 1a, 2, 3a, and 3b), and three in the intermediate/basal cells (KRT17, PCP4L1 and ATP1A4). 61 of the 90 elevated genes have not been previously described in the context of urinary bladder and the corresponding proteins are interesting targets for more in-depth studies. In summary, an integrated omics approach using transcriptomics and antibody-based profiling has been used to define a comprehensive list of proteins elevated in the urinary bladder. PMID:26694548

Transcriptomics Profiling of Alzheimer’s Disease Reveal Neurovascular Defects, Altered Amyloid-β Homeostasis, and Deregulated Expression of Long Noncoding RNAs

PubMed Central

Magistri, Marco; Velmeshev, Dmitry; Makhmutova, Madina; Faghihi, Mohammad Ali

2015-01-01

Abstract The underlying genetic variations of late-onset Alzheimer’s disease (LOAD) cases remain largely unknown. A combination of genetic variations with variable penetrance and lifetime epigenetic factors may converge on transcriptomic alterations that drive LOAD pathological process. Transcriptome profiling using deep sequencing technology offers insight into common altered pathways regardless of underpinning genetic or epigenetic factors and thus represents an ideal tool to investigate molecular mechanisms related to the pathophysiology of LOAD. We performed directional RNA sequencing on high quality RNA samples extracted from hippocampi of LOAD and age-matched controls. We further validated our data using qRT-PCR on a larger set of postmortem brain tissues, confirming downregulation of the gene encoding substance P (TAC1) and upregulation of the gene encoding the plasminogen activator inhibitor-1 (SERPINE1). Pathway analysis indicates dysregulation in neural communication, cerebral vasculature, and amyloid-β clearance. Beside protein coding genes, we identified several annotated and non-annotated long noncoding RNAs that are differentially expressed in LOAD brain tissues, three of them are activity-dependent regulated and one is induced by Aβ1 - 42 exposure of human neural cells. Our data provide a comprehensive list of transcriptomics alterations in LOAD hippocampi and warrant holistic approach including both coding and non-coding RNAs in functional studies aimed to understand the pathophysiology of LOAD. PMID:26402107

A generic Transcriptomics Reporting Framework (TRF) for 'omics data processing and analysis.

PubMed

Gant, Timothy W; Sauer, Ursula G; Zhang, Shu-Dong; Chorley, Brian N; Hackermüller, Jörg; Perdichizzi, Stefania; Tollefsen, Knut E; van Ravenzwaay, Ben; Yauk, Carole; Tong, Weida; Poole, Alan

2017-12-01

A generic Transcriptomics Reporting Framework (TRF) is presented that lists parameters that should be reported in 'omics studies used in a regulatory context. The TRF encompasses the processes from transcriptome profiling from data generation to a processed list of differentially expressed genes (DEGs) ready for interpretation. Included within the TRF is a reference baseline analysis (RBA) that encompasses raw data selection; data normalisation; recognition of outliers; and statistical analysis. The TRF itself does not dictate the methodology for data processing, but deals with what should be reported. Its principles are also applicable to sequencing data and other 'omics. In contrast, the RBA specifies a simple data processing and analysis methodology that is designed to provide a comparison point for other approaches and is exemplified here by a case study. By providing transparency on the steps applied during 'omics data processing and analysis, the TRF will increase confidence processing of 'omics data, and regulatory use. Applicability of the TRF is ensured by its simplicity and generality. The TRF can be applied to all types of regulatory 'omics studies, and it can be executed using different commonly available software tools. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

De novo assembly and analysis of the Artemisia argyi transcriptome and identification of genes involved in terpenoid biosynthesis.

PubMed

Liu, Miaomiao; Zhu, Jinhang; Wu, Shengbing; Wang, Chenkai; Guo, Xingyi; Wu, Jiawen; Zhou, Meiqi

2018-04-11

Artemisia argyi Lev. et Vant. (A. argyi) is widely utilized for moxibustion in Chinese medicine, and the mechanism underlying terpenoid biosynthesis in its leaves is suggested to play an important role in its medicinal use. However, the A. argyi transcriptome has not been sequenced. Herein, we performed RNA sequencing for A. argyi leaf, root and stem tissues to identify as many as possible of the transcribed genes. In total, 99,807 unigenes were assembled by analysing the expression profiles generated from the three tissue types, and 67,446 of those unigenes were annotated in public databases. We further performed differential gene expression analysis to compare leaf tissue with the other two tissue types and identified numerous genes that were specifically expressed or up-regulated in leaf tissue. Specifically, we identified multiple genes encoding significant enzymes or transcription factors related to terpenoid synthesis. This study serves as a valuable resource for transcriptome information, as many transcribed genes related to terpenoid biosynthesis were identified in the A. argyi transcriptome, providing a functional genomic basis for additional studies on molecular mechanisms underlying the medicinal use of A. argyi.

Transcriptome Sequencing of Gracilariopsis lemaneiformis to Analyze the Genes Related to Optically Active Phycoerythrin Synthesis.

PubMed

Huang, Xiaoyun; Zang, Xiaonan; Wu, Fei; Jin, Yuming; Wang, Haitao; Liu, Chang; Ding, Yating; He, Bangxiang; Xiao, Dongfang; Song, Xinwei; Liu, Zhu

2017-01-01

Gracilariopsis lemaneiformis (aka Gracilaria lemaneiformis) is a red macroalga rich in phycoerythrin, which can capture light efficiently and transfer it to photosystemⅡ. However, little is known about the synthesis of optically active phycoerythrinin in G. lemaneiformis at the molecular level. With the advent of high-throughput sequencing technology, analysis of genetic information for G. lemaneiformis by transcriptome sequencing is an effective means to get a deeper insight into the molecular mechanism of phycoerythrin synthesis. Illumina technology was employed to sequence the transcriptome of two strains of G. lemaneiformis- the wild type and a green-pigmented mutant. We obtained a total of 86915 assembled unigenes as a reference gene set, and 42884 unigenes were annotated in at least one public database. Taking the above transcriptome sequencing as a reference gene set, 4041 differentially expressed genes were screened to analyze and compare the gene expression profiles of the wild type and green mutant. By GO and KEGG pathway analysis, we concluded that three factors, including a reduction in the expression level of apo-phycoerythrin, an increase of chlorophyll light-harvesting complex synthesis, and reduction of phycoerythrobilin by competitive inhibition, caused the reduction of optically active phycoerythrin in the green-pigmented mutant.

The transcriptional landscape of age in human peripheral blood

PubMed Central

Peters, Marjolein J.; Joehanes, Roby; Pilling, Luke C.; Schurmann, Claudia; Conneely, Karen N.; Powell, Joseph; Reinmaa, Eva; Sutphin, George L.; Zhernakova, Alexandra; Schramm, Katharina; Wilson, Yana A.; Kobes, Sayuko; Tukiainen, Taru; Nalls, Michael A.; Hernandez, Dena G.; Cookson, Mark R.; Gibbs, Raphael J.; Hardy, John; Ramasamy, Adaikalavan; Zonderman, Alan B.; Dillman, Allissa; Traynor, Bryan; Smith, Colin; Longo, Dan L.; Trabzuni, Daniah; Troncoso, Juan; van der Brug, Marcel; Weale, Michael E.; O'Brien, Richard; Johnson, Robert; Walker, Robert; Zielke, Ronald H.; Arepalli, Sampath; Ryten, Mina; Singleton, Andrew B.; Ramos, Yolande F.; Göring, Harald H. H.; Fornage, Myriam; Liu, Yongmei; Gharib, Sina A.; Stranger, Barbara E.; De Jager, Philip L.; Aviv, Abraham; Levy, Daniel; Murabito, Joanne M.; Munson, Peter J.; Huan, Tianxiao; Hofman, Albert; Uitterlinden, André G.; Rivadeneira, Fernando; van Rooij, Jeroen; Stolk, Lisette; Broer, Linda; Verbiest, Michael M. P. J.; Jhamai, Mila; Arp, Pascal; Metspalu, Andres; Tserel, Liina; Milani, Lili; Samani, Nilesh J.; Peterson, Pärt; Kasela, Silva; Codd, Veryan; Peters, Annette; Ward-Caviness, Cavin K.; Herder, Christian; Waldenberger, Melanie; Roden, Michael; Singmann, Paula; Zeilinger, Sonja; Illig, Thomas; Homuth, Georg; Grabe, Hans-Jörgen; Völzke, Henry; Steil, Leif; Kocher, Thomas; Murray, Anna; Melzer, David; Yaghootkar, Hanieh; Bandinelli, Stefania; Moses, Eric K.; Kent, Jack W.; Curran, Joanne E.; Johnson, Matthew P.; Williams-Blangero, Sarah; Westra, Harm-Jan; McRae, Allan F.; Smith, Jennifer A.; Kardia, Sharon L. R.; Hovatta, Iiris; Perola, Markus; Ripatti, Samuli; Salomaa, Veikko; Henders, Anjali K.; Martin, Nicholas G.; Smith, Alicia K.; Mehta, Divya; Binder, Elisabeth B.; Nylocks, K Maria; Kennedy, Elizabeth M.; Klengel, Torsten; Ding, Jingzhong; Suchy-Dicey, Astrid M.; Enquobahrie, Daniel A.; Brody, Jennifer; Rotter, Jerome I.; Chen, Yii-Der I.; Houwing-Duistermaat, Jeanine; Kloppenburg, Margreet; Slagboom, P. Eline; Helmer, Quinta; den Hollander, Wouter; Bean, Shannon; Raj, Towfique; Bakhshi, Noman; Wang, Qiao Ping; Oyston, Lisa J.; Psaty, Bruce M.; Tracy, Russell P.; Montgomery, Grant W.; Turner, Stephen T.; Blangero, John; Meulenbelt, Ingrid; Ressler, Kerry J.; Yang, Jian; Franke, Lude; Kettunen, Johannes; Visscher, Peter M.; Neely, G. Gregory; Korstanje, Ron; Hanson, Robert L.; Prokisch, Holger; Ferrucci, Luigi; Esko, Tonu; Teumer, Alexander; van Meurs, Joyce B. J.; Johnson, Andrew D.

2015-01-01

Disease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication) and identify 1,497 genes that are differentially expressed with chronological age. The age-associated genes do not harbor more age-associated CpG-methylation sites than other genes, but are instead enriched for the presence of potentially functional CpG-methylation sites in enhancer and insulator regions that associate with both chronological age and gene expression levels. We further used the gene expression profiles to calculate the ‘transcriptomic age' of an individual, and show that differences between transcriptomic age and chronological age are associated with biological features linked to ageing, such as blood pressure, cholesterol levels, fasting glucose, and body mass index. The transcriptomic prediction model adds biological relevance and complements existing epigenetic prediction models, and can be used by others to calculate transcriptomic age in external cohorts. PMID:26490707

An evaluation of the basis and consequences of a stay-green mutation in the navel negra (nan) citrus mutant using transcriptomic and proteomic profiling and metabolite analysis

USDA-ARS?s Scientific Manuscript database

A Citrus sinensis spontaneous mutant, navel negra (nan), produces fruit with an abnormal brown colored flavedo during ripening. Analysis of pigment composition in the wild type (WT) and nan flavedo suggested that typical ripening-related chlorophyll (Chl) degradation, but not carotenoid biosynthesis...

Poplar trees reconfigure the transcriptome and metabolome in response to drought in a genotype- and time-of-day-dependent manner.

PubMed

Hamanishi, Erin T; Barchet, Genoa L H; Dauwe, Rebecca; Mansfield, Shawn D; Campbell, Malcolm M

2015-04-21

Drought has a major impact on tree growth and survival. Understanding tree responses to this stress can have important application in both conservation of forest health, and in production forestry. Trees of the genus Populus provide an excellent opportunity to explore the mechanistic underpinnings of forest tree drought responses, given the growing molecular resources that are available for this taxon. Here, foliar tissue of six water-deficit stressed P. balsamifera genotypes was analysed for variation in the metabolome in response to drought and time of day by using an untargeted metabolite profiling technique, gas chromatography/mass-spectrometry (GC/MS). Significant variation in the metabolome was observed in response the imposition of water-deficit stress. Notably, organic acid intermediates such as succinic and malic acid had lower concentrations in leaves exposed to drought, whereas galactinol and raffinose were found in increased concentrations. A number of metabolites with significant difference in accumulation under water-deficit conditions exhibited intraspecific variation in metabolite accumulation. Large magnitude fold-change accumulation was observed in three of the six genotypes. In order to understand the interaction between the transcriptome and metabolome, an integrated analysis of the drought-responsive transcriptome and the metabolome was performed. One P. balsamifera genotype, AP-1006, demonstrated a lack of congruence between the magnitude of the drought transcriptome response and the magnitude of the metabolome response. More specifically, metabolite profiles in AP-1006 demonstrated the smallest changes in response to water-deficit conditions. Pathway analysis of the transcriptome and metabolome revealed specific genotypic responses with respect to primary sugar accumulation, citric acid metabolism, and raffinose family oligosaccharide biosynthesis. The intraspecific variation in the molecular strategies that underpin the responses to drought among genotypes may have an important role in the maintenance of forest health and productivity.

Comparative Analysis of Vertebrate Diurnal/Circadian Transcriptomes

PubMed Central

Boyle, Greg; Richter, Kerstin; Priest, Henry D.; Traver, David; Mockler, Todd C.; Chang, Jeffrey T.; Kay, Steve A.

2017-01-01

From photosynthetic bacteria to mammals, the circadian clock evolved to track diurnal rhythms and enable organisms to anticipate daily recurring changes such as temperature and light. It orchestrates a broad spectrum of physiology such as the sleep/wake and eating/fasting cycles. While we have made tremendous advances in our understanding of the molecular details of the circadian clock mechanism and how it is synchronized with the environment, we still have rudimentary knowledge regarding its connection to help regulate diurnal physiology. One potential reason is the sheer size of the output network. Diurnal/circadian transcriptomic studies are reporting that around 10% of the expressed genome is rhythmically controlled. Zebrafish is an important model system for the study of the core circadian mechanism in vertebrate. As Zebrafish share more than 70% of its genes with human, it could also be an additional model in addition to rodent for exploring the diurnal/circadian output with potential for translational relevance. Here we performed comparative diurnal/circadian transcriptome analysis with established mouse liver and other tissue datasets. First, by combining liver tissue sampling in a 48h time series, transcription profiling using oligonucleotide arrays and bioinformatics analysis, we profiled rhythmic transcripts and identified 2609 rhythmic genes. The comparative analysis revealed interesting features of the output network regarding number of rhythmic genes, proportion of tissue specific genes and the extent of transcription factor family expression. Undoubtedly, the Zebrafish model system will help identify new vertebrate outputs and their regulators and provides leads for further characterization of the diurnal cis-regulatory network. PMID:28076377

Analysis of experience-regulated transcriptome and imprintome during critical periods of mouse visual system development reveals spatiotemporal dynamics.

PubMed

Hsu, Chi-Lin; Chou, Chih-Hsuan; Huang, Shih-Chuan; Lin, Chia-Yi; Lin, Meng-Ying; Tung, Chun-Che; Lin, Chun-Yen; Lai, Ivan Pochou; Zou, Yan-Fang; Youngson, Neil A; Lin, Shau-Ping; Yang, Chang-Hao; Chen, Shih-Kuo; Gau, Susan Shur-Fen; Huang, Hsien-Sung

2018-03-15

Visual system development is light-experience dependent, which strongly implicates epigenetic mechanisms in light-regulated maturation. Among many epigenetic processes, genomic imprinting is an epigenetic mechanism through which monoallelic gene expression occurs in a parent-of-origin-specific manner. It is unknown if genomic imprinting contributes to visual system development. We profiled the transcriptome and imprintome during critical periods of mouse visual system development under normal- and dark-rearing conditions using B6/CAST F1 hybrid mice. We identified experience-regulated, isoform-specific and brain-region-specific imprinted genes. We also found imprinted microRNAs were predominantly clustered into the Dlk1-Dio3 imprinted locus with light experience affecting some imprinted miRNA expression. Our findings provide the first comprehensive analysis of light-experience regulation of the transcriptome and imprintome during critical periods of visual system development. Our results may contribute to therapeutic strategies for visual impairments and circadian rhythm disorders resulting from a dysfunctional imprintome.

Temporal network analysis identifies early physiological and transcriptomic indicators of mild drought in Brassica rapa

PubMed Central

Gehan, Malia A; Mockler, Todd C; Weinig, Cynthia; Ewers, Brent E

2017-01-01

The dynamics of local climates make development of agricultural strategies challenging. Yield improvement has progressed slowly, especially in drought-prone regions where annual crop production suffers from episodic aridity. Underlying drought responses are circadian and diel control of gene expression that regulate daily variations in metabolic and physiological pathways. To identify transcriptomic changes that occur in the crop Brassica rapa during initial perception of drought, we applied a co-expression network approach to associate rhythmic gene expression changes with physiological responses. Coupled analysis of transcriptome and physiological parameters over a two-day time course in control and drought-stressed plants provided temporal resolution necessary for correlation of network modules with dynamic changes in stomatal conductance, photosynthetic rate, and photosystem II efficiency. This approach enabled the identification of drought-responsive genes based on their differential rhythmic expression profiles in well-watered versus droughted networks and provided new insights into the dynamic physiological changes that occur during drought. PMID:28826479

Transcriptomic profiling as a screening tool to detect trenbolone treatment in beef cattle.

PubMed

Pegolo, S; Cannizzo, F T; Biolatti, B; Castagnaro, M; Bargelloni, L

2014-06-01

The effects of steroid hormone implants containing trenbolone alone (Finaplix-H), combined with 17β-oestradiol (17β-E; Revalor-H), or with 17β-E and dexamethasone (Revalor-H plus dexamethasone per os) on the bovine muscle transcriptome were examined by DNA-microarray. Overall, large sets of genes were shown to be modulated by the different growth promoters (GPs) and the regulated pathways and biological processes were mostly shared among the treatment groups. Using the Prediction Analysis of Microarray program, GP-treated animals were accurately identified by a small number of predictive genes. A meta-analysis approach was also carried out for the Revalor group to potentially increase the robustness of class prediction analysis. After data pre-processing, a high level of accuracy (90%) was obtained in the classification of samples, using 105 predictive gene markers. Transcriptomics could thus help in the identification of indirect biomarkers for anabolic treatment in beef cattle to be applied for the screening of muscle samples collected after slaughtering. Copyright © 2014 Elsevier Ltd. All rights reserved.

Time-series analysis of the transcriptome and proteome of Escherichia coli upon glucose repression.

PubMed

Borirak, Orawan; Rolfe, Matthew D; de Koning, Leo J; Hoefsloot, Huub C J; Bekker, Martijn; Dekker, Henk L; Roseboom, Winfried; Green, Jeffrey; de Koster, Chris G; Hellingwerf, Klaas J

2015-10-01

Time-series transcript- and protein-profiles were measured upon initiation of carbon catabolite repression in Escherichia coli, in order to investigate the extent of post-transcriptional control in this prototypical response. A glucose-limited chemostat culture was used as the CCR-free reference condition. Stopping the pump and simultaneously adding a pulse of glucose, that saturated the cells for at least 1h, was used to initiate the glucose response. Samples were collected and subjected to quantitative time-series analysis of both the transcriptome (using microarray analysis) and the proteome (through a combination of 15N-metabolic labeling and mass spectrometry). Changes in the transcriptome and corresponding proteome were analyzed using statistical procedures designed specifically for time-series data. By comparison of the two sets of data, a total of 96 genes were identified that are post-transcriptionally regulated. This gene list provides candidates for future in-depth investigation of the molecular mechanisms involved in post-transcriptional regulation during carbon catabolite repression in E. coli, like the involvement of small RNAs. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Integrated Transcriptomic and Epigenomic Analysis of Primary Human Lung Epithelial Cell Differentiation

PubMed Central

Marconett, Crystal N.; Zhou, Beiyun; Rieger, Megan E.; Selamat, Suhaida A.; Dubourd, Mickael; Fang, Xiaohui; Lynch, Sean K.; Stueve, Theresa Ryan; Siegmund, Kimberly D.; Berman, Benjamin P.

2013-01-01

Elucidation of the epigenetic basis for cell-type specific gene regulation is key to gaining a full understanding of how the distinct phenotypes of differentiated cells are achieved and maintained. Here we examined how epigenetic changes are integrated with transcriptional activation to determine cell phenotype during differentiation. We performed epigenomic profiling in conjunction with transcriptomic profiling using in vitro differentiation of human primary alveolar epithelial cells (AEC). This model recapitulates an in vivo process in which AEC transition from one differentiated cell type to another during regeneration following lung injury. Interrogation of histone marks over time revealed enrichment of specific transcription factor binding motifs within regions of changing chromatin structure. Cross-referencing of these motifs with pathways showing transcriptional changes revealed known regulatory pathways of distal alveolar differentiation, such as the WNT and transforming growth factor beta (TGFB) pathways, and putative novel regulators of adult AEC differentiation including hepatocyte nuclear factor 4 alpha (HNF4A), and the retinoid X receptor (RXR) signaling pathways. Inhibition of the RXR pathway confirmed its functional relevance for alveolar differentiation. Our incorporation of epigenetic data allowed specific identification of transcription factors that are potential direct upstream regulators of the differentiation process, demonstrating the power of this approach. Integration of epigenomic data with transcriptomic profiling has broad application for the identification of regulatory pathways in other models of differentiation. PMID:23818859

Global transcriptome analysis of the C57BL/6J mouse testis by SAGE: evidence for nonrandom gene order.

PubMed

Divina, Petr; Vlcek, Cestmír; Strnad, Petr; Paces, Václav; Forejt, Jirí

2005-03-05

We generated the gene expression profile of the total testis from the adult C57BL/6J male mice using serial analysis of gene expression (SAGE). Two high-quality SAGE libraries containing a total of 76 854 tags were constructed. An extensive bioinformatic analysis and comparison of SAGE transcriptomes of the total testis, testicular somatic cells and other mouse tissues was performed and the theory of male-biased gene accumulation on the X chromosome was tested. We sorted out 829 genes predominantly expressed from the germinal part and 944 genes from the somatic part of the testis. The genes preferentially and specifically expressed in total testis and testicular somatic cells were identified by comparing the testis SAGE transcriptomes to the available transcriptomes of seven non-testis tissues. We uncovered chromosomal clusters of adjacent genes with preferential expression in total testis and testicular somatic cells by a genome-wide search and found that the clusters encompassed a significantly higher number of genes than expected by chance. We observed a significant 3.2-fold enrichment of the proportion of X-linked genes specific for testicular somatic cells, while the proportions of X-linked genes specific for total testis and for other tissues were comparable. In contrast to the tissue-specific genes, an under-representation of X-linked genes in the total testis transcriptome but not in the transcriptomes of testicular somatic cells and other tissues was detected. Our results provide new evidence in favor of the theory of male-biased genes accumulation on the X chromosome in testicular somatic cells and indicate the opposite action of the meiotic X-inactivation in testicular germ cells.

Global transcriptome analysis of the C57BL/6J mouse testis by SAGE: evidence for nonrandom gene order

PubMed Central

Divina, Petr; Vlček, Čestmír; Strnad, Petr; Pačes, Václav; Forejt, Jiří

2005-01-01

Background We generated the gene expression profile of the total testis from the adult C57BL/6J male mice using serial analysis of gene expression (SAGE). Two high-quality SAGE libraries containing a total of 76 854 tags were constructed. An extensive bioinformatic analysis and comparison of SAGE transcriptomes of the total testis, testicular somatic cells and other mouse tissues was performed and the theory of male-biased gene accumulation on the X chromosome was tested. Results We sorted out 829 genes predominantly expressed from the germinal part and 944 genes from the somatic part of the testis. The genes preferentially and specifically expressed in total testis and testicular somatic cells were identified by comparing the testis SAGE transcriptomes to the available transcriptomes of seven non-testis tissues. We uncovered chromosomal clusters of adjacent genes with preferential expression in total testis and testicular somatic cells by a genome-wide search and found that the clusters encompassed a significantly higher number of genes than expected by chance. We observed a significant 3.2-fold enrichment of the proportion of X-linked genes specific for testicular somatic cells, while the proportions of X-linked genes specific for total testis and for other tissues were comparable. In contrast to the tissue-specific genes, an under-representation of X-linked genes in the total testis transcriptome but not in the transcriptomes of testicular somatic cells and other tissues was detected. Conclusion Our results provide new evidence in favor of the theory of male-biased genes accumulation on the X chromosome in testicular somatic cells and indicate the opposite action of the meiotic X-inactivation in testicular germ cells. PMID:15748293

Determining the optimal number of independent components for reproducible transcriptomic data analysis.

PubMed

Kairov, Ulykbek; Cantini, Laura; Greco, Alessandro; Molkenov, Askhat; Czerwinska, Urszula; Barillot, Emmanuel; Zinovyev, Andrei

2017-09-11

Independent Component Analysis (ICA) is a method that models gene expression data as an action of a set of statistically independent hidden factors. The output of ICA depends on a fundamental parameter: the number of components (factors) to compute. The optimal choice of this parameter, related to determining the effective data dimension, remains an open question in the application of blind source separation techniques to transcriptomic data. Here we address the question of optimizing the number of statistically independent components in the analysis of transcriptomic data for reproducibility of the components in multiple runs of ICA (within the same or within varying effective dimensions) and in multiple independent datasets. To this end, we introduce ranking of independent components based on their stability in multiple ICA computation runs and define a distinguished number of components (Most Stable Transcriptome Dimension, MSTD) corresponding to the point of the qualitative change of the stability profile. Based on a large body of data, we demonstrate that a sufficient number of dimensions is required for biological interpretability of the ICA decomposition and that the most stable components with ranks below MSTD have more chances to be reproduced in independent studies compared to the less stable ones. At the same time, we show that a transcriptomics dataset can be reduced to a relatively high number of dimensions without losing the interpretability of ICA, even though higher dimensions give rise to components driven by small gene sets. We suggest a protocol of ICA application to transcriptomics data with a possibility of prioritizing components with respect to their reproducibility that strengthens the biological interpretation. Computing too few components (much less than MSTD) is not optimal for interpretability of the results. The components ranked within MSTD range have more chances to be reproduced in independent studies.

The root transcriptome for North American ginseng assembled and profiled across seasonal development

PubMed Central

2013-01-01

Background Ginseng including North American ginseng (Panax quinquefolius L.) is one of the most widely used medicinal plants. Its success is thought to be due to a diverse collection of ginsenosides that serve as its major bioactive compounds. However, few genomic resources exist and the details concerning its various biosynthetic pathways remain poorly understood. As the root is the primary tissue harvested commercially for ginsenosides, next generation sequencing was applied to the characterization and assembly of the root transcriptome throughout seasonal development. Transcripts showing homology to ginsenoside biosynthesis enzymes were profiled in greater detail. Results RNA extracts from root samples from seven development stages of North American ginseng were subjected to 454 sequencing, filtered for quality and used in the de novo assembly of a collective root reference transcriptome consisting of 41,623 transcripts. Annotation efforts using a number of public databases resulted in detailed annotation information for 34,801 (84%) transcripts. In addition, 3,955 genes were assigned to metabolic pathways using the Kyoto Encyclopedia of Genes and Genomes. Among our results, we found all of the known enzymes involved in the ginsenoside backbone biosynthesis and used co-expression analysis to identify a number of candidate sequences involved in the latter stages ginsenoside biosynthesis pathway. Transcript profiles suggest ginsenoside biosynthesis occurs at distinct stages of development. Conclusions The assembly generated provides a comprehensive annotated reference for future transcriptomic study of North American ginseng. A collection of putative ginsenoside biosynthesis genes were identified and candidate genes predicted from the lesser understood downstream stages of biosynthesis. Transcript expression profiles across seasonal development suggest a primary dammarane-type ginsenoside biosynthesis occurs just prior to plant senescence, with secondary ginsenoside production occurring throughout development. Data from the study provide a valuable resource for conducting future ginsenoside biosynthesis research in this important medicinal plant. PMID:23957709

Comparative Transcriptome Analysis of Cultivated and Wild Watermelon during Fruit Development

PubMed Central

Guo, Shaogui; Sun, Honghe; Zhang, Haiying; Liu, Jingan; Ren, Yi; Gong, Guoyi; Jiao, Chen; Zheng, Yi; Yang, Wencai; Fei, Zhangjun; Xu, Yong

2015-01-01

Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] is an important vegetable crop world-wide. Watermelon fruit quality is a complex trait determined by various factors such as sugar content, flesh color and flesh texture. Fruit quality and developmental process of cultivated and wild watermelon are highly different. To systematically understand the molecular basis of these differences, we compared transcriptome profiles of fruit tissues of cultivated watermelon 97103 and wild watermelon PI296341-FR. We identified 2,452, 826 and 322 differentially expressed genes in cultivated flesh, cultivated mesocarp and wild flesh, respectively, during fruit development. Gene ontology enrichment analysis of these genes indicated that biological processes and metabolic pathways related to fruit quality such as sweetness and flavor were significantly changed only in the flesh of 97103 during fruit development, while those related to abiotic stress response were changed mainly in the flesh of PI296341-FR. Our comparative transcriptome profiling analysis identified critical genes potentially involved in controlling fruit quality traits including α-galactosidase, invertase, UDP-galactose/glucose pyrophosphorylase and sugar transporter genes involved in the determination of fruit sugar content, phytoene synthase, β-carotene hydroxylase, 9-cis-epoxycarotenoid dioxygenase and carotenoid cleavage dioxygenase genes involved in carotenoid metabolism, and 4-coumarate:coenzyme A ligase, cellulose synthase, pectinesterase, pectinesterase inhibitor, polygalacturonase inhibitor and α-mannosidase genes involved in the regulation of flesh texture. In addition, we found that genes in the ethylene biosynthesis and signaling pathway including ACC oxidase, ethylene receptor and ethylene responsive factor showed highly ripening-associated expression patterns, indicating a possible role of ethylene in fruit development and ripening of watermelon, a non-climacteric fruit. Our analysis provides novel insights into watermelon fruit quality and ripening biology. Furthermore, the comparative expression profile data we developed provides a valuable resource to accelerate functional studies in watermelon and facilitate watermelon crop improvement. PMID:26079257

Transcriptome profile and cytogenetic analysis of immortalized neuronally restricted progenitor cells derived from the porcine olfactory bulb

USDA-ARS?s Scientific Manuscript database

Recently, we established and phenotypically characterized an immortalized porcine olfactory bulb neuroblast cell line, OBGF400 (Uebing-Czipura et al., 2008). To facilitate the future application of these cells in studies of neurological dysfunction and neuronal replacement therapies, a comprehensive...

Transcriptomic Analysis of Differentially Expressed Genes During Larval Development of Rapana venosa by Digital Gene Expression Profiling.

PubMed

Song, Hao; Yu, Zheng-Lin; Sun, Li-Na; Xue, Dong-Xiu; Zhang, Tao; Wang, Hai-Yan

2016-07-07

During the life cycle of shellfish, larval development, especially metamorphosis, has a vital influence on the dynamics, distribution, and recruitment of natural populations, as well as seed breeding. Rapana venosa, a carnivorous gastropod, is an important commercial shellfish in China, and is an ecological invader in the United States, Argentina, and France. However, information about the mechanism of its early development is still limited, because research in this area has long suffered from a lack of genomic resources. In this study, 15 digital gene expression (DGE) libraries from five developmental stages of R. venosa were constructed and sequenced on the IIIumina Hi-Sequation 2500 platform. Bioinformaticsanalysis identified numerous differentially and specifically expressed genes, which revealed that genes associated with growth, nervous system, digestive system, immune system, and apoptosis participate in important developmental processes. The functional analysis of differentially expressed genes was further implemented by gene ontology, and Kyoto encyclopedia of genes and genomes enrichment. DGE profiling provided a general picture of the transcriptomic activities during the early development of R. venosa, which may provide interesting hints for further study. Our data represent the first comparative transcriptomic information available for the early development of R. venosa, which is a prerequisite for a better understanding of the physiological traits controlling development. Copyright © 2016 Song et al.

Metabolic modeling helps interpret transcriptomic changes during malaria.

PubMed

Tang, Yan; Gupta, Anuj; Garimalla, Swetha; Galinski, Mary R; Styczynski, Mark P; Fonseca, Luis L; Voit, Eberhard O

2018-06-01

Disease represents a specific case of malfunctioning within a complex system. Whereas it is often feasible to observe and possibly treat the symptoms of a disease, it is much more challenging to identify and characterize its molecular root causes. Even in infectious diseases that are caused by a known parasite, it is often impossible to pinpoint exactly which molecular profiles of components or processes are directly or indirectly altered. However, a deep understanding of such profiles is a prerequisite for rational, efficacious treatments. Modern omics methodologies are permitting large-scale scans of some molecular profiles, but these scans often yield results that are not intuitive and difficult to interpret. For instance, the comparison of healthy and diseased transcriptome profiles may point to certain sets of involved genes, but a host of post-transcriptional processes and regulatory mechanisms renders predictions regarding metabolic or physiological consequences of the observed changes in gene expression unreliable. Here we present proof of concept that dynamic models of metabolic pathway systems may offer a tool for interpreting transcriptomic profiles measured during disease. We illustrate this strategy with the interpretation of expression data of genes coding for enzymes associated with purine metabolism. These data were obtained during infections of rhesus macaques (Macaca mulatta) with the malaria parasite Plasmodium cynomolgi or P. coatneyi. The model-based interpretation reveals clear patterns of flux redistribution within the purine pathway that are consistent between the two malaria pathogens and are even reflected in data from humans infected with P. falciparum. This article is part of a Special Issue entitled: Accelerating Precision Medicine through Genetic and Genomic Big Data Analysis edited by Yudong Cai & Tao Huang. Copyright © 2017 Elsevier B.V. All rights reserved.

Lessons from single-cell transcriptome analysis of oxygen-sensing cells.

PubMed

Zhou, Ting; Matsunami, Hiroaki

2018-05-01

The advent of single-cell RNA-sequencing (RNA-Seq) technology has enabled transcriptome profiling of individual cells. Comprehensive gene expression analysis at the single-cell level has proven to be effective in characterizing the most fundamental aspects of cellular function and identity. This unbiased approach is revolutionary for small and/or heterogeneous tissues like oxygen-sensing cells in identifying key molecules. Here, we review the major methods of current single-cell RNA-Seq technology. We discuss how this technology has advanced the understanding of oxygen-sensing glomus cells in the carotid body and helped uncover novel oxygen-sensing cells and mechanisms in the mice olfactory system. We conclude by providing our perspective on future single-cell RNA-Seq research directed at oxygen-sensing cells.

Transcriptome analysis of genes involved in anthocyanins biosynthesis and transport in berries of black and white spine grapes (Vitis davidii).

PubMed

Sun, Lei; Fan, Xiucai; Zhang, Ying; Jiang, Jianfu; Sun, Haisheng; Liu, Chonghuai

2016-01-01

The color of berry skin is an important economic trait for grape and is essentially determined by the components and content of anthocyanins. The fruit color of Chinese wild grapes is generally black, and the profile of anthocyanins in Chinese wild grapes is significantly different from that of Vitis vinifera . However, V. davidii is the only species that possesses white berry varieties among Chinese wild grape species. Thus, we performed a transcriptomic analysis to compare the difference of transcriptional level in black and white V. davidii , in order to find some key genes that are related to anthocyanins accumulation in V. davidii . The results of anthocyanins detection revealed that 3,5- O -diglucoside anthocyanins is the predominant anthocyanins in V. davidii . It showed obvious differences from V. vinifera in the profile of the composition of anthocyanins. The transcriptome sequencing by Illumina mRNA-Seq technology generated an average of 57 million 100-base pair clean reads from each sample. Differential gene expression analysis revealed thousands of differential expression genes (DEGs) in the pairwise comparison of different fruit developmental stages between and within black and white V. davidii . After the analysis of functional category enrichment and differential expression patterns of DEGs, 46 genes were selected as the candidate genes. Some genes have been reported as being related to anthocyanins accumulation, and some genes were newly found in our study as probably being related to anthocyanins accumulation. We inferred that 3AT (VIT_03s0017g00870) played an important role in anthocyanin acylation, GST4 (VIT_04s0079g00690) and AM2 (VIT_16s0050g00910) played important roles in anthocyanins transport in V. davidii . The expression of some selected DEGs was further confirmed by quantitative real-time PCR (qRT-PCR). The present study investigated the transcriptomic profiles of berry skin from black and white spine grapes at three fruit developmental stages by Illumina mRNA-Seq technology. It revealed the variety specificity of anthocyanins accumulation in V. davidi at the transcriptional level. The data reported here will provide a valuable resource for understanding anthocyanins accumulation in grapes, especially in V. davidii .

RNA-Seq transcriptome profiling of mouse oocytes after in vitro maturation and/or vitrification.

PubMed

Gao, Lei; Jia, Gongxue; Li, Ai; Ma, Haojia; Huang, Zhengyuan; Zhu, Shien; Hou, Yunpeng; Fu, Xiangwei

2017-10-16

In vitro maturation (IVM) and vitrification have been widely used to prepare oocytes before fertilization; however, potential effects of these procedures, such as expression profile changes, are poorly understood. In this study, mouse oocytes were divided into four groups and subjected to combinations of in vitro maturation and/or vitrification treatments. RNA-seq and in silico pathway analysis were used to identify differentially expressed genes (DEGs) that may be involved in oocyte viability after in vitro maturation and/or vitrification. Our results showed that 1) 69 genes were differentially expressed after IVM, 66 of which were up-regulated. Atp5e and Atp5o were enriched in the most significant gene ontology term "mitochondrial membrane part"; thus, these genes may be promising candidate biomarkers for oocyte viability after IVM. 2) The influence of vitrification on the transcriptome of oocytes was negligible, as no DEGs were found between vitrified and fresh oocytes. 3) The MII stage is more suitable for oocyte vitrification with respect to the transcriptome. This study provides a valuable new theoretical basis to further improve the efficiency of in vitro maturation and/or oocyte vitrification.

Transcriptome analysis of Cronobacter sakazakii ATCC BAA-894 after interaction with human intestinal epithelial cell line HCT-8.

PubMed

Jing, Chun-e; Du, Xin-jun; Li, Ping; Wang, Shuo

2016-01-01

Cronobacter spp. are opportunistic pathogens that are responsible for infections including severe meningitis, septicemia, and necrotizing enterocolitis in neonates and infants. To date, questions still remain regarding the mechanisms of pathogenicity and virulence determinants for each bacterial strain. In this study, we established an in vitro model for Cronobacter sakazakii ATCC BAA-894 infection of HCT-8 human colorectal epithelial cells. The transcriptome profile of C. sakazakii ATCC BAA-894 after interaction with HCT-8 cells was determined using high-throughput whole-transcriptome sequencing (RNA sequencing (RNA-seq)). Gene expression profiles indicated that 139 genes were upregulated and 72 genes were downregulated in the adherent C. sakazakii ATCC BAA-894 strain on HCT-8 cells compared to the cultured bacteria in the cell-free medium. Expressions of some flagella genes and virulence factors involved in adherence were upregulated. High osmolarity and osmotic stress-associated genes were highly upregulated, as well as genes responsible for the synthesis of lipopolysaccharides and outer membrane proteins, iron acquisition systems, and glycerol and glycerophospholipid metabolism. In sum, our study provides further insight into the mechanisms underlying C. sakazakii pathogenesis in the human gastrointestinal tract.

Genomic methylation and transcriptomic profiling provides insights into heading depression in inbred Brassica rapa L. ssp. pekinensis.

PubMed

Liu, Yan; Xu, Cui; Tang, Xuebing; Pei, Surui; Jin, Di; Guo, Minghao; Yang, Meng; Zhang, Yaowei

2018-07-30

Inbreeding depression is the reduction in fitness observed in inbred populations. In plants, it leads to disease, weaker resistance to adverse environmental conditions, inhibition of growth, and decrease of yield. To elucidate molecular mechanisms behind inbreeding depression, we compared global DNA methylation and transcriptome profiles of a normal and a highly inbred heading degenerated variety of the Chinese cabbage (Brassica rapa L. ssp. pekinensis). DNA methylation was reduced in inbred plants, suggesting a change in the epigenetic landscape. Transcriptome analysis by RNA-Seq revealed that genes in auxin-response and synthesis pathways were differentially expressed in the inbreeding depression lines. Interestingly, methylation levels of some of those genes were also changed. Furthermore, endogenous IAA content was decreased in inbred plants, in agreement with expression and methylation data. Chemical inhibition of auxin also replicated the degenerated phenotype in normal plants, while exogenous IAA application had no effect in inbred depression plants, suggesting a more complex mechanism. These data indicate DNA methylation-regulated auxin pathways play a role in establishing inbred depression phenotypes in plants. Our findings reveal new insights into inbreeding depression and leafy head development in Chinese cabbage. Copyright © 2018 Elsevier B.V. All rights reserved.

De novo assembly and comparative transcriptome analysis of the foot from Chinese green mussel (Perna viridis) in response to cadmium stimulation

PubMed Central

You, Xinxin; Wang, Jintu; Chen, Jieming; Peng, Chao; Shi, Qiong

2017-01-01

The Chinese green mussel, Perna viridis, is a marine bivalve with important economic values as well as biomonitoring roles for aquatic pollution. Byssus, secreted by the foot gland, has been proved to bind heavy metals effectively. In this study, using the RNA sequencing technology, we performed comparative transcriptomic analysis on the mussel feet with or without inducing by cadmium (Cd). Our current work is aiming at providing insights into the molecular mechanisms of byssus binding to heavy metal ions. The transcriptome sequencing generated a total of 26.13-Gb raw data. After a careful assembly of clean data, we obtained a primary set of 105,127 unigenes, in which 32,268 unigenes were annotated. Based on the expression profiles, we identified 9,048 differentially expressed genes (DEGs) between Cd treatment (50 or 100 μg/L) at 48 h and the control, suggesting an extensive transcriptome response of the mussels during the Cd stimulation. Moreover, we observed that the expression levels of 54 byssus protein coding genes increased significantly after the 48-h Cd stimulation. In addition, 16 critical byssus protein coding genes were picked for profiling by quantitative real-time PCR (qRT-PCR). Finally, we reached a primary conclusion that high content of tyrosine (Tyr), cysteine (Cys), histidine (His) residues or the special motif plays an important role in the accumulation of heavy metals in byssus. We also proposed an interesting model for the confirmed byssal Cd accumulation, in which biosynthesis of byssus proteins may play simultaneously critical roles since their transcription levels were significantly elevated. PMID:28520756

Physiology of Pseudomonas aeruginosa in biofilms as revealed by transcriptome analysis

PubMed Central

2010-01-01

Background Transcriptome analysis was applied to characterize the physiological activities of Pseudomonas aeruginosa grown for three days in drip-flow biofilm reactors. Conventional applications of transcriptional profiling often compare two paired data sets that differ in a single experimentally controlled variable. In contrast this study obtained the transcriptome of a single biofilm state, ranked transcript signals to make the priorities of the population manifest, and compared ranki ngs for a priori identified physiological marker genes between the biofilm and published data sets. Results Biofilms tolerated exposure to antibiotics, harbored steep oxygen concentration gradients, and exhibited stratified and heterogeneous spatial patterns of protein synthetic activity. Transcriptional profiling was performed and the signal intensity of each transcript was ranked to gain insight into the physiological state of the biofilm population. Similar rankings were obtained from data sets published in the GEO database http://www.ncbi.nlm.nih.gov/geo. By comparing the rank of genes selected as markers for particular physiological activities between the biofilm and comparator data sets, it was possible to infer qualitative features of the physiological state of the biofilm bacteria. These biofilms appeared, from their transcriptome, to be glucose nourished, iron replete, oxygen limited, and growing slowly or exhibiting stationary phase character. Genes associated with elaboration of type IV pili were strongly expressed in the biofilm. The biofilm population did not indicate oxidative stress, homoserine lactone mediated quorum sensing, or activation of efflux pumps. Using correlations with transcript ranks, the average specific growth rate of biofilm cells was estimated to be 0.08 h-1. Conclusions Collectively these data underscore the oxygen-limited, slow-growing nature of the biofilm population and are consistent with antimicrobial tolerance due to low metabolic activity. PMID:21083928

A Cross-Species Analysis in Pancreatic Neuroendocrine Tumors Reveals Molecular Subtypes with Distinctive Clinical, Metastatic, Developmental, and Metabolic Characteristics

PubMed Central

Sadanandam, Anguraj; Wullschleger, Stephan; Lyssiotis, Costas A.; Grötzinger, Carsten; Barbi, Stefano; Bersani, Samantha; Körner, Jan; Wafy, Ismael; Mafficini, Andrea; Lawlor, Rita T.; Simbolo, Michele; Asara, John M.; Bläker, Hendrik; Cantley, Lewis C.; Wiedenmann, Bertram; Scarpa, Aldo; Hanahan, Douglas

2016-01-01

Seeking to assess the representative and instructive value of an engineered mouse model of pancreatic neuroendocrine tumors (PanNET) for its cognate human cancer, we profiled and compared mRNA and miRNA transcriptomes of tumors from both. Mouse PanNET tumors could be classified into two distinctive subtypes, well-differentiated islet/insulinoma tumors (IT) and poorly differentiated tumors associated with liver metastases, dubbed metastasis-like primary (MLP). Human PanNETs were independently classified into these same two subtypes, along with a third, specific gene mutation–enriched subtype. The MLP subtypes in human and mouse were similar to liver metastases in terms of miRNA and mRNA transcriptome profiles and signature genes. The human/mouse MLP subtypes also similarly expressed genes known to regulate early pancreas development, whereas the IT subtypes expressed genes characteristic of mature islet cells, suggesting different tumorigenesis pathways. In addition, these subtypes exhibit distinct metabolic profiles marked by differential pyruvate metabolism, substantiating the significance of their separate identities. SIGNIFICANCE This study involves a comprehensive cross-species integrated analysis of multi-omics profiles and histology to stratify PanNETs into subtypes with distinctive characteristics. We provide support for the RIP1-TAG2 mouse model as representative of its cognate human cancer with prospects to better understand PanNET heterogeneity and consider future applications of personalized cancer therapy. PMID:26446169

A Cross-Species Analysis in Pancreatic Neuroendocrine Tumors Reveals Molecular Subtypes with Distinctive Clinical, Metastatic, Developmental, and Metabolic Characteristics.

PubMed

Sadanandam, Anguraj; Wullschleger, Stephan; Lyssiotis, Costas A; Grötzinger, Carsten; Barbi, Stefano; Bersani, Samantha; Körner, Jan; Wafy, Ismael; Mafficini, Andrea; Lawlor, Rita T; Simbolo, Michele; Asara, John M; Bläker, Hendrik; Cantley, Lewis C; Wiedenmann, Bertram; Scarpa, Aldo; Hanahan, Douglas

2015-12-01

Seeking to assess the representative and instructive value of an engineered mouse model of pancreatic neuroendocrine tumors (PanNET) for its cognate human cancer, we profiled and compared mRNA and miRNA transcriptomes of tumors from both. Mouse PanNET tumors could be classified into two distinctive subtypes, well-differentiated islet/insulinoma tumors (IT) and poorly differentiated tumors associated with liver metastases, dubbed metastasis-like primary (MLP). Human PanNETs were independently classified into these same two subtypes, along with a third, specific gene mutation-enriched subtype. The MLP subtypes in human and mouse were similar to liver metastases in terms of miRNA and mRNA transcriptome profiles and signature genes. The human/mouse MLP subtypes also similarly expressed genes known to regulate early pancreas development, whereas the IT subtypes expressed genes characteristic of mature islet cells, suggesting different tumorigenesis pathways. In addition, these subtypes exhibit distinct metabolic profiles marked by differential pyruvate metabolism, substantiating the significance of their separate identities. This study involves a comprehensive cross-species integrated analysis of multi-omics profiles and histology to stratify PanNETs into subtypes with distinctive characteristics. We provide support for the RIP1-TAG2 mouse model as representative of its cognate human cancer with prospects to better understand PanNET heterogeneity and consider future applications of personalized cancer therapy. ©2015 American Association for Cancer Research.

De novo transcriptomic analysis of hydrogen production in the green alga Chlamydomonas moewusii through RNA-Seq

PubMed Central

2013-01-01

Background Microalgae can make a significant contribution towards meeting global renewable energy needs in both carbon-based and hydrogen (H2) biofuel. The development of energy-related products from algae could be accelerated with improvements in systems biology tools, and recent advances in sequencing technology provide a platform for enhanced transcriptomic analyses. However, these techniques are still heavily reliant upon available genomic sequence data. Chlamydomonas moewusii is a unicellular green alga capable of evolving molecular H2 under both dark and light anaerobic conditions, and has high hydrogenase activity that can be rapidly induced. However, to date, there is no systematic investigation of transcriptomic profiling during induction of H2 photoproduction in this organism. Results In this work, RNA-Seq was applied to investigate transcriptomic profiles during the dark anaerobic induction of H2 photoproduction. 156 million reads generated from 7 samples were then used for de novo assembly after data trimming. BlastX results against NCBI database and Blast2GO results were used to interpret the functions of the assembled 34,136 contigs, which were then used as the reference contigs for RNA-Seq analysis. Our results indicated that more contigs were differentially expressed during the period of early and higher H2 photoproduction, and fewer contigs were differentially expressed when H2-photoproduction rates decreased. In addition, C. moewusii and C. reinhardtii share core functional pathways, and transcripts for H2 photoproduction and anaerobic metabolite production were identified in both organisms. C. moewusii also possesses similar metabolic flexibility as C. reinhardtii, and the difference between C. moewusii and C. reinhardtii on hydrogenase expression and anaerobic fermentative pathways involved in redox balancing may explain their different profiles of hydrogenase activity and secreted anaerobic metabolites. Conclusions Herein, we have described a workflow using commercial software to analyze RNA-Seq data without reference genome sequence information, which can be applied to other unsequenced microorganisms. This study provided biological insights into the anaerobic fermentation and H2 photoproduction of C. moewusii, and the first transcriptomic RNA-Seq dataset of C. moewusii generated in this study also offer baseline data for further investigation (e.g. regulatory proteins related to fermentative pathway discussed in this study) of this organism as a H2-photoproduction strain. PMID:23971877

Transcriptome analysis of Petunia axillaris flowers reveals genes involved in morphological differentiation and metabolite transport

PubMed Central

Amano, Ikuko; Kitajima, Sakihito; Suzuki, Hideyuki; Koeduka, Takao

2018-01-01

The biosynthesis of plant secondary metabolites is associated with morphological and metabolic differentiation. As a consequence, gene expression profiles can change drastically, and primary and secondary metabolites, including intermediate and end-products, move dynamically within and between cells. However, little is known about the molecular mechanisms underlying differentiation and transport mechanisms. In this study, we performed a transcriptome analysis of Petunia axillaris subsp. parodii, which produces various volatiles in its corolla limbs and emits metabolites to attract pollinators. RNA-sequencing from leaves, buds, and limbs identified 53,243 unigenes. Analysis of differentially expressed genes, combined with gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses, showed that many biological processes were highly enriched in limbs. These included catabolic processes and signaling pathways of hormones, such as gibberellins, and metabolic pathways, including phenylpropanoids and fatty acids. Moreover, we identified five transporter genes that showed high expression in limbs, and we performed spatiotemporal expression analyses and homology searches to infer their putative functions. Our systematic analysis provides comprehensive transcriptomic information regarding morphological differentiation and metabolite transport in the Petunia flower and lays the foundation for establishing the specific mechanisms that control secondary metabolite biosynthesis in plants. PMID:29902274

Nephron Toxicity Profiling via Untargeted Metabolome Analysis Employing a High Performance Liquid Chromatography-Mass Spectrometry-based Experimental and Computational Pipeline*

PubMed Central

Ranninger, Christina; Rurik, Marc; Limonciel, Alice; Ruzek, Silke; Reischl, Roland; Wilmes, Anja; Jennings, Paul; Hewitt, Philip; Dekant, Wolfgang; Kohlbacher, Oliver; Huber, Christian G.

2015-01-01

Untargeted metabolomics has the potential to improve the predictivity of in vitro toxicity models and therefore may aid the replacement of expensive and laborious animal models. Here we describe a long term repeat dose nephrotoxicity study conducted on the human renal proximal tubular epithelial cell line, RPTEC/TERT1, treated with 10 and 35 μmol·liter−1 of chloroacetaldehyde, a metabolite of the anti-cancer drug ifosfamide. Our study outlines the establishment of an automated and easy to use untargeted metabolomics workflow for HPLC-high resolution mass spectrometry data. Automated data analysis workflows based on open source software (OpenMS, KNIME) enabled a comprehensive and reproducible analysis of the complex and voluminous metabolomics data produced by the profiling approach. Time- and concentration-dependent responses were clearly evident in the metabolomic profiles. To obtain a more comprehensive picture of the mode of action, transcriptomics and proteomics data were also integrated. For toxicity profiling of chloroacetaldehyde, 428 and 317 metabolite features were detectable in positive and negative modes, respectively, after stringent removal of chemical noise and unstable signals. Changes upon treatment were explored using principal component analysis, and statistically significant differences were identified using linear models for microarray assays. The analysis revealed toxic effects only for the treatment with 35 μmol·liter−1 for 3 and 14 days. The most regulated metabolites were glutathione and metabolites related to the oxidative stress response of the cells. These findings are corroborated by proteomics and transcriptomics data, which show, among other things, an activation of the Nrf2 and ATF4 pathways. PMID:26055719

Tissue-Specific Profiling Reveals Transcriptome Alterations in Arabidopsis Mutants Lacking Morphological Phenotypes[C][W

PubMed Central

Simon, Marissa; Bruex, Angela; Kainkaryam, Raghunandan M.; Zheng, Xiaohua; Huang, Ling; Woolf, Peter J.; Schiefelbein, John

2013-01-01

Traditional genetic analysis relies on mutants with observable phenotypes. Mutants lacking visible abnormalities may nevertheless exhibit molecular differences useful for defining gene function. To examine this, we analyzed tissue-specific transcript profiles from Arabidopsis thaliana transcription factor gene mutants with known roles in root epidermis development, but lacking a single-gene mutant phenotype due to genetic redundancy. We discovered substantial transcriptional changes in each mutant, preferentially affecting root epidermal genes in a manner consistent with the known double mutant effects. Furthermore, comparing transcript profiles of single and double mutants, we observed remarkable variation in the sensitivity of target genes to the loss of one or both paralogous genes, including preferential effects on specific branches of the epidermal gene network, likely reflecting the pathways of paralog subfunctionalization during evolution. In addition, we analyzed the root epidermal transcriptome of the transparent testa glabra2 mutant to clarify its role in the network. These findings provide insight into the molecular basis of genetic redundancy and duplicate gene diversification at the level of a specific gene regulatory network, and they demonstrate the usefulness of tissue-specific transcript profiling to define gene function in mutants lacking informative visible changes in phenotype. PMID:24014549

Comprehensive RNA-Seq Expression Analysis of Sensory Ganglia with a Focus on Ion Channels and GPCRs in Trigeminal Ganglia

PubMed Central

Manteniotis, Stavros; Lehmann, Ramona; Flegel, Caroline; Vogel, Felix; Hofreuter, Adrian; Schreiner, Benjamin S. P.; Altmüller, Janine; Becker, Christian; Schöbel, Nicole; Hatt, Hanns; Gisselmann, Günter

2013-01-01

The specific functions of sensory systems depend on the tissue-specific expression of genes that code for molecular sensor proteins that are necessary for stimulus detection and membrane signaling. Using the Next Generation Sequencing technique (RNA-Seq), we analyzed the complete transcriptome of the trigeminal ganglia (TG) and dorsal root ganglia (DRG) of adult mice. Focusing on genes with an expression level higher than 1 FPKM (fragments per kilobase of transcript per million mapped reads), we detected the expression of 12984 genes in the TG and 13195 in the DRG. To analyze the specific gene expression patterns of the peripheral neuronal tissues, we compared their gene expression profiles with that of the liver, brain, olfactory epithelium, and skeletal muscle. The transcriptome data of the TG and DRG were scanned for virtually all known G-protein-coupled receptors (GPCRs) as well as for ion channels. The expression profile was ranked with regard to the level and specificity for the TG. In total, we detected 106 non-olfactory GPCRs and 33 ion channels that had not been previously described as expressed in the TG. To validate the RNA-Seq data, in situ hybridization experiments were performed for several of the newly detected transcripts. To identify differences in expression profiles between the sensory ganglia, the RNA-Seq data of the TG and DRG were compared. Among the differentially expressed genes (> 1 FPKM), 65 and 117 were expressed at least 10-fold higher in the TG and DRG, respectively. Our transcriptome analysis allows a comprehensive overview of all ion channels and G protein-coupled receptors that are expressed in trigeminal ganglia and provides additional approaches for the investigation of trigeminal sensing as well as for the physiological and pathophysiological mechanisms of pain. PMID:24260241

Digital sorting of complex tissues for cell type-specific gene expression profiles.

PubMed

Zhong, Yi; Wan, Ying-Wooi; Pang, Kaifang; Chow, Lionel M L; Liu, Zhandong

2013-03-07

Cellular heterogeneity is present in almost all gene expression profiles. However, transcriptome analysis of tissue specimens often ignores the cellular heterogeneity present in these samples. Standard deconvolution algorithms require prior knowledge of the cell type frequencies within a tissue or their in vitro expression profiles. Furthermore, these algorithms tend to report biased estimations. Here, we describe a Digital Sorting Algorithm (DSA) for extracting cell-type specific gene expression profiles from mixed tissue samples that is unbiased and does not require prior knowledge of cell type frequencies. The results suggest that DSA is a specific and sensitivity algorithm in gene expression profile deconvolution and will be useful in studying individual cell types of complex tissues.

Mycobacterium tuberculosis Transcriptome Profiling in Mice with Genetically Different Susceptibility to Tuberculosis.

PubMed

Skvortsov, T A; Ignatov, D V; Majorov, K B; Apt, A S; Azhikina, T L

2013-04-01

Whole transcriptome profiling is now almost routinely used in various fields of biology, including microbiology. In vivo transcriptome studies usually provide relevant information about the biological processes in the organism and thus are indispensable for the formulation of hypotheses, testing, and correcting. In this study, we describe the results of genome-wide transcriptional profiling of the major human bacterial pathogen M. tuberculosis during its persistence in lungs. Two mouse strains differing in their susceptibility to tuberculosis were used for experimental infection with M. tuberculosis. Mycobacterial transcriptomes obtained from the infected tissues of the mice at two different time points were analyzed by deep sequencing and compared. It was hypothesized that the changes in the M. tuberculosis transcriptome may attest to the activation of the metabolism of lipids and amino acids, transition to anaerobic respiration, and increased expression of the factors modulating the immune response. A total of 209 genes were determined whose expression increased with disease progression in both host strains (commonly upregulated genes, CUG). Among them, the genes related to the functional categories of lipid metabolism, cell wall, and cell processes are of great interest. It was assumed that the products of these genes are involved in M. tuberculosis adaptation to the host immune system defense, thus being potential targets for drug development.

Intra-tumor heterogeneity in breast cancer has limited impact on transcriptomic-based molecular profiling.

PubMed

Karthik, Govindasamy-Muralidharan; Rantalainen, Mattias; Stålhammar, Gustav; Lövrot, John; Ullah, Ikram; Alkodsi, Amjad; Ma, Ran; Wedlund, Lena; Lindberg, Johan; Frisell, Jan; Bergh, Jonas; Hartman, Johan

2017-11-29

Transcriptomic profiling of breast tumors provides opportunity for subtyping and molecular-based patient stratification. In diagnostic applications the specimen profiled should be representative of the expression profile of the whole tumor and ideally capture properties of the most aggressive part of the tumor. However, breast cancers commonly exhibit intra-tumor heterogeneity at molecular, genomic and in phenotypic level, which can arise during tumor evolution. Currently it is not established to what extent a random sampling approach may influence molecular breast cancer diagnostics. In this study we applied RNA-sequencing to quantify gene expression in 43 pieces (2-5 pieces per tumor) from 12 breast tumors (Cohort 1). We determined molecular subtype and transcriptomic grade for all tumor pieces and analysed to what extent pieces originating from the same tumors are concordant or discordant with each other. Additionally, we validated our finding in an independent cohort consisting of 19 pieces (2-6 pieces per tumor) from 6 breast tumors (Cohort 2) profiled using microarray technique. Exome sequencing was also performed on this cohort, to investigate the extent of intra-tumor genomic heterogeneity versus the intra-tumor molecular subtype classifications. Molecular subtyping was consistent in 11 out of 12 tumors and transcriptomic grade assignments were consistent in 11 out of 12 tumors as well. Molecular subtype predictions revealed consistent subtypes in four out of six patients in this cohort 2. Interestingly, we observed extensive intra-tumor genomic heterogeneity in these tumor pieces but not in their molecular subtype classifications. Our results suggest that macroscopic intra-tumoral transcriptomic heterogeneity is limited and unlikely to have an impact on molecular diagnostics for most patients.

Transcriptome analysis of Cymbidium sinense and its application to the identification of genes associated with floral development

PubMed Central

2013-01-01

Background Cymbidium sinense belongs to the Orchidaceae, which is one of the most abundant angiosperm families. C. sinense, a high-grade traditional potted flower, is most prevalent in China and some Southeast Asian countries. The control of flowering time is a major bottleneck in the industrialized development of C. sinense. Little is known about the mechanisms responsible for floral development in this orchid. Moreover, genome references for entire transcriptome sequences do not currently exist for C. sinense. Thus, transcriptome and expression profiling data for this species are needed as an important resource to identify genes and to better understand the biological mechanisms of floral development in C. sinense. Results In this study, de novo transcriptome assembly and gene expression analysis using Illumina sequencing technology were performed. Transcriptome analysis assembles gene-related information related to vegetative and reproductive growth of C. sinense. Illumina sequencing generated 54,248,006 high quality reads that were assembled into 83,580 unigenes with an average sequence length of 612 base pairs, including 13,315 clusters and 70,265 singletons. A total of 41,687 (49.88%) unique sequences were annotated, 23,092 of which were assigned to specific metabolic pathways by the Kyoto Encyclopedia of Genes and Genomes (KEGG). Gene Ontology (GO) analysis of the annotated unigenes revealed that the majority of sequenced genes were associated with metabolic and cellular processes, cell and cell parts, catalytic activity and binding. Furthermore, 120 flowering-associated unigenes, 73 MADS-box unigenes and 28 CONSTANS-LIKE (COL) unigenes were identified from our collection. In addition, three digital gene expression (DGE) libraries were constructed for the vegetative phase (VP), floral differentiation phase (FDP) and reproductive phase (RP). The specific expression of many genes in the three development phases was also identified. 32 genes among three sub-libraries with high differential expression were selected as candidates connected with flower development. Conclusion RNA-seq and DGE profiling data provided comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the molecular mechanisms of floral development at three development phases of C. sinense. This data could be used as an important resource for investigating the genetics of the flowering pathway and various biological mechanisms in this orchid. PMID:23617896

Transcriptome analysis of Cymbidium sinense and its application to the identification of genes associated with floral development.

PubMed

Zhang, Jianxia; Wu, Kunlin; Zeng, Songjun; Teixeira da Silva, Jaime A; Zhao, Xiaolan; Tian, Chang-En; Xia, Haoqiang; Duan, Jun

2013-04-24

Cymbidium sinense belongs to the Orchidaceae, which is one of the most abundant angiosperm families. C. sinense, a high-grade traditional potted flower, is most prevalent in China and some Southeast Asian countries. The control of flowering time is a major bottleneck in the industrialized development of C. sinense. Little is known about the mechanisms responsible for floral development in this orchid. Moreover, genome references for entire transcriptome sequences do not currently exist for C. sinense. Thus, transcriptome and expression profiling data for this species are needed as an important resource to identify genes and to better understand the biological mechanisms of floral development in C. sinense. In this study, de novo transcriptome assembly and gene expression analysis using Illumina sequencing technology were performed. Transcriptome analysis assembles gene-related information related to vegetative and reproductive growth of C. sinense. Illumina sequencing generated 54,248,006 high quality reads that were assembled into 83,580 unigenes with an average sequence length of 612 base pairs, including 13,315 clusters and 70,265 singletons. A total of 41,687 (49.88%) unique sequences were annotated, 23,092 of which were assigned to specific metabolic pathways by the Kyoto Encyclopedia of Genes and Genomes (KEGG). Gene Ontology (GO) analysis of the annotated unigenes revealed that the majority of sequenced genes were associated with metabolic and cellular processes, cell and cell parts, catalytic activity and binding. Furthermore, 120 flowering-associated unigenes, 73 MADS-box unigenes and 28 CONSTANS-LIKE (COL) unigenes were identified from our collection. In addition, three digital gene expression (DGE) libraries were constructed for the vegetative phase (VP), floral differentiation phase (FDP) and reproductive phase (RP). The specific expression of many genes in the three development phases was also identified. 32 genes among three sub-libraries with high differential expression were selected as candidates connected with flower development. RNA-seq and DGE profiling data provided comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the molecular mechanisms of floral development at three development phases of C. sinense. This data could be used as an important resource for investigating the genetics of the flowering pathway and various biological mechanisms in this orchid.

Combined Analysis of the Chloroplast Genome and Transcriptome of the Antarctic Vascular Plant Deschampsia antarctica Desv

PubMed Central

Lee, Jungeun; Kang, Yoonjee; Shin, Seung Chul; Park, Hyun; Lee, Hyoungseok

2014-01-01

Background Antarctic hairgrass (Deschampsia antarctica Desv.) is the only natural grass species in the maritime Antarctic. It has been researched as an important ecological marker and as an extremophile plant for studies on stress tolerance. Despite its importance, little genomic information is available for D. antarctica. Here, we report the complete chloroplast genome, transcriptome profiles of the coding/noncoding genes, and the posttranscriptional processing by RNA editing in the chloroplast system. Results The complete chloroplast genome of D. antarctica is 135,362 bp in length with a typical quadripartite structure, including the large (LSC: 79,881 bp) and small (SSC: 12,519 bp) single-copy regions, separated by a pair of identical inverted repeats (IR: 21,481 bp). It contains 114 unique genes, including 81 unique protein-coding genes, 29 tRNA genes, and 4 rRNA genes. Sequence divergence analysis with other plastomes from the BEP clade of the grass family suggests a sister relationship between D. antarctica, Festuca arundinacea and Lolium perenne of the Poeae tribe, based on the whole plastome. In addition, we conducted high-resolution mapping of the chloroplast-derived transcripts. Thus, we created an expression profile for 81 protein-coding genes and identified ndhC, psbJ, rps19, psaJ, and psbA as the most highly expressed chloroplast genes. Small RNA-seq analysis identified 27 small noncoding RNAs of chloroplast origin that were preferentially located near the 5′- or 3′-ends of genes. We also found >30 RNA-editing sites in the D. antarctica chloroplast genome, with a dominance of C-to-U conversions. Conclusions We assembled and characterized the complete chloroplast genome sequence of D. antarctica and investigated the features of the plastid transcriptome. These data may contribute to a better understanding of the evolution of D. antarctica within the Poaceae family for use in molecular phylogenetic studies and may also help researchers understand the characteristics of the chloroplast transcriptome. PMID:24647560

Combined analysis of the chloroplast genome and transcriptome of the Antarctic vascular plant Deschampsia antarctica Desv.

PubMed

Lee, Jungeun; Kang, Yoonjee; Shin, Seung Chul; Park, Hyun; Lee, Hyoungseok

2014-01-01

Antarctic hairgrass (Deschampsia antarctica Desv.) is the only natural grass species in the maritime Antarctic. It has been researched as an important ecological marker and as an extremophile plant for studies on stress tolerance. Despite its importance, little genomic information is available for D. antarctica. Here, we report the complete chloroplast genome, transcriptome profiles of the coding/noncoding genes, and the posttranscriptional processing by RNA editing in the chloroplast system. The complete chloroplast genome of D. antarctica is 135,362 bp in length with a typical quadripartite structure, including the large (LSC: 79,881 bp) and small (SSC: 12,519 bp) single-copy regions, separated by a pair of identical inverted repeats (IR: 21,481 bp). It contains 114 unique genes, including 81 unique protein-coding genes, 29 tRNA genes, and 4 rRNA genes. Sequence divergence analysis with other plastomes from the BEP clade of the grass family suggests a sister relationship between D. antarctica, Festuca arundinacea and Lolium perenne of the Poeae tribe, based on the whole plastome. In addition, we conducted high-resolution mapping of the chloroplast-derived transcripts. Thus, we created an expression profile for 81 protein-coding genes and identified ndhC, psbJ, rps19, psaJ, and psbA as the most highly expressed chloroplast genes. Small RNA-seq analysis identified 27 small noncoding RNAs of chloroplast origin that were preferentially located near the 5'- or 3'-ends of genes. We also found >30 RNA-editing sites in the D. antarctica chloroplast genome, with a dominance of C-to-U conversions. We assembled and characterized the complete chloroplast genome sequence of D. antarctica and investigated the features of the plastid transcriptome. These data may contribute to a better understanding of the evolution of D. antarctica within the Poaceae family for use in molecular phylogenetic studies and may also help researchers understand the characteristics of the chloroplast transcriptome.

The impact of oxygen on the transcriptome of recombinant S. cerevisiae and P. pastoris - a comparative analysis.

PubMed

Baumann, Kristin; Dato, Laura; Graf, Alexandra B; Frascotti, Gianni; Dragosits, Martin; Porro, Danilo; Mattanovich, Diethard; Ferrer, Pau; Branduardi, Paola

2011-05-09

Saccharomyces cerevisiae and Pichia pastoris are two of the most relevant microbial eukaryotic platforms for the production of recombinant proteins. Their known genome sequences enabled several transcriptomic profiling studies under many different environmental conditions, thus mimicking not only perturbations and adaptations which occur in their natural surroundings, but also in industrial processes. Notably, the majority of such transcriptome analyses were performed using non-engineered strains.In this comparative study, the gene expression profiles of S. cerevisiae and P. pastoris, a Crabtree positive and Crabtree negative yeast, respectively, were analyzed for three different oxygenation conditions (normoxic, oxygen-limited and hypoxic) under recombinant protein producing conditions in chemostat cultivations. The major differences in the transcriptomes of S. cerevisiae and P. pastoris were observed between hypoxic and normoxic conditions, where the availability of oxygen strongly affected ergosterol biosynthesis, central carbon metabolism and stress responses, particularly the unfolded protein response. Steady state conditions under low oxygen set-points seemed to perturb the transcriptome of S. cerevisiae to a much lesser extent than the one of P. pastoris, reflecting the major tolerance of the baker's yeast towards oxygen limitation, and a higher fermentative capacity. Further important differences were related to Fab production, which was not significantly affected by oxygen availability in S. cerevisiae, while a clear productivity increase had been previously reported for hypoxically grown P. pastoris. The effect of three different levels of oxygen availability on the physiology of P. pastoris and S. cerevisiae revealed a very distinct remodelling of the transcriptional program, leading to novel insights into the different adaptive responses of Crabtree negative and positive yeasts to oxygen availability. Moreover, the application of such comparative genomic studies to recombinant hosts grown in different environments might lead to the identification of key factors for efficient protein production.

The genome- and transcriptome-wide analysis of innate immunity in the brown planthopper, Nilaparvata lugens

PubMed Central

2013-01-01

Background The brown planthopper (Nilaparvata lugens) is one of the most serious rice plant pests in Asia. N. lugens causes extensive rice damage by sucking rice phloem sap, which results in stunted plant growth and the transmission of plant viruses. Despite the importance of this insect pest, little is known about the immunological mechanisms occurring in this hemimetabolous insect species. Results In this study, we performed a genome- and transcriptome-wide analysis aiming at the immune-related genes. The transcriptome datasets include the N. lugens intestine, the developmental stage, wing formation, and sex-specific expression information that provided useful gene expression sequence data for the genome-wide analysis. As a result, we identified a large number of genes encoding N. lugens pattern recognition proteins, modulation proteins in the prophenoloxidase (proPO) activating cascade, immune effectors, and the signal transduction molecules involved in the immune pathways, including the Toll, Immune deficiency (Imd) and Janus kinase signal transducers and activators of transcription (JAK-STAT) pathways. The genome scale analysis revealed detailed information of the gene structure, distribution and transcription orientations in scaffolds. A comparison of the genome-available hemimetabolous and metabolous insect species indicate the differences in the immune-related gene constitution. We investigated the gene expression profiles with regards to how they responded to bacterial infections and tissue, as well as development and sex expression specificity. Conclusions The genome- and transcriptome-wide analysis of immune-related genes including pattern recognition and modulation molecules, immune effectors, and the signal transduction molecules involved in the immune pathways is an important step in determining the overall architecture and functional network of the immune components in N. lugens. Our findings provide the comprehensive gene sequence resource and expression profiles of the immune-related genes of N. lugens, which could facilitate the understanding of the innate immune mechanisms in the hemimetabolous insect species. These data give insight into clarifying the potential functional roles of the immune-related genes involved in the biological processes of development, reproduction, and virus transmission in N. lugens. PMID:23497397

Antennal Transcriptome Analysis and Comparison of Chemosensory Gene Families in Two Closely Related Noctuidae Moths, Helicoverpa armigera and H. assulta

PubMed Central

Zhang, Jin; Wang, Bing; Dong, Shuanglin; Cao, Depan; Dong, Junfeng; Walker, William B.; Liu, Yang; Wang, Guirong

2015-01-01

To better understand the olfactory mechanisms in the two lepidopteran pest model species, the Helicoverpa armigera and H. assulta, we conducted transcriptome analysis of the adult antennae using Illumina sequencing technology and compared the chemosensory genes between these two related species. Combined with the chemosensory genes we had identified previously in H. armigera by 454 sequencing, we identified 133 putative chemosensory unigenes in H. armigera including 60 odorant receptors (ORs), 19 ionotropic receptors (IRs), 34 odorant binding proteins (OBPs), 18 chemosensory proteins (CSPs), and 2 sensory neuron membrane proteins (SNMPs). Consistent with these results, 131 putative chemosensory genes including 64 ORs, 19 IRs, 29 OBPs, 17 CSPs, and 2 SNMPs were identified through male and female antennal transcriptome analysis in H. assulta. Reverse Transcription-PCR (RT-PCR) was conducted in H. assulta to examine the accuracy of the assembly and annotation of the transcriptome and the expression profile of these unigenes in different tissues. Most of the ORs, IRs and OBPs were enriched in adult antennae, while almost all the CSPs were expressed in antennae as well as legs. We compared the differences of the chemosensory genes between these two species in detail. Our work will surely provide valuable information for further functional studies of pheromones and host volatile recognition genes in these two related species. PMID:25659090

SPARTA: Simple Program for Automated reference-based bacterial RNA-seq Transcriptome Analysis.

PubMed

Johnson, Benjamin K; Scholz, Matthew B; Teal, Tracy K; Abramovitch, Robert B

2016-02-04

Many tools exist in the analysis of bacterial RNA sequencing (RNA-seq) transcriptional profiling experiments to identify differentially expressed genes between experimental conditions. Generally, the workflow includes quality control of reads, mapping to a reference, counting transcript abundance, and statistical tests for differentially expressed genes. In spite of the numerous tools developed for each component of an RNA-seq analysis workflow, easy-to-use bacterially oriented workflow applications to combine multiple tools and automate the process are lacking. With many tools to choose from for each step, the task of identifying a specific tool, adapting the input/output options to the specific use-case, and integrating the tools into a coherent analysis pipeline is not a trivial endeavor, particularly for microbiologists with limited bioinformatics experience. To make bacterial RNA-seq data analysis more accessible, we developed a Simple Program for Automated reference-based bacterial RNA-seq Transcriptome Analysis (SPARTA). SPARTA is a reference-based bacterial RNA-seq analysis workflow application for single-end Illumina reads. SPARTA is turnkey software that simplifies the process of analyzing RNA-seq data sets, making bacterial RNA-seq analysis a routine process that can be undertaken on a personal computer or in the classroom. The easy-to-install, complete workflow processes whole transcriptome shotgun sequencing data files by trimming reads and removing adapters, mapping reads to a reference, counting gene features, calculating differential gene expression, and, importantly, checking for potential batch effects within the data set. SPARTA outputs quality analysis reports, gene feature counts and differential gene expression tables and scatterplots. SPARTA provides an easy-to-use bacterial RNA-seq transcriptional profiling workflow to identify differentially expressed genes between experimental conditions. This software will enable microbiologists with limited bioinformatics experience to analyze their data and integrate next generation sequencing (NGS) technologies into the classroom. The SPARTA software and tutorial are available at sparta.readthedocs.org.

Persistent foot-and-mouth disease virus infection in the nasopharynx of cattle: tissue-specific distribution and local cytokine expression

USDA-ARS?s Scientific Manuscript database

Tissues obtained post-mortem from cattle persistently infected with foot-and-mouth disease virus (FMDV) were analyzed to characterize the tissue-specific localization of FMDV and partial transcriptome profiles for selected immunoregulatory cytokines. Analysis of 28 distinct anatomic sites from 21 st...

Analysis of petunia hybrida in response to salt stress using high throughput RNA sequencing

USDA-ARS?s Scientific Manuscript database

Salt and drought are among the greatest challenges to crop and native plants in meeting their yield and reproductive potentials. DNA sequencing-enabled transcriptome profiling provides a means of assessing what genes are responding to salt or drought stress so as to better understand the molecular ...

Serum Metabolomic Profiling and Liver Transcriptomic Analysis Provide Mechanistic Evidence of Ozone (O3)- Induced Systemic Metabolic Impairment

EPA Science Inventory

Recently, air pollution has been linked to insulin resistance and obesity but the mechanisms remain to be elucidated. We have recently shown that acute O3 exposure induces glucose intolerance, hyperglycemia and increases in leptin and epinephrine in rats. Here, we hypothesized th...

Transcriptome analysis reveals persistent effects of neonatal diet on small intestine gene expression profile in a porcine model

USDA-ARS?s Scientific Manuscript database

Breastfeeding is associated with several benefits affecting gut development and immune function. Compared to breast feeding, infant formula feeding is linked to a greater risk for gut dysfunction, ear and respiratory tract infections, and allergies. The beneficial effects appear to last at least thr...

Transcriptome Analysis of Thermal Parthenogenesis of the Domesticated Silkworm.

PubMed

Liu, Peigang; Wang, Yongqiang; Du, Xin; Yao, Lusong; Li, Fengbo; Meng, Zhiqi

2015-01-01

Thermal induction of parthenogenesis (also known as thermal parthenogenesis) in silkworms is an important technique that has been used in artificial insemination, expansion of hybridization, transgenesis and sericultural production; however, the exact mechanisms of this induction remain unclear. This study aimed to investigate the gene expression profile in silkworms undergoing thermal parthenogenesis using RNA-seq analysis. The transcriptome profiles indicated that in non-induced and induced eggs, the numbers of differentially expressed genes (DEGs) for the parthenogenetic line (PL) and amphigenetic line (AL) were 538 and 545, respectively, as determined by fold-change ≥ 2. Gene ontology (GO) analysis showed that DEGs between two lines were mainly involved in reproduction, formation of chorion, female gamete generation and cell development pathways. Upregulation of many chorion genes in AL suggests that the maturation rate of AL eggs was slower than PL eggs. Some DEGs related to reactive oxygen species removal, DNA repair and heat shock response were differentially expressed between the two lines, such as MPV-17, REV1 and HSP68. These results supported the view that a large fraction of genes are differentially expressed between PL and AL, which offers a new approach to identifying the molecular mechanism of silkworm thermal parthenogenesis.

Transcriptome Analysis of Thermal Parthenogenesis of the Domesticated Silkworm

PubMed Central

Du, Xin; Yao, Lusong; Li, Fengbo; Meng, Zhiqi

2015-01-01

Thermal induction of parthenogenesis (also known as thermal parthenogenesis) in silkworms is an important technique that has been used in artificial insemination, expansion of hybridization, transgenesis and sericultural production; however, the exact mechanisms of this induction remain unclear. This study aimed to investigate the gene expression profile in silkworms undergoing thermal parthenogenesis using RNA-seq analysis. The transcriptome profiles indicated that in non-induced and induced eggs, the numbers of differentially expressed genes (DEGs) for the parthenogenetic line (PL) and amphigenetic line (AL) were 538 and 545, respectively, as determined by fold-change ≥ 2. Gene ontology (GO) analysis showed that DEGs between two lines were mainly involved in reproduction, formation of chorion, female gamete generation and cell development pathways. Upregulation of many chorion genes in AL suggests that the maturation rate of AL eggs was slower than PL eggs. Some DEGs related to reactive oxygen species removal, DNA repair and heat shock response were differentially expressed between the two lines, such as MPV-17, REV1 and HSP68. These results supported the view that a large fraction of genes are differentially expressed between PL and AL, which offers a new approach to identifying the molecular mechanism of silkworm thermal parthenogenesis. PMID:26274803

Root Type-Specific Reprogramming of Maize Pericycle Transcriptomes by Local High Nitrate Results in Disparate Lateral Root Branching Patterns1[OPEN

PubMed Central

Lithio, Andrew

2016-01-01

The adaptability of root system architecture to unevenly distributed mineral nutrients in soil is a key determinant of plant performance. The molecular mechanisms underlying nitrate dependent plasticity of lateral root branching across the different root types of maize are only poorly understood. In this study, detailed morphological and anatomical analyses together with cell type-specific transcriptome profiling experiments combining laser capture microdissection with RNA-seq were performed to unravel the molecular signatures of lateral root formation in primary, seminal, crown, and brace roots of maize (Zea mays) upon local high nitrate stimulation. The four maize root types displayed divergent branching patterns of lateral roots upon local high nitrate stimulation. In particular, brace roots displayed an exceptional architectural plasticity compared to other root types. Transcriptome profiling revealed root type-specific transcriptomic reprogramming of pericycle cells upon local high nitrate stimulation. The alteration of the transcriptomic landscape of brace root pericycle cells in response to local high nitrate stimulation was most significant. Root type-specific transcriptome diversity in response to local high nitrate highlighted differences in the functional adaptability and systemic shoot nitrogen starvation response during development. Integration of morphological, anatomical, and transcriptomic data resulted in a framework underscoring similarity and diversity among root types grown in heterogeneous nitrate environments. PMID:26811190

Integrating Genomic Analysis with the Genetic Basis of Gene Expression: Preliminary Evidence of the Identification of Causal Genes for Cardiovascular and Metabolic Traits Related to Nutrition in Mexicans123

PubMed Central

Bastarrachea, Raúl A.; Gallegos-Cabriales, Esther C.; Nava-González, Edna J.; Haack, Karin; Voruganti, V. Saroja; Charlesworth, Jac; Laviada-Molina, Hugo A.; Veloz-Garza, Rosa A.; Cardenas-Villarreal, Velia Margarita; Valdovinos-Chavez, Salvador B.; Gomez-Aguilar, Patricia; Meléndez, Guillermo; López-Alvarenga, Juan Carlos; Göring, Harald H. H.; Cole, Shelley A.; Blangero, John; Comuzzie, Anthony G.; Kent, Jack W.

2012-01-01

Whole-transcriptome expression profiling provides novel phenotypes for analysis of complex traits. Gene expression measurements reflect quantitative variation in transcript-specific messenger RNA levels and represent phenotypes lying close to the action of genes. Understanding the genetic basis of gene expression will provide insight into the processes that connect genotype to clinically significant traits representing a central tenet of system biology. Synchronous in vivo expression profiles of lymphocytes, muscle, and subcutaneous fat were obtained from healthy Mexican men. Most genes were expressed at detectable levels in multiple tissues, and RNA levels were correlated between tissue types. A subset of transcripts with high reliability of expression across tissues (estimated by intraclass correlation coefficients) was enriched for cis-regulated genes, suggesting that proximal sequence variants may influence expression similarly in different cellular environments. This integrative global gene expression profiling approach is proving extremely useful for identifying genes and pathways that contribute to complex clinical traits. Clearly, the coincidence of clinical trait quantitative trait loci and expression quantitative trait loci can help in the prioritization of positional candidate genes. Such data will be crucial for the formal integration of positional and transcriptomic information characterized as genetical genomics. PMID:22797999

Transcriptome assembly and expression profiling of the molecular responses to cadmium toxicity in cerebral ganglia of wolf spider Pardosa pseudoannulata (Araneae: Lycosidae).

PubMed

Yang, Huilin; Peng, Yuande; Shi, Yixue; Tian, Jianxiang; Wang, Juan; Peng, Xianjin; Xie, Chunliang; Xu, Xiang; Song, Qisheng; Wang, Zhi; Lv, Zhiyue

2018-03-01

Cadmium (Cd) is a heavy metal that can cause irreversible toxicity to animals, and is an environmental pollutant in farmlands. Spiders are considered to be an excellent model for investigating the impacts of heavy metals on the environment. To date, the changes at the molecular level in the cerebral ganglia of spiders are poorly understood. Cd exposure leads to strong damage in the nervous system, such as apoptosis and necrosis of nerve cells, therefore we conducted a transcriptomic analysis of Pardosa pseudoannulata cerebral ganglia under Cd stress to profile differential gene expression (DGE). We obtained a total of 123,328 assembled unigenes, and 1441 Cd stress-associated DEGs between the Cd-treated and control groups. Expression profile analysis demonstrated that many genes involved in calcium signaling, cGMP-PKG signaling, tyrosine metabolism, phototransduction-fly, melanogenesis and isoquinoline alkaloid biosynthesis were up-regulated under Cd stress, whereas oxidative phosphorylation-related, nervous disease-associated, non-alcoholic fatty liver disease-associated, and ribosomal-associated genes were down-regulated. Here, we provide a comprehensive set of DEGs influenced by Cd stress, and heavy metal stress, and provide new information for elucidating the neurotoxic mechanisms of Cd stress in spiders.

Comparative transcriptome sequencing and de novo analysis of Vaccinium corymbosum during fruit and color development.

PubMed

Li, Lingli; Zhang, Hehua; Liu, Zhongshuai; Cui, Xiaoyue; Zhang, Tong; Li, Yanfang; Zhang, Lingyun

2016-10-12

Blueberry is an economically important fruit crop in Ericaceae family. The substantial quantities of flavonoids in blueberry have been implicated in a broad range of health benefits. However, the information regarding fruit development and flavonoid metabolites based on the transcriptome level is still limited. In the present study, the transcriptome and gene expression profiling over berry development, especially during color development were initiated. A total of approximately 13.67 Gbp of data were obtained and assembled into 186,962 transcripts and 80,836 unigenes from three stages of blueberry fruit and color development. A large number of simple sequence repeats (SSRs) and candidate genes, which are potentially involved in plant development, metabolic and hormone pathways, were identified. A total of 6429 sequences containing 8796 SSRs were characterized from 15,457 unigenes and 1763 unigenes contained more than one SSR. The expression profiles of key genes involved in anthocyanin biosynthesis were also studied. In addition, a comparison between our dataset and other published results was carried out. Our high quality reads produced in this study are an important advancement and provide a new resource for the interpretation of high-throughput data for blueberry species whether regarding sequencing data depth or species extension. The use of this transcriptome data will serve as a valuable public information database for the studies of blueberry genome and would greatly boost the research of fruit and color development, flavonoid metabolisms and regulation and breeding of more healthful blueberries.

Benchmarking Water Quality from Wastewater to Drinking Waters Using Reduced Transcriptome of Human Cells.

PubMed

Xia, Pu; Zhang, Xiaowei; Zhang, Hanxin; Wang, Pingping; Tian, Mingming; Yu, Hongxia

2017-08-15

One of the major challenges in environmental science is monitoring and assessing the risk of complex environmental mixtures. In vitro bioassays with limited key toxicological end points have been shown to be suitable to evaluate mixtures of organic pollutants in wastewater and recycled water. Omics approaches such as transcriptomics can monitor biological effects at the genome scale. However, few studies have applied omics approach in the assessment of mixtures of organic micropollutants. Here, an omics approach was developed for profiling bioactivity of 10 water samples ranging from wastewater to drinking water in human cells by a reduced human transcriptome (RHT) approach and dose-response modeling. Transcriptional expression of 1200 selected genes were measured by an Ampliseq technology in two cell lines, HepG2 and MCF7, that were exposed to eight serial dilutions of each sample. Concentration-effect models were used to identify differentially expressed genes (DEGs) and to calculate effect concentrations (ECs) of DEGs, which could be ranked to investigate low dose response. Furthermore, molecular pathways disrupted by different samples were evaluated by Gene Ontology (GO) enrichment analysis. The ability of RHT for representing bioactivity utilizing both HepG2 and MCF7 was shown to be comparable to the results of previous in vitro bioassays. Finally, the relative potencies of the mixtures indicated by RHT analysis were consistent with the chemical profiles of the samples. RHT analysis with human cells provides an efficient and cost-effective approach to benchmarking mixture of micropollutants and may offer novel insight into the assessment of mixture toxicity in water.

Determination of male strobilus developmental stages by cytological and gene expression analyses in Japanese cedar (Cryptomeria japonica).

PubMed

Tsubomura, Miyoko; Kurita, Manabu; Watanabe, Atsushi

2016-05-01

The molecular mechanisms that control male strobilus development in conifers are largely unknown because the developmental stages and related genes have not yet been characterized. The determination of male strobilus developmental stages will contribute to genetic research and reproductive biology in conifers. Our objectives in this study were to determine the developmental stages of male strobili by cytological and transcriptome analysis, and to determine the stages at which aberrant morphology is observed in a male-sterile mutant of Cryptomeria japonica D. Don to better understand the molecular mechanisms that control male strobilus and pollen development. Male strobilus development was observed for 8 months, from initiation to pollen dispersal. A set of 19,209 expressed sequence tags (ESTs) collected from a male reproductive library and a pollen library was used for microarray analysis. We divided male strobilus development into 10 stages by cytological and transcriptome analysis. Eight clusters (7324 ESTs) exhibited major changes in transcriptome profiles during male strobili and pollen development in C. japonica Two clusters showed a gradual increase and decline in transcript abundance, respectively, while the other six clusters exhibited stage-specific changes. The stages at which the male sterility trait of Sosyun was expressed were identified using information on male strobilus and pollen developmental stages and gene expression profiles. Aberrant morphology was observed cytologically at Stage 6 (microspore stage), and differences in expression patterns compared with wild type were observed at Stage 4 (tetrad stage). © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Gonad Transcriptome Analysis of High-Temperature-Treated Females and High-Temperature-Induced Sex-Reversed Neomales in Nile Tilapia

PubMed Central

Sun, Li Xue; Teng, Jian; Zhao, Yan; Li, Ning; Wang, Hui

2018-01-01

Background: Nowadays, the molecular mechanisms governing TSD (temperature-dependent sex determination) or GSD + TE (genotypic sex determination + temperature effects) remain a mystery in fish. Methods: We developed three all-female families of Nile tilapia (Oreochromis niloticus), and the family with the highest male ratio after high-temperature treatment was used for transcriptome analysis. Results: First, gonadal histology analysis indicated that the histological morphology of control females (CF) was not significantly different from that of high-temperature-treated females (TF) at various development stages. However, the high-temperature treatment caused a lag of spermatogenesis in high-temperature-induced neomales (IM). Next, we sequenced the transcriptome of CF, TF, and IM Nile tilapia. 79, 11,117, and 11,000 differentially expressed genes (DEGs) were detected in the CF–TF, CF–IM, and TF–IM comparisons, respectively, and 44 DEGs showed identical expression changes in the CF–TF and CF–IM comparisons. Principal component analysis (PCA) indicated that three individuals in CF and three individuals in TF formed a cluster, and three individuals in IM formed a distinct cluster, which confirmed that the gonad transcriptome profile of TF was similar to that of CF and different from that of IM. Finally, six sex-related genes were validated by qRT-PCR. Conclusions: This study identifies a number of genes that may be involved in GSD + TE, which will be useful for investigating the molecular mechanisms of TSD or GSD + TE in fish. PMID:29495590

Gonad Transcriptome Analysis of High-Temperature-Treated Females and High-Temperature-Induced Sex-Reversed Neomales in Nile Tilapia.

PubMed

Sun, Li Xue; Teng, Jian; Zhao, Yan; Li, Ning; Wang, Hui; Ji, Xiang Shan

2018-02-28

Nowadays, the molecular mechanisms governing TSD (temperature-dependent sex determination) or GSD + TE (genotypic sex determination + temperature effects) remain a mystery in fish. We developed three all-female families of Nile tilapia ( Oreochromis niloticus ), and the family with the highest male ratio after high-temperature treatment was used for transcriptome analysis. First, gonadal histology analysis indicated that the histological morphology of control females (CF) was not significantly different from that of high-temperature-treated females (TF) at various development stages. However, the high-temperature treatment caused a lag of spermatogenesis in high-temperature-induced neomales (IM). Next, we sequenced the transcriptome of CF, TF, and IM Nile tilapia. 79, 11,117, and 11,000 differentially expressed genes (DEGs) were detected in the CF-TF, CF-IM, and TF-IM comparisons, respectively, and 44 DEGs showed identical expression changes in the CF-TF and CF-IM comparisons. Principal component analysis (PCA) indicated that three individuals in CF and three individuals in TF formed a cluster, and three individuals in IM formed a distinct cluster, which confirmed that the gonad transcriptome profile of TF was similar to that of CF and different from that of IM. Finally, six sex-related genes were validated by qRT-PCR. This study identifies a number of genes that may be involved in GSD + TE, which will be useful for investigating the molecular mechanisms of TSD or GSD + TE in fish.

Transcriptome Profile Analysis of Breast Muscle Tissues from High or Low Levels of Atmospheric Ammonia Exposed Broilers (Gallus gallus)

PubMed Central

Sa, Renna; Zhong, Ruqing; Xing, Huan; Zhang, Hongfu

2016-01-01

Atmospheric ammonia is a common problem in poultry industry. High concentrations of aerial ammonia cause great harm to broilers' health and production. For the consideration of human health, the limit exposure concentration of ammonia in houses is set at 25 ppm. Previous reports have shown that 25 ppm is still detrimental to livestock, especially the gastrointestinal tract and respiratory tract, but the negative relationship between ammonia exposure and the tissue of breast muscle of broilers is still unknown. In the present study, 25 ppm ammonia in poultry houses was found to lower slaughter performance and breast yield. Then, high-throughput RNA sequencing was utilized to identify differentially expressed genes in breast muscle of broiler chickens exposed to high (25 ppm) or low (3 ppm) levels of atmospheric ammonia. The transcriptome analysis showed that 163 genes (fold change ≥ 2 or ≤ 0.5; P-value < 0.05) were differentially expressed between Ammonia25 (treatment group) and Ammonia3 (control group), including 96 down-regulated and 67 up-regulated genes. qRT-PCR analysis validated the transcriptomic results of RNA sequencing. Gene Ontology (GO) functional annotation analysis revealed potential genes, processes and pathways with putative involvement in growth and development inhibition of breast muscle in broilers caused by aerial ammonia exposure. This study facilitates understanding of the genetic architecture of the chicken breast muscle transcriptome, and has identified candidate genes for breast muscle response to atmospheric ammonia exposure. PMID:27611572

De Novo Sequencing and Analysis of Lemongrass Transcriptome Provide First Insights into the Essential Oil Biosynthesis of Aromatic Grasses.

PubMed

Meena, Seema; Kumar, Sarma R; Venkata Rao, D K; Dwivedi, Varun; Shilpashree, H B; Rastogi, Shubhra; Shasany, Ajit K; Nagegowda, Dinesh A

2016-01-01

Aromatic grasses of the genus Cymbopogon (Poaceae family) represent unique group of plants that produce diverse composition of monoterpene rich essential oils, which have great value in flavor, fragrance, cosmetic, and aromatherapy industries. Despite the commercial importance of these natural aromatic oils, their biosynthesis at the molecular level remains unexplored. As the first step toward understanding the essential oil biosynthesis, we performed de novo transcriptome assembly and analysis of C. flexuosus (lemongrass) by employing Illumina sequencing. Mining of transcriptome data and subsequent phylogenetic analysis led to identification of terpene synthases, pyrophosphatases, alcohol dehydrogenases, aldo-keto reductases, carotenoid cleavage dioxygenases, alcohol acetyltransferases, and aldehyde dehydrogenases, which are potentially involved in essential oil biosynthesis. Comparative essential oil profiling and mRNA expression analysis in three Cymbopogon species (C. flexuosus, aldehyde type; C. martinii, alcohol type; and C. winterianus, intermediate type) with varying essential oil composition indicated the involvement of identified candidate genes in the formation of alcohols, aldehydes, and acetates. Molecular modeling and docking further supported the role of identified protein sequences in aroma formation in Cymbopogon. Also, simple sequence repeats were found in the transcriptome with many linked to terpene pathway genes including the genes potentially involved in aroma biosynthesis. This work provides the first insights into the essential oil biosynthesis of aromatic grasses, and the identified candidate genes and markers can be a great resource for biotechnological and molecular breeding approaches to modulate the essential oil composition.

Transcriptomic insights on the ABC transporter gene family in the salmon louse Caligus rogercresseyi.

PubMed

Valenzuela-Muñoz, Valentina; Sturm, Armin; Gallardo-Escárate, Cristian

2015-04-09

ATP-binding cassette (ABC) protein family encode for membrane proteins involved in the transport of various biomolecules through the cellular membrane. These proteins have been identified in all taxa and present important physiological functions, including the process of insecticide detoxification in arthropods. For that reason the ectoparasite Caligus rogercresseyi represents a model species for understanding the molecular underpinnings involved in insecticide drug resistance. llumina sequencing was performed using sea lice exposed to 2 and 3 ppb of deltamethrin and azamethiphos. Contigs obtained from de novo assembly were annotated by Blastx. RNA-Seq analysis was performed and validated by qPCR analysis. From the transcriptome database of C. rogercresseyi, 57 putative members of ABC protein sequences were identified and phylogenetically classified into the eight subfamilies described for ABC transporters in arthropods. Transcriptomic profiles for ABC proteins subfamilies were evaluated throughout C. rogercresseyi development. Moreover, RNA-Seq analysis was performed for adult male and female salmon lice exposed to the delousing drugs azamethiphos and deltamethrin. High transcript levels of the ABCB and ABCC subfamilies were evidenced. Furthermore, SNPs mining was carried out for the ABC proteins sequences, revealing pivotal genomic information. The present study gives a comprehensive transcriptome analysis of ABC proteins from C. rogercresseyi, providing relevant information about transporter roles during ontogeny and in relation to delousing drug responses in salmon lice. This genomic information represents a valuable tool for pest management in the Chilean salmon aquaculture industry.

De Novo Sequencing and Analysis of Lemongrass Transcriptome Provide First Insights into the Essential Oil Biosynthesis of Aromatic Grasses

PubMed Central

Meena, Seema; Kumar, Sarma R.; Venkata Rao, D. K.; Dwivedi, Varun; Shilpashree, H. B.; Rastogi, Shubhra; Shasany, Ajit K.; Nagegowda, Dinesh A.

2016-01-01

Aromatic grasses of the genus Cymbopogon (Poaceae family) represent unique group of plants that produce diverse composition of monoterpene rich essential oils, which have great value in flavor, fragrance, cosmetic, and aromatherapy industries. Despite the commercial importance of these natural aromatic oils, their biosynthesis at the molecular level remains unexplored. As the first step toward understanding the essential oil biosynthesis, we performed de novo transcriptome assembly and analysis of C. flexuosus (lemongrass) by employing Illumina sequencing. Mining of transcriptome data and subsequent phylogenetic analysis led to identification of terpene synthases, pyrophosphatases, alcohol dehydrogenases, aldo-keto reductases, carotenoid cleavage dioxygenases, alcohol acetyltransferases, and aldehyde dehydrogenases, which are potentially involved in essential oil biosynthesis. Comparative essential oil profiling and mRNA expression analysis in three Cymbopogon species (C. flexuosus, aldehyde type; C. martinii, alcohol type; and C. winterianus, intermediate type) with varying essential oil composition indicated the involvement of identified candidate genes in the formation of alcohols, aldehydes, and acetates. Molecular modeling and docking further supported the role of identified protein sequences in aroma formation in Cymbopogon. Also, simple sequence repeats were found in the transcriptome with many linked to terpene pathway genes including the genes potentially involved in aroma biosynthesis. This work provides the first insights into the essential oil biosynthesis of aromatic grasses, and the identified candidate genes and markers can be a great resource for biotechnological and molecular breeding approaches to modulate the essential oil composition. PMID:27516768

From root to fruit: RNA-Seq analysis shows that arbuscular mycorrhizal symbiosis may affect tomato fruit metabolism.

PubMed

Zouari, Inès; Salvioli, Alessandra; Chialva, Matteo; Novero, Mara; Miozzi, Laura; Tenore, Gian Carlo; Bagnaresi, Paolo; Bonfante, Paola

2014-03-21

Tomato (Solanum lycopersicum) establishes a beneficial symbiosis with arbuscular mycorrhizal (AM) fungi. The formation of the mycorrhizal association in the roots leads to plant-wide modulation of gene expression. To understand the systemic effect of the fungal symbiosis on the tomato fruit, we used RNA-Seq to perform global transcriptome profiling on Moneymaker tomato fruits at the turning ripening stage. Fruits were collected at 55 days after flowering, from plants colonized with Funneliformis mosseae and from control plants, which were fertilized to avoid responses related to nutrient deficiency. Transcriptome analysis identified 712 genes that are differentially expressed in fruits from mycorrhizal and control plants. Gene Ontology (GO) enrichment analysis of these genes showed 81 overrepresented functional GO classes. Up-regulated GO classes include photosynthesis, stress response, transport, amino acid synthesis and carbohydrate metabolism functions, suggesting a general impact of fungal symbiosis on primary metabolisms and, particularly, on mineral nutrition. Down-regulated GO classes include cell wall, metabolism and ethylene response pathways. Quantitative RT-PCR validated the RNA-Seq results for 12 genes out of 14 when tested at three fruit ripening stages, mature green, breaker and turning. Quantification of fruit nutraceutical and mineral contents produced values consistent with the expression changes observed by RNA-Seq analysis. This RNA-Seq profiling produced a novel data set that explores the intersection of mycorrhization and fruit development. We found that the fruits of mycorrhizal plants show two transcriptomic "signatures": genes characteristic of a climacteric fleshy fruit, and genes characteristic of mycorrhizal status, like phosphate and sulphate transporters. Moreover, mycorrhizal plants under low nutrient conditions produce fruits with a nutrient content similar to those from non-mycorrhizal plants under high nutrient conditions, indicating that AM fungi can help replace exogenous fertilizer for fruit crops.

Comparative transcriptome analysis reveals vertebrate phylotypic period during organogenesis

PubMed Central

Irie, Naoki; Kuratani, Shigeru

2011-01-01

One of the central issues in evolutionary developmental biology is how we can formulate the relationships between evolutionary and developmental processes. Two major models have been proposed: the 'funnel-like' model, in which the earliest embryo shows the most conserved morphological pattern, followed by diversifying later stages, and the 'hourglass' model, in which constraints are imposed to conserve organogenesis stages, which is called the phylotypic period. Here we perform a quantitative comparative transcriptome analysis of several model vertebrate embryos and show that the pharyngula stage is most conserved, whereas earlier and later stages are rather divergent. These results allow us to predict approximate developmental timetables between different species, and indicate that pharyngula embryos have the most conserved gene expression profiles, which may be the source of the basic body plan of vertebrates. PMID:21427719

Composite transcriptome assembly of RNA-seq data in a sheep model for delayed bone healing.

PubMed

Jäger, Marten; Ott, Claus-Eric; Grünhagen, Johannes; Hecht, Jochen; Schell, Hanna; Mundlos, Stefan; Duda, Georg N; Robinson, Peter N; Lienau, Jasmin

2011-03-24

The sheep is an important model organism for many types of medically relevant research, but molecular genetic experiments in the sheep have been limited by the lack of knowledge about ovine gene sequences. Prior to our study, mRNA sequences for only 1,556 partial or complete ovine genes were publicly available. Therefore, we developed a composite de novo transcriptome assembly method for next-generation sequence data to combine known ovine mRNA and EST sequences, mRNA sequences from mouse and cow, and sequences assembled de novo from short read RNA-Seq data into a composite reference transcriptome, and identified transcripts from over 12 thousand previously undescribed ovine genes. Gene expression analysis based on these data revealed substantially different expression profiles in standard versus delayed bone healing in an ovine tibial osteotomy model. Hundreds of transcripts were differentially expressed between standard and delayed healing and between the time points of the standard and delayed healing groups. We used the sheep sequences to design quantitative RT-PCR assays with which we validated the differential expression of 26 genes that had been identified by RNA-seq analysis. A number of clusters of characteristic expression profiles could be identified, some of which showed striking differences between the standard and delayed healing groups. Gene Ontology (GO) analysis showed that the differentially expressed genes were enriched in terms including extracellular matrix, cartilage development, contractile fiber, and chemokine activity. Our results provide a first atlas of gene expression profiles and differentially expressed genes in standard and delayed bone healing in a large-animal model and provide a number of clues as to the shifts in gene expression that underlie delayed bone healing. In the course of our study, we identified transcripts of 13,987 ovine genes, including 12,431 genes for which no sequence information was previously available. This information will provide a basis for future molecular research involving the sheep as a model organism.

Composite transcriptome assembly of RNA-seq data in a sheep model for delayed bone healing

PubMed Central

2011-01-01

Background The sheep is an important model organism for many types of medically relevant research, but molecular genetic experiments in the sheep have been limited by the lack of knowledge about ovine gene sequences. Results Prior to our study, mRNA sequences for only 1,556 partial or complete ovine genes were publicly available. Therefore, we developed a composite de novo transcriptome assembly method for next-generation sequence data to combine known ovine mRNA and EST sequences, mRNA sequences from mouse and cow, and sequences assembled de novo from short read RNA-Seq data into a composite reference transcriptome, and identified transcripts from over 12 thousand previously undescribed ovine genes. Gene expression analysis based on these data revealed substantially different expression profiles in standard versus delayed bone healing in an ovine tibial osteotomy model. Hundreds of transcripts were differentially expressed between standard and delayed healing and between the time points of the standard and delayed healing groups. We used the sheep sequences to design quantitative RT-PCR assays with which we validated the differential expression of 26 genes that had been identified by RNA-seq analysis. A number of clusters of characteristic expression profiles could be identified, some of which showed striking differences between the standard and delayed healing groups. Gene Ontology (GO) analysis showed that the differentially expressed genes were enriched in terms including extracellular matrix, cartilage development, contractile fiber, and chemokine activity. Conclusions Our results provide a first atlas of gene expression profiles and differentially expressed genes in standard and delayed bone healing in a large-animal model and provide a number of clues as to the shifts in gene expression that underlie delayed bone healing. In the course of our study, we identified transcripts of 13,987 ovine genes, including 12,431 genes for which no sequence information was previously available. This information will provide a basis for future molecular research involving the sheep as a model organism. PMID:21435219

Transcriptome complexity in cardiac development and diseases--an expanding universe between genome and phenome.

PubMed

Gao, Chen; Wang, Yibin

2014-01-01

With the advancement of transcriptome profiling by micro-arrays and high-throughput RNA-sequencing, transcriptome complexity and its dynamics are revealed at different levels in cardiovascular development and diseases. In this review, we will highlight the recent progress in our knowledge of cardiovascular transcriptome complexity contributed by RNA splicing, RNA editing and noncoding RNAs. The emerging importance of many of these previously under-explored aspects of gene regulation in cardiovascular development and pathology will be discussed.

RNA-Seq Transcriptome Profiling of Upland Cotton (Gossypium hirsutum L.) Root Tissue under Water-Deficit Stress

PubMed Central

Bowman, Megan J.; Park, Wonkeun; Bauer, Philip J.; Udall, Joshua A.; Page, Justin T.; Raney, Joshua; Scheffler, Brian E.; Jones, Don. C.; Campbell, B. Todd

2013-01-01

An RNA-Seq experiment was performed using field grown well-watered and naturally rain fed cotton plants to identify differentially expressed transcripts under water-deficit stress. Our work constitutes the first application of the newly published diploid D5 Gossypium raimondii sequence in the study of tetraploid AD1 upland cotton RNA-seq transcriptome analysis. A total of 1,530 transcripts were differentially expressed between well-watered and water-deficit stressed root tissues, in patterns that confirm the accuracy of this technique for future studies in cotton genomics. Additionally, putative sequence based genome localization of differentially expressed transcripts detected A2 genome specific gene expression under water-deficit stress. These data will facilitate efforts to understand the complex responses governing transcriptomic regulatory mechanisms and to identify candidate genes that may benefit applied plant breeding programs. PMID:24324815

A joint analysis of transcriptomic and metabolomic data uncovers enhanced enzyme-metabolite coupling in breast cancer

NASA Astrophysics Data System (ADS)

Auslander, Noam; Yizhak, Keren; Weinstock, Adam; Budhu, Anuradha; Tang, Wei; Wang, Xin Wei; Ambs, Stefan; Ruppin, Eytan

2016-07-01

Disrupted regulation of cellular processes is considered one of the hallmarks of cancer. We analyze metabolomic and transcriptomic profiles jointly collected from breast cancer and hepatocellular carcinoma patients to explore the associations between the expression of metabolic enzymes and the levels of the metabolites participating in the reactions they catalyze. Surprisingly, both breast cancer and hepatocellular tumors exhibit an increase in their gene-metabolites associations compared to noncancerous adjacent tissues. Following, we build predictors of metabolite levels from the expression of the enzyme genes catalyzing them. Applying these predictors to a large cohort of breast cancer samples we find that depleted levels of key cancer-related metabolites including glucose, glycine, serine and acetate are significantly associated with improved patient survival. Thus, we show that the levels of a wide range of metabolites in breast cancer can be successfully predicted from the transcriptome, going beyond the limited set of those measured.

Characterization of the cork oak transcriptome dynamics during acorn development.

PubMed

Miguel, Andreia; de Vega-Bartol, José; Marum, Liliana; Chaves, Inês; Santo, Tatiana; Leitão, José; Varela, Maria Carolina; Miguel, Célia M

2015-06-25

Cork oak (Quercus suber L.) has a natural distribution across western Mediterranean regions and is a keystone forest tree species in these ecosystems. The fruiting phase is especially critical for its regeneration but the molecular mechanisms underlying the biochemical and physiological changes during cork oak acorn development are poorly understood. In this study, the transcriptome of the cork oak acorn, including the seed, was characterized in five stages of development, from early development to acorn maturation, to identify the dominant processes in each stage and reveal transcripts with important functions in gene expression regulation and response to water. A total of 80,357 expressed sequence tags (ESTs) were de novo assembled from RNA-Seq libraries representative of the several acorn developmental stages. Approximately 7.6 % of the total number of transcripts present in Q. suber transcriptome was identified as acorn specific. The analysis of expression profiles during development returned 2,285 differentially expressed (DE) transcripts, which were clustered into six groups. The stage of development corresponding to the mature acorn exhibited an expression profile markedly different from other stages. Approximately 22 % of the DE transcripts putatively code for transcription factors (TF) or transcriptional regulators, and were found almost equally distributed among the several expression profile clusters, highlighting their major roles in controlling the whole developmental process. On the other hand, carbohydrate metabolism, the biological pathway most represented during acorn development, was especially prevalent in mid to late stages as evidenced by enrichment analysis. We further show that genes related to response to water, water deprivation and transport were mostly represented during the early (S2) and the last stage (S8) of acorn development, when tolerance to water desiccation is possibly critical for acorn viability. To our knowledge this work represents the first report of acorn development transcriptomics in oaks. The obtained results provide novel insights into the developmental biology of cork oak acorns, highlighting transcripts putatively involved in the regulation of the gene expression program and in specific processes likely essential for adaptation. It is expected that this knowledge can be transferred to other oak species of great ecological value.

Transcriptomic analysis revealed the mechanism of oil dynamic accumulation during developing Siberian apricot (Prunus sibirica L.) seed kernels for the development of woody biodiesel.

PubMed

Niu, Jun; An, Jiyong; Wang, Libing; Fang, Chengliang; Ha, Denglong; Fu, Chengyu; Qiu, Lin; Yu, Haiyan; Zhao, Haiyan; Hou, Xinyu; Xiang, Zheng; Zhou, Sufan; Zhang, Zhixiang; Feng, Xinyi; Lin, Shanzhi

2015-01-01

Siberian apricot (Prunus sibirica L.) has emerged as a novel potential source of biodiesel in China, but the molecular regulatory mechanism of oil accumulation in Siberian apricot seed kernels (SASK) is still unknown at present. To better develop SASK oil as woody biodiesel, it is essential to profile transcriptome and to identify the full repertoire of potential unigenes involved in the formation and accumulation of oil SASK during the different developing stages. We firstly detected the temporal patterns for oil content and fatty acid (FA) compositions of SASK in 7 different developing stages. The best time for obtaining the high quality and quantity of SASK oil was characterized at 60 days after flowering (DAF), and the representative periods (10, 30, 50, 60, and 70 DAF) were selected for transcriptomic analysis. By Illumina/Solexa sequencings, approximately 65 million short reads (average length = 96 bp) were obtained, and then assembled into 124,070 unigenes by Trinity strategy (mean size = 829.62 bp). A total of 3,000, 2,781, 2,620, and 2,675 differentially expressed unigenes were identified at 30, 50, 60, and 70 DAF (10 DAF as the control) by DESeq method, respectively. The relationship between the unigene transcriptional profiles and the oil dynamic patterns in developing SASK was comparatively analyzed, and the specific unigenes encoding some known enzymes and transcription factors involved in acetyl-coenzyme A (acetyl-CoA) formation and oil accumulation were determined. Additionally, 5 key metabolic genes implicated in SASK oil accumulation were experimentally validated by quantitative real-time PCR (qRT-PCR). Our findings could help to construction of oil accumulated pathway and to elucidate the molecular regulatory mechanism of increased oil production in developing SASK. This is the first study of oil temporal patterns, transcriptome sequencings, and differential profiles in developing SASK. All our results will serve as the important foundation to further deeply explore the regulatory mechanism of SASK high-quality oil accumulation, and may also provide some reference for researching the woody biodiesel plants.

Comparative analysis of the transcriptome responses of zebrafish embryos after exposure to low concentrations of cadmium, cobalt and copper.

PubMed

Sonnack, Laura; Klawonn, Thorsten; Kriehuber, Ralf; Hollert, Henner; Schäfers, Christoph; Fenske, Martina

2018-03-01

Metal toxicity is a global environmental challenge. Fish are particularly prone to metal exposure, which can be lethal or cause sublethal physiological impairments. The objective of this study was to investigate how adverse effects of chronic exposure to non-toxic levels of essential and non-essential metals in early life stage zebrafish may be explained by changes in the transcriptome. We therefore studied the effects of three different metals at low concentrations in zebrafish embryos by transcriptomics analysis. The study design compared exposure effects caused by different metals at different developmental stages (pre-hatch and post-hatch). Wild-type embryos were exposed to solutions of low concentrations of copper (CuSO 4 ), cadmium (CdCl 2 ) and cobalt (CoSO 4 ) until 96h post-fertilization (hpf) and microarray experiments were carried out to determine transcriptome profiles at 48 and 96hpf. We found that the toxic metal cadmium affected the expression of more genes at 96hpf than 48hpf. The opposite effect was observed for the essential metals cobalt and copper, which also showed enrichment of different GO terms. Genes involved in neuromast and motor neuron development were significantly enriched, agreeing with our previous results showing motor neuron and neuromast damage in the embryos. Our data provide evidence that the response of the transcriptome of fish embryos to metal exposure differs for essential and non-essential metals. Copyright © 2017 Elsevier Inc. All rights reserved.

TRAM (Transcriptome Mapper): database-driven creation and analysis of transcriptome maps from multiple sources

PubMed Central

2011-01-01

Background Several tools have been developed to perform global gene expression profile data analysis, to search for specific chromosomal regions whose features meet defined criteria as well as to study neighbouring gene expression. However, most of these tools are tailored for a specific use in a particular context (e.g. they are species-specific, or limited to a particular data format) and they typically accept only gene lists as input. Results TRAM (Transcriptome Mapper) is a new general tool that allows the simple generation and analysis of quantitative transcriptome maps, starting from any source listing gene expression values for a given gene set (e.g. expression microarrays), implemented as a relational database. It includes a parser able to assign univocal and updated gene symbols to gene identifiers from different data sources. Moreover, TRAM is able to perform intra-sample and inter-sample data normalization, including an original variant of quantile normalization (scaled quantile), useful to normalize data from platforms with highly different numbers of investigated genes. When in 'Map' mode, the software generates a quantitative representation of the transcriptome of a sample (or of a pool of samples) and identifies if segments of defined lengths are over/under-expressed compared to the desired threshold. When in 'Cluster' mode, the software searches for a set of over/under-expressed consecutive genes. Statistical significance for all results is calculated with respect to genes localized on the same chromosome or to all genome genes. Transcriptome maps, showing differential expression between two sample groups, relative to two different biological conditions, may be easily generated. We present the results of a biological model test, based on a meta-analysis comparison between a sample pool of human CD34+ hematopoietic progenitor cells and a sample pool of megakaryocytic cells. Biologically relevant chromosomal segments and gene clusters with differential expression during the differentiation toward megakaryocyte were identified. Conclusions TRAM is designed to create, and statistically analyze, quantitative transcriptome maps, based on gene expression data from multiple sources. The release includes FileMaker Pro database management runtime application and it is freely available at http://apollo11.isto.unibo.it/software/, along with preconfigured implementations for mapping of human, mouse and zebrafish transcriptomes. PMID:21333005

Comprehensive Analysis of the Triterpenoid Saponins Biosynthetic Pathway in Anemone flaccida by Transcriptome and Proteome Profiling

PubMed Central

Zhan, Chuansong; Li, Xiaohua; Zhao, Zeying; Yang, Tewu; Wang, Xuekui; Luo, Biaobiao; Zhang, Qiyun; Hu, Yanru; Hu, Xuebo

2016-01-01

Background: Anemone flaccida Fr. Shmidt (Ranunculaceae), commonly known as ‘Di Wu’ in China, is a perennial herb with limited distribution. The rhizome of A. flaccida has long been used to treat arthritis as a tradition in China. Studies disclosed that the plant contains a rich source of triterpenoid saponins. However, little is known about triterpenoid saponins biosynthesis in A. flaccida. Results: In this study, we conducted the tandem transcriptome and proteome profiling of a non-model medicinal plant, A. flaccida. Using Illumina HiSeq 2000 sequencing and iTRAQ technique, a total of 46,962 high-quality unigenes were obtained with an average sequence length of 1,310 bp, along with 1473 unique proteins from A. flaccida. Among the A. flaccida transcripts, 36,617 (77.97%) showed significant similarity (E-value < 1e-5) to the known proteins in the public database. Of the total 46,962 unigenes, 36,617 open reading frame (ORFs) were predicted. By the fragments per kilobases per million reads (FPKM) statistics, 14,004 isoforms/unigenes were found to be upregulated, and 14,090 isoforms/unigenes were down-regulated in the rhizomes as compared to those in the leaves. Based on the bioinformatics analysis, all possible enzymes involved in the triterpenoid saponins biosynthetic pathway of A. flaccida were identified, including cytosolic mevalonate pathway (MVA) and the plastidial methylerythritol pathway (MEP). Additionally, a total of 126 putative cytochrome P450 (CYP450) and 32 putative UDP glycosyltransferases were selected as the candidates of triterpenoid saponins modifiers. Among them, four of them were annotated as the gene of CYP716A subfamily, the key enzyme in the oleanane-type triterpenoid saponins biosynthetic pathway. Furthermore, based on RNA-Seq and proteome analysis, as well as quantitative RT-PCR verification, the expression level of gene and protein committed to triterpenoids biosynthesis in the leaf versus the rhizome was compared. Conclusion: A combination of the de novo transcriptome and proteome profiling based on the Illumina HiSeq 2000 sequencing platform and iTRAQ technique was shown to be a powerful method for the discovery of candidate genes, which encoded enzymes that were responsible for the biosynthesis of novel secondary metabolites in a non-model plant. The transcriptome data of our study provides a very important resource for the understanding of the triterpenoid saponins biosynthesis of A. flaccida. PMID:27504115

Comparative Transcriptomes and EVO-DEVO Studies Depending on Next Generation Sequencing.

PubMed

Liu, Tiancheng; Yu, Lin; Liu, Lei; Li, Hong; Li, Yixue

2015-01-01

High throughput technology has prompted the progressive omics studies, including genomics and transcriptomics. We have reviewed the improvement of comparative omic studies, which are attributed to the high throughput measurement of next generation sequencing technology. Comparative genomics have been successfully applied to evolution analysis while comparative transcriptomics are adopted in comparison of expression profile from two subjects by differential expression or differential coexpression, which enables their application in evolutionary developmental biology (EVO-DEVO) studies. EVO-DEVO studies focus on the evolutionary pressure affecting the morphogenesis of development and previous works have been conducted to illustrate the most conserved stages during embryonic development. Old measurements of these studies are based on the morphological similarity from macro view and new technology enables the micro detection of similarity in molecular mechanism. Evolutionary model of embryo development, which includes the "funnel-like" model and the "hourglass" model, has been evaluated by combination of these new comparative transcriptomic methods with prior comparative genomic information. Although the technology has promoted the EVO-DEVO studies into a new era, technological and material limitation still exist and further investigations require more subtle study design and procedure.

Detecting early signs of heat and drought stress in Phoenix dactylifera (date palm)

PubMed Central

Safronov, Omid; Kreuzwieser, Jürgen; Haberer, Georg; Alyousif, Mohamed S.; Schulze, Waltraud; Al-Harbi, Naif; Arab, Leila; Ache, Peter; Stempfl, Thomas; Kruse, Joerg; Mayer, Klaus X.; Hedrich, Rainer; Rennenberg, Heinz

2017-01-01

Plants adapt to the environment by either long-term genome evolution or by acclimatization processes where the cellular processes and metabolism of the plant are adjusted within the existing potential in the genome. Here we studied the adaptation strategies in date palm, Phoenix dactylifera, under mild heat, drought and combined heat and drought by transcriptomic and metabolomic profiling. In transcriptomics data, combined heat and drought resembled heat response, whereas in metabolomics data it was more similar to drought. In both conditions, soluble carbohydrates, such as fucose, and glucose derivatives, were increased, suggesting a switch to carbohydrate metabolism and cell wall biogenesis. This result is consistent with the evidence from transcriptomics and cis-motif analysis. In addition, transcriptomics data showed transcriptional activation of genes related to reactive oxygen species in all three conditions (drought, heat, and combined heat and drought), suggesting increased activity of enzymatic antioxidant systems in cytosol, chloroplast and peroxisome. Finally, the genes that were differentially expressed in heat and combined heat and drought stresses were significantly enriched for circadian and diurnal rhythm motifs, suggesting new stress avoidance strategies. PMID:28570677

Comparative Transcriptome Analysis of Bacillus subtilis Responding to Dissolved Oxygen in Adenosine Fermentation

PubMed Central

Yin, Chun-Yun; Zhou, Ying; Ye, Bang-Ce

2011-01-01

Dissolved oxygen (DO) is an important factor for adenosine fermentation. Our previous experiments have shown that low oxygen supply in the growth period was optimal for high adenosine yield. Herein, to better understand the link between oxygen supply and adenosine productivity in B. subtilis (ATCC21616), we sought to systematically explore the effect of DO on genetic regulation and metabolism through transcriptome analysis. The microarrays representing 4,106 genes were used to study temporal transcript profiles of B. subtilis fermentation in response to high oxygen supply (agitation 700 r/min) and low oxygen supply (agitation 450 r/min). The transcriptome data analysis revealed that low oxygen supply has three major effects on metabolism: enhance carbon metabolism (glucose metabolism, pyruvate metabolism and carbon overflow), inhibit degradation of nitrogen sources (glutamate family amino acids and xanthine) and purine synthesis. Inhibition of xanthine degradation was the reason that low oxygen supply enhanced adenosine production. These provide us with potential targets, which can be modified to achieve higher adenosine yield. Expression of genes involved in energy, cell type differentiation, protein synthesis was also influenced by oxygen supply. These results provided new insights into the relationship between oxygen supply and metabolism. PMID:21625606

Transcriptomic data analysis and differential gene expression of antioxidant pathways in king penguin juveniles (Aptenodytes patagonicus) before and after acclimatization to marine life.

PubMed

Rey, Benjamin; Dégletagne, Cyril; Duchamp, Claude

2016-12-01

In this article, we present differentially expressed gene profiles in the pectoralis muscle of wild juvenile king penguins that were either naturally acclimated to cold marine environment or experimentally immersed in cold water as compared with penguin juveniles that never experienced cold water immersion. Transcriptomic data were obtained by hybridizing penguins total cDNA on Affymetrix GeneChip Chicken Genome arrays and analyzed using maxRS algorithm , " Transcriptome analysis in non-model species: a new method for the analysis of heterologous hybridization on microarrays " (Dégletagne et al., 2010) [1] . We focused on genes involved in multiple antioxidant pathways. For better clarity, these differentially expressed genes were clustered into six functional groups according to their role in controlling redox homeostasis. The data are related to a comprehensive research study on the ontogeny of antioxidant functions in king penguins, "Hormetic response triggers multifaceted anti-oxidant strategies in immature king penguins (Aptenodytes patagonicus)" (Rey et al., 2016) [2] . The raw microarray dataset supporting the present analyses has been deposited at the Gene Expression Omnibus (GEO) repository under accessions GEO: GSE17725 and GEO: GSE82344.

Antennal transcriptome analysis of the Asian longhorned beetle Anoplophora glabripennis

PubMed Central

Hu, Ping; Wang, Jingzhen; Cui, Mingming; Tao, Jing; Luo, Youqing

2016-01-01

Olfactory proteins form the basis of insect olfactory recognition, which is crucial for host identification, mating, and oviposition. Using transcriptome analysis of Anoplophora glabripennis antenna, we identified 42 odorant-binding proteins (OBPs), 12 chemosensory proteins (CSPs), 14 pheromone-degrading enzymes (PDEs), 1 odorant-degrading enzymes (ODE), 37 odorant receptors (ORs), 11 gustatory receptors (GRs), 2 sensory neuron membrane proteins (SNMPs), and 4 ionotropic receptor (IR). All CSPs and PBPs were expressed in antennae, confirming the authenticity of the transcriptome data. CSP expression profiles showed that AglaCSP3, AglaCSP6, and AglaCSP12 were expressed preferentially in maxillary palps and AglaCSP7 and AglaCSP9 were strongly expressed in antennae. The vast majority of CSPs were highly expressed in multiple chemosensory tissues, suggesting their participation in olfactory recognition in almost all olfactory tissues. Intriguingly, the PBP AglaPBP2 was preferentially expressed in antenna, indicating that it is the main protein involved in efficient and sensitive pheromone recognition. Phylogenetic analysis of olfactory proteins indicated AglaGR1 may detect CO2. This study establishes a foundation for determining the chemoreception molecular mechanisms of A. glabripennis, which would provide a new perspective for controlling pest populations, especially those of borers. PMID:27222053

Whole transcriptome analysis of the fasting and fed Burmese python heart: insights into extreme physiological cardiac adaptation.

PubMed

Wall, Christopher E; Cozza, Steven; Riquelme, Cecilia A; McCombie, W Richard; Heimiller, Joseph K; Marr, Thomas G; Leinwand, Leslie A

2011-01-01

The infrequently feeding Burmese python (Python molurus) experiences significant and rapid postprandial cardiac hypertrophy followed by regression as digestion is completed. To begin to explore the molecular mechanisms of this response, we have sequenced and assembled the fasted and postfed Burmese python heart transcriptomes with Illumina technology using the chicken (Gallus gallus) genome as a reference. In addition, we have used RNA-seq analysis to identify differences in the expression of biological processes and signaling pathways between fasted, 1 day postfed (DPF), and 3 DPF hearts. Out of a combined transcriptome of ∼2,800 mRNAs, 464 genes were differentially expressed. Genes showing differential expression at 1 DPF compared with fasted were enriched for biological processes involved in metabolism and energetics, while genes showing differential expression at 3 DPF compared with fasted were enriched for processes involved in biogenesis, structural remodeling, and organization. Moreover, we present evidence for the activation of physiological and not pathological signaling pathways in this rapid, novel model of cardiac growth in pythons. Together, our data provide the first comprehensive gene expression profile for a reptile heart.

Transcriptome analysis of eyestalk and hemocytes in the ridgetail white prawn Exopalaemon carinicauda: assembly, annotation and marker discovery.

PubMed

Li, Jitao; Li, Jian; Chen, Ping; Liu, Ping; He, Yuying

2015-01-01

The ridgetail white prawn Exopalaemon carinicauda is one of major economic mariculture species in eastern China. The deficiency of genomic and transcriptomic data is becoming the bottleneck of further researches on its good traits. In the present study, 454 pyrosequencing was undertaken to investigate the transcriptome profiles of E. carinicauda. A collection of 1,028,710 sequence reads (459.59 Mb) obtained from cDNA prepared from eyestalk and hemocytes was assembled into 162,056 expressed sequence tags (ESTs). Of these, 29.88 % of 48,428 contigs and 70.12 % of 113,628 singlets possessed high similarities to sequences in the GenBank non-redundant database, with most significant (E value <1e(-10)) unigenes matches occurring with crustacean and insect sequences. KEGG analysis of unigenes identified putative members of biological pathways related to growth and immunity. In addition, we obtained a total of putative 125,112 SNPs and 13,467 microsatellites. These results will contribute to the understanding of the genome makeup and provide useful information for future functional genomic research in E. carinicauda.

Transcriptomic Analysis of the Adaptation of Listeria monocytogenes to Lagoon and Soil Matrices Associated with a Piggery Environment: Comparison of Expression Profiles

PubMed Central

Vivant, Anne-Laure; Desneux, Jeremy; Pourcher, Anne-Marie; Piveteau, Pascal

2017-01-01

Understanding how Listeria monocytogenes, the causative agent of listeriosis, adapts to the environment is crucial. Adaptation to new matrices requires regulation of gene expression. To determine how the pathogen adapts to lagoon effluent and soil, two matrices where L. monocytogenes has been isolated, we compared the transcriptomes of L. monocytogenes CIP 110868 20 min and 24 h after its transfer to effluent and soil extract. Results showed major variations in the transcriptome of L. monocytogenes in the lagoon effluent but only minor modifications in the soil. In both the lagoon effluent and in the soil, genes involved in mobility and chemotaxis and in the transport of carbohydrates were the most frequently represented in the set of genes with higher transcript levels, and genes with phage-related functions were the most represented in the set of genes with lower transcript levels. A modification of the cell envelop was only found in the lagoon environment. Finally, the differential analysis included a large proportion of regulators, regulons, and ncRNAs. PMID:29018416

Transcriptomic Analysis of the Adaptation of Listeria monocytogenes to Lagoon and Soil Matrices Associated with a Piggery Environment: Comparison of Expression Profiles.

PubMed

Vivant, Anne-Laure; Desneux, Jeremy; Pourcher, Anne-Marie; Piveteau, Pascal

2017-01-01

Understanding how Listeria monocytogenes , the causative agent of listeriosis, adapts to the environment is crucial. Adaptation to new matrices requires regulation of gene expression. To determine how the pathogen adapts to lagoon effluent and soil, two matrices where L. monocytogenes has been isolated, we compared the transcriptomes of L. monocytogenes CIP 110868 20 min and 24 h after its transfer to effluent and soil extract. Results showed major variations in the transcriptome of L. monocytogenes in the lagoon effluent but only minor modifications in the soil. In both the lagoon effluent and in the soil, genes involved in mobility and chemotaxis and in the transport of carbohydrates were the most frequently represented in the set of genes with higher transcript levels, and genes with phage-related functions were the most represented in the set of genes with lower transcript levels. A modification of the cell envelop was only found in the lagoon environment. Finally, the differential analysis included a large proportion of regulators, regulons, and ncRNAs.

ARG1 Functions in the Physiological Adaptation of Undifferentiated Plant Cells to Spaceflight

NASA Astrophysics Data System (ADS)

Zupanska, Agata K.; Schultz, Eric R.; Yao, JiQiang; Sng, Natasha J.; Zhou, Mingqi; Callaham, Jordan B.; Ferl, Robert J.; Paul, Anna-Lisa

2017-11-01

Scientific access to spaceflight and especially the International Space Station has revealed that physiological adaptation to spaceflight is accompanied or enabled by changes in gene expression that significantly alter the transcriptome of cells in spaceflight. A wide range of experiments have shown that plant physiological adaptation to spaceflight involves gene expression changes that alter cell wall and other metabolisms. However, while transcriptome profiling aptly illuminates changes in gene expression that accompany spaceflight adaptation, mutation analysis is required to illuminate key elements required for that adaptation. Here we report how transcriptome profiling was used to gain insight into the spaceflight adaptation role of Altered response to gravity 1 (Arg1), a gene known to affect gravity responses in plants on Earth. The study compared expression profiles of cultured lines of Arabidopsis thaliana derived from wild-type (WT) cultivar Col-0 to profiles from a knock-out line deficient in the gene encoding ARG1 (ARG1 KO), both on the ground and in space. The cell lines were launched on SpaceX CRS-2 as part of the Cellular Expression Logic (CEL) experiment of the BRIC-17 spaceflight mission. The cultured cell lines were grown within 60 mm Petri plates in Petri Dish Fixation Units (PDFUs) that were housed within the Biological Research In Canisters (BRIC) hardware. Spaceflight samples were fixed on orbit. Differentially expressed genes were identified between the two environments (spaceflight and comparable ground controls) and the two genotypes (WT and ARG1 KO). Each genotype engaged unique genes during physiological adaptation to the spaceflight environment, with little overlap. Most of the genes altered in expression in spaceflight in WT cells were found to be Arg1-dependent, suggesting a major role for that gene in the physiological adaptation of undifferentiated cells to spaceflight.

Temporal transcriptome changes induced by methyl jasmonate in Salvia sclarea.

PubMed

Hao, Da Cheng; Chen, Shi Lin; Osbourn, Anne; Kontogianni, Vassiliki G; Liu, Li Wei; Jordán, Maria J

2015-03-01

Salvia sclarea is a traditional medicinal and aromatic plant that grows in Europe and produces various economically important compounds, including phenylpropanoid derivatives and terpenoids. Methyl jasmonate (MeJA) is commonly used to elicit plant stress responses. However, how MeJA enhances production of secondary metabolites in S. sclarea is not well understood. We performed a genome-wide analysis of temporal gene expression in S. sclarea leaves and roots. The transcriptome profiles 0, 10 and 26 h after MeJA treatment were analyzed by Illumina RNA-Seq. A total of 16,142 isogenes (average length 866bp; N50 1035bp) were obtained by de novo assembly of 35,757,567 raw sequencing reads. When these sequencing reads were mapped onto the assembled Unigenes, 3236, 2792 and 798 Unigenes were found to be expressed differentially between 0 and 10h, 0 and 26 h, and 10 and 26h, respectively. These included many secondary metabolite biosynthesis, stress and defense-related genes. A qRT-PCR analysis confirmed the expression profiles of selected differentially expressed genes (DEGs) revealed by RNA-Seq data, and also extended our analysis of differential gene expression to 73 h. Our investigations revealed temporal differences in the responses of S. sclarea to MeJA treatment. MeJA treatment induced the expression of a large number of genes involved in phenylpropanoid biosynthesis, especially between 0 and 10h, and 0 and 26 h. Additionally, many genes encoding transcription factors, cytochrome P450s, glycosyltransferases, methyltransferases and transporters were shown to respond to MeJA elicitation. DEGs related to structural molecule activity and cell death showed a significant temporal variation. A chromatographic analysis of metabolites at 26h, 73h and six days after MeJA treatment indicated that these transcriptomic changes precede MeJA-induced changes in secondary metabolite content. This study sheds light on the molecular mechanisms of MeJA elicitation and is helpful in understanding how exogenous MeJA treatment mediates extensive plant transcriptome reprogramming/remodeling. Our results can be utilized to characterize genes related to secondary metabolism and their regulation, and in breeding S. sclarea for desirable chemotypes. Copyright © 2014 Elsevier B.V. All rights reserved.

De novo transcriptome assembly and RNA-Seq expression analysis in blood from beluga whales of Bristol Bay, AK.

PubMed

Morey, Jeanine S; Burek Huntington, Kathy A; Campbell, Michelle; Clauss, Tonya M; Goertz, Caroline E; Hobbs, Roderick C; Lunardi, Denise; Moors, Amanda J; Neely, Marion G; Schwacke, Lori H; Van Dolah, Frances M

2017-10-01

Assessing the health of marine mammal sentinel species is crucial to understanding the impacts of environmental perturbations on marine ecosystems and human health. In Arctic regions, beluga whales, Delphinapterus leucas, are upper level predators that may serve as a sentinel species, potentially forecasting impacts on human health. While gene expression profiling from blood transcriptomes has widely been used to assess health status and environmental exposures in human and veterinary medicine, its use in wildlife has been limited due to the lack of available genomes and baseline data. To this end we constructed the first beluga whale blood transcriptome de novo from samples collected during annual health assessments of the healthy Bristol Bay, AK stock during 2012-2014 to establish baseline information on the content and variation of the beluga whale blood transcriptome. The Trinity transcriptome assembly from beluga was comprised of 91,325 transcripts that represented a wide array of cellular functions and processes and was extremely similar in content to the blood transcriptome of another cetacean, the bottlenose dolphin. Expression of hemoglobin transcripts was much lower in beluga (25.6% of TPM, transcripts per million) than has been observed in many other mammals. A T12A amino acid substitution in the HBB sequence of beluga whales, but not bottlenose dolphins, was identified and may play a role in low temperature adaptation. The beluga blood transcriptome was extremely stable between sex and year, with no apparent clustering of samples by principle components analysis and <4% of genes differentially expressed (EBseq, FDR<0.05). While the impacts of season, sexual maturity, disease, and geography on the beluga blood transcriptome must be established, the presence of transcripts involved in stress, detoxification, and immune functions indicate that blood gene expression analyses may provide information on health status and exposure. This study provides a wealth of transcriptomic data on beluga whales and provides a sizeable pool of preliminary data for comparison with other studies in beluga whale. Copyright © 2017 Elsevier B.V. All rights reserved.

Laser Capture Microdissection Revisited as a Tool for Transcriptomic Analysis: Application of an Excel-Based qPCR Preparation Software (PREXCEL-Q)

USDA-ARS?s Scientific Manuscript database

The ability to reliably analyze cellular and molecular profiles of normal or diseased tissues is frequently obfuscated by the inherent heterogeneous nature of tissues. Laser Capture Microdissection (LCM) is an innovative technique that allows the isolation and enrichment of pure subpopulations of c...

Transcriptome analysis of Aspergillus flavus reveals isolate specific gene profiles in the response to oxidative stresses and carbon sources in vitro

USDA-ARS?s Scientific Manuscript database

Aflatoxin contamination of peanut and maize is exacerbated by drought stress. Reactive oxygen species (ROS) are produced in host plants during drought/heat stress, and are hypothesized to stimulate aflatoxin production. In order to better understand why Aspergillus flavus produces aflatoxin and the ...

Linear Regression Links Transcriptomic Data and Cellular Raman Spectra.

PubMed

Kobayashi-Kirschvink, Koseki J; Nakaoka, Hidenori; Oda, Arisa; Kamei, Ken-Ichiro F; Nosho, Kazuki; Fukushima, Hiroko; Kanesaki, Yu; Yajima, Shunsuke; Masaki, Haruhiko; Ohta, Kunihiro; Wakamoto, Yuichi

2018-06-08

Raman microscopy is an imaging technique that has been applied to assess molecular compositions of living cells to characterize cell types and states. However, owing to the diverse molecular species in cells and challenges of assigning peaks to specific molecules, it has not been clear how to interpret cellular Raman spectra. Here, we provide firm evidence that cellular Raman spectra and transcriptomic profiles of Schizosaccharomyces pombe and Escherichia coli can be computationally connected and thus interpreted. We find that the dimensions of high-dimensional Raman spectra and transcriptomes measured by RNA sequencing can be reduced and connected linearly through a shared low-dimensional subspace. Accordingly, we were able to predict global gene expression profiles by applying the calculated transformation matrix to Raman spectra, and vice versa. Highly expressed non-coding RNAs contributed to the Raman-transcriptome linear correspondence more significantly than mRNAs in S. pombe. This demonstration of correspondence between cellular Raman spectra and transcriptomes is a promising step toward establishing spectroscopic live-cell omics studies. Copyright © 2018 Elsevier Inc. All rights reserved.

Dual Transcriptome Profiling of Leishmania-Infected Human Macrophages Reveals Distinct Reprogramming Signatures

PubMed Central

Fernandes, Maria Cecilia; Dillon, Laura A. L.; Belew, Ashton Trey; Bravo, Hector Corrada; Mosser, David M.

2016-01-01

ABSTRACT Macrophages are mononuclear phagocytes that constitute a first line of defense against pathogens. While lethal to many microbes, they are the primary host cells of Leishmania spp. parasites, the obligate intracellular pathogens that cause leishmaniasis. We conducted transcriptomic profiling of two Leishmania species and the human macrophage over the course of intracellular infection by using high-throughput RNA sequencing to characterize the global gene expression changes and reprogramming events that underlie the interactions between the pathogen and its host. A systematic exclusion of the generic effects of large-particle phagocytosis revealed a vigorous, parasite-specific response of the human macrophage early in the infection that was greatly tempered at later time points. An analogous temporal expression pattern was observed with the parasite, suggesting that much of the reprogramming that occurs as parasites transform into intracellular forms generally stabilizes shortly after entry. Following that, the parasite establishes an intracellular niche within macrophages, with minimal communication between the parasite and the host cell later during the infection. No significant difference was observed between parasite species transcriptomes or in the transcriptional response of macrophages infected with each species. Our comparative analysis of gene expression changes that occur as mouse and human macrophages are infected by Leishmania spp. points toward a general signature of the Leishmania-macrophage infectome. PMID:27165796

Discovering Functions of Unannotated Genes from a Transcriptome Survey of Wild Fungal Isolates

PubMed Central

Ellison, Christopher E.; Kowbel, David; Glass, N. Louise; Taylor, John W.

2014-01-01

ABSTRACT Most fungal genomes are poorly annotated, and many fungal traits of industrial and biomedical relevance are not well suited to classical genetic screens. Assigning genes to phenotypes on a genomic scale thus remains an urgent need in the field. We developed an approach to infer gene function from expression profiles of wild fungal isolates, and we applied our strategy to the filamentous fungus Neurospora crassa. Using transcriptome measurements in 70 strains from two well-defined clades of this microbe, we first identified 2,247 cases in which the expression of an unannotated gene rose and fell across N. crassa strains in parallel with the expression of well-characterized genes. We then used image analysis of hyphal morphologies, quantitative growth assays, and expression profiling to test the functions of four genes predicted from our population analyses. The results revealed two factors that influenced regulation of metabolism of nonpreferred carbon and nitrogen sources, a gene that governed hyphal architecture, and a gene that mediated amino acid starvation resistance. These findings validate the power of our population-transcriptomic approach for inference of novel gene function, and we suggest that this strategy will be of broad utility for genome-scale annotation in many fungal systems. PMID:24692637

Characterization of the transcriptome profiles related to globin gene switching during in vitro erythroid maturation

PubMed Central

2012-01-01

Background The fetal and adult globin genes in the human β-globin cluster on chromosome 11 are sequentially expressed to achieve normal hemoglobin switching during human development. The pharmacological induction of fetal γ-globin (HBG) to replace abnormal adult sickle βS-globin is a successful strategy to treat sickle cell disease; however the molecular mechanism of γ-gene silencing after birth is not fully understood. Therefore, we performed global gene expression profiling using primary erythroid progenitors grown from human peripheral blood mononuclear cells to characterize gene expression patterns during the γ-globin to β-globin (γ/β) switch observed throughout in vitro erythroid differentiation. Results We confirmed erythroid maturation in our culture system using cell morphologic features defined by Giemsa staining and the γ/β-globin switch by reverse transcription-quantitative PCR (RT-qPCR) analysis. We observed maximal γ-globin expression at day 7 with a switch to a predominance of β-globin expression by day 28 and the γ/β-globin switch occurred around day 21. Expression patterns for transcription factors including GATA1, GATA2, KLF1 and NFE2 confirmed our system produced the expected pattern of expression based on the known function of these factors in globin gene regulation. Subsequent gene expression profiling was performed with RNA isolated from progenitors harvested at day 7, 14, 21, and 28 in culture. Three major gene profiles were generated by Principal Component Analysis (PCA). For profile-1 genes, where expression decreased from day 7 to day 28, we identified 2,102 genes down-regulated > 1.5-fold. Ingenuity pathway analysis (IPA) for profile-1 genes demonstrated involvement of the Cdc42, phospholipase C, NF-Kβ, Interleukin-4, and p38 mitogen activated protein kinase (MAPK) signaling pathways. Transcription factors known to be involved in γ-and β-globin regulation were identified. The same approach was used to generate profile-2 genes where expression was up-regulated over 28 days in culture. IPA for the 2,437 genes with > 1.5-fold induction identified the mitotic roles of polo-like kinase, aryl hydrocarbon receptor, cell cycle control, and ATM (Ataxia Telangiectasia Mutated Protein) signaling pathways; transcription factors identified included KLF1, GATA1 and NFE2 among others. Finally, profile-3 was generated from 1,579 genes with maximal expression at day 21, around the time of the γ/β-globin switch. IPA identified associations with cell cycle control, ATM, and aryl hydrocarbon receptor signaling pathways. Conclusions The transcriptome analysis completed with erythroid progenitors grown in vitro identified groups of genes with distinct expression profiles, which function in metabolic pathways associated with cell survival, hematopoiesis, blood cells activation, and inflammatory responses. This study represents the first report of a transcriptome analysis in human primary erythroid progenitors to identify transcription factors involved in hemoglobin switching. Our results also demonstrate that the in vitro liquid culture system is an excellent model to define mechanisms of global gene expression and the DNA-binding protein and signaling pathways involved in globin gene regulation. PMID:22537182

Transcriptome assembly, profiling and differential gene expression analysis of the halophyte Suaeda fruticosa provides insights into salt tolerance.

PubMed

Diray-Arce, Joann; Clement, Mark; Gul, Bilquees; Khan, M Ajmal; Nielsen, Brent L

2015-05-06

Improvement of crop production is needed to feed the growing world population as the amount and quality of agricultural land decreases and soil salinity increases. This has stimulated research on salt tolerance in plants. Most crops tolerate a limited amount of salt to survive and produce biomass, while halophytes (salt-tolerant plants) have the ability to grow with saline water utilizing specific biochemical mechanisms. However, little is known about the genes involved in salt tolerance. We have characterized the transcriptome of Suaeda fruticosa, a halophyte that has the ability to sequester salts in its leaves. Suaeda fruticosa is an annual shrub in the family Chenopodiaceae found in coastal and inland regions of Pakistan and Mediterranean shores. This plant is an obligate halophyte that grows optimally from 200-400 mM NaCl and can grow at up to 1000 mM NaCl. High throughput sequencing technology was performed to provide understanding of genes involved in the salt tolerance mechanism. De novo assembly of the transcriptome and analysis has allowed identification of differentially expressed and unique genes present in this non-conventional crop. Twelve sequencing libraries prepared from control (0 mM NaCl treated) and optimum (300 mM NaCl treated) plants were sequenced using Illumina Hiseq 2000 to investigate differential gene expression between shoots and roots of Suaeda fruticosa. The transcriptome was assembled de novo using Velvet and Oases k-45 and clustered using CDHIT-EST. There are 54,526 unigenes; among these 475 genes are downregulated and 44 are upregulated when samples from plants grown under optimal salt are compared with those grown without salt. BLAST analysis identified the differentially expressed genes, which were categorized in gene ontology terms and their pathways. This work has identified potential genes involved in salt tolerance in Suaeda fruticosa, and has provided an outline of tools to use for de novo transcriptome analysis. The assemblies that were used provide coverage of a considerable proportion of the transcriptome, which allows analysis of differential gene expression and identification of genes that may be involved in salt tolerance. The transcriptome may serve as a reference sequence for study of other succulent halophytes.

Integrated analysis of whole-exome sequencing and transcriptome profiling in males with autism spectrum disorders.

PubMed

Codina-Solà, Marta; Rodríguez-Santiago, Benjamín; Homs, Aïda; Santoyo, Javier; Rigau, Maria; Aznar-Laín, Gemma; Del Campo, Miguel; Gener, Blanca; Gabau, Elisabeth; Botella, María Pilar; Gutiérrez-Arumí, Armand; Antiñolo, Guillermo; Pérez-Jurado, Luis Alberto; Cuscó, Ivon

2015-01-01

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders with high heritability. Recent findings support a highly heterogeneous and complex genetic etiology including rare de novo and inherited mutations or chromosomal rearrangements as well as double or multiple hits. We performed whole-exome sequencing (WES) and blood cell transcriptome by RNAseq in a subset of male patients with idiopathic ASD (n = 36) in order to identify causative genes, transcriptomic alterations, and susceptibility variants. We detected likely monogenic causes in seven cases: five de novo (SCN2A, MED13L, KCNV1, CUL3, and PTEN) and two inherited X-linked variants (MAOA and CDKL5). Transcriptomic analyses allowed the identification of intronic causative mutations missed by the usual filtering of WES and revealed functional consequences of some rare mutations. These included aberrant transcripts (PTEN, POLR3C), deregulated expression in 1.7% of mutated genes (that is, SEMA6B, MECP2, ANK3, CREBBP), allele-specific expression (FUS, MTOR, TAF1C), and non-sense-mediated decay (RIT1, ALG9). The analysis of rare inherited variants showed enrichment in relevant pathways such as the PI3K-Akt signaling and the axon guidance. Integrative analysis of WES and blood RNAseq data has proven to be an efficient strategy to identify likely monogenic forms of ASD (19% in our cohort), as well as additional rare inherited mutations that can contribute to ASD risk in a multifactorial manner. Blood transcriptomic data, besides validating 88% of expressed variants, allowed the identification of missed intronic mutations and revealed functional correlations of genetic variants, including changes in splicing, expression levels, and allelic expression.

Comparative Transcriptome Analysis of the Accessory Sex Gland and Testis from the Chinese Mitten Crab (Eriocheir sinensis)

PubMed Central

He, Lin; Jiang, Hui; Cao, Dandan; Liu, Lihua; Hu, Songnian; Wang, Qun

2013-01-01

The accessory sex gland (ASG) is an important component of the male reproductive system, which functions to enhance the fertility of spermatozoa during male reproduction. Certain proteins secreted by the ASG are known to bind to the spermatozoa membrane and affect its function. The ASG gene expression profile in Chinese mitten crab (Eriocheir sinensis) has not been extensively studied, and limited genetic research has been conducted on this species. The advent of high-throughput sequencing technologies enables the generation of genomic resources within a short period of time and at minimal cost. In the present study, we performed de novo transcriptome sequencing to produce a comprehensive transcript dataset for the ASG of E. sinensis using Illumina sequencing technology. This analysis yielded a total of 33,221,284 sequencing reads, including 2.6 Gb of total nucleotides. Reads were assembled into 85,913 contigs (average 218 bp), or 58,567 scaffold sequences (average 292 bp), that identified 37,955 unigenes (average 385 bp). We assembled all unigenes and compared them with the published testis transcriptome from E. sinensis. In order to identify which genes may be involved in ASG function, as it pertains to modification of spermatozoa, we compared the ASG and testis transcriptome of E. sinensis. Our analysis identified specific genes with both higher and lower tissue expression levels in the two tissues, and the functions of these genes were analyzed to elucidate their potential roles during maturation of spermatozoa. Availability of detailed transcriptome data from ASG and testis in E. sinensis can assist our understanding of the molecular mechanisms involved with spermatozoa conservation, transport, maturation and capacitation and potentially acrosome activation. PMID:23342039

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schwender, Jorg; Konig, Christina; Klapperstuck, Matthias

An attempt has been made to define the extent to which metabolic flux in central plant metabolism is reflected by changes in the transcriptome and metabolome, based on an analysis of in vitro cultured immature embryos of two oilseed rape (Brassica napus) accessions which contrast for seed lipid accumulation. Metabolic flux analysis (MFA) was used to constrain a flux balance metabolic model which included 671 biochemical and transport reactions within the central metabolism. This highly confident flux information was eventually used for comparative analysis of flux vs. transcript (metabolite). Metabolite profiling succeeded in identifying 79 intermediates within the central metabolism,more » some of which differed quantitatively between the two accessions and displayed a significant shift corresponding to flux. An RNA-Seq based transcriptome analysis revealed a large number of genes which were differentially transcribed in the two accessions, including some enzymes/proteins active in major metabolic pathways. With a few exceptions, differential activity in the major pathways (glycolysis, TCA cycle, amino acid, and fatty acid synthesis) was not reflected in contrasting abundances of the relevant transcripts. The conclusion was that transcript abundance on its own cannot be used to infer metabolic activity/fluxes in central plant metabolism. Lastly, this limitation needs to be borne in mind in evaluating transcriptome data and designing metabolic engineering experiments.« less

Integrated Quantitative Transcriptome Maps of Human Trisomy 21 Tissues and Cells

PubMed Central

Pelleri, Maria Chiara; Cattani, Chiara; Vitale, Lorenza; Antonaros, Francesca; Strippoli, Pierluigi; Locatelli, Chiara; Cocchi, Guido; Piovesan, Allison; Caracausi, Maria

2018-01-01

Down syndrome (DS) is due to the presence of an extra full or partial chromosome 21 (Hsa21). The identification of genes contributing to DS pathogenesis could be the key to any rational therapy of the associated intellectual disability. We aim at generating quantitative transcriptome maps in DS integrating all gene expression profile datasets available for any cell type or tissue, to obtain a complete model of the transcriptome in terms of both expression values for each gene and segmental trend of gene expression along each chromosome. We used the TRAM (Transcriptome Mapper) software for this meta-analysis, comparing transcript expression levels and profiles between DS and normal brain, lymphoblastoid cell lines, blood cells, fibroblasts, thymus and induced pluripotent stem cells, respectively. TRAM combined, normalized, and integrated datasets from different sources and across diverse experimental platforms. The main output was a linear expression value that may be used as a reference for each of up to 37,181 mapped transcripts analyzed, related to both known genes and expression sequence tag (EST) clusters. An independent example in vitro validation of fibroblast transcriptome map data was performed through “Real-Time” reverse transcription polymerase chain reaction showing an excellent correlation coefficient (r = 0.93, p < 0.0001) with data obtained in silico. The availability of linear expression values for each gene allowed the testing of the gene dosage hypothesis of the expected 3:2 DS/normal ratio for Hsa21 as well as other human genes in DS, in addition to listing genes differentially expressed with statistical significance. Although a fraction of Hsa21 genes escapes dosage effects, Hsa21 genes are selectively over-expressed in DS samples compared to genes from other chromosomes, reflecting a decisive role in the pathogenesis of the syndrome. Finally, the analysis of chromosomal segments reveals a high prevalence of Hsa21 over-expressed segments over the other genomic regions, suggesting, in particular, a specific region on Hsa21 that appears to be frequently over-expressed (21q22). Our complete datasets are released as a new framework to investigate transcription in DS for individual genes as well as chromosomal segments in different cell types and tissues. PMID:29740474

The technology and biology of single-cell RNA sequencing.

PubMed

Kolodziejczyk, Aleksandra A; Kim, Jong Kyoung; Svensson, Valentine; Marioni, John C; Teichmann, Sarah A

2015-05-21

The differences between individual cells can have profound functional consequences, in both unicellular and multicellular organisms. Recently developed single-cell mRNA-sequencing methods enable unbiased, high-throughput, and high-resolution transcriptomic analysis of individual cells. This provides an additional dimension to transcriptomic information relative to traditional methods that profile bulk populations of cells. Already, single-cell RNA-sequencing methods have revealed new biology in terms of the composition of tissues, the dynamics of transcription, and the regulatory relationships between genes. Rapid technological developments at the level of cell capture, phenotyping, molecular biology, and bioinformatics promise an exciting future with numerous biological and medical applications. Copyright © 2015 Elsevier Inc. All rights reserved.

Digital transcriptome profiling using selective hexamer priming for cDNA synthesis.

PubMed

Armour, Christopher D; Castle, John C; Chen, Ronghua; Babak, Tomas; Loerch, Patrick; Jackson, Stuart; Shah, Jyoti K; Dey, John; Rohl, Carol A; Johnson, Jason M; Raymond, Christopher K

2009-09-01

We developed a procedure for the preparation of whole transcriptome cDNA libraries depleted of ribosomal RNA from only 1 microg of total RNA. The method relies on a collection of short, computationally selected oligonucleotides, called 'not-so-random' (NSR) primers, to obtain full-length, strand-specific representation of nonribosomal RNA transcripts. In this study we validated the technique by profiling human whole brain and universal human reference RNA using ultra-high-throughput sequencing.

Comparative transcriptome response in swine tracheobronchial lymph nodes to viral infection

USDA-ARS?s Scientific Manuscript database

The tracheobronchial lymph node (TBLN) transcriptome response was evaluated following viral infection using Digital Gene Expression Tag Profiling (DGETP). Pigs were sham-treated or infected intranasally with porcine reproductive and respiratory syndrome virus, porcine circovirus type 2, pseudorabies...

Potential effect of exercise in ameliorating insulin resistance at transcriptome level.

PubMed

Hu, Zhigang; Zhou, Lei; He, Tingting

2017-10-24

Insulin resistance can lead to the pathogenesis of type 2 diabetes and exercise can increase insulin sensitivity. And different exercises may have different influences on the mitigation of insulin resistance. It's still unclear how exercise affects inherited insulin resistance at transcriptome level. The purpose of our study was to analyze the potential effects of exercise in ameliorating insulin resistance at transcriptome level. Herein, we analyzed two skeletal muscle transcriptome profiles, including gene profiles between inherited insulin resistant patients and matched healthy controls, and between trained and sedentary subjects (young and old subjects, respectively). Analysis of differentially expressed genes revealed that 12 genes (SGK1, LOC101929876, MYL5, COL6A3, MLF1, LUM, MSTN, COL1A2, COL3A1, IL32, IRS2 and ID1) associated with insulin resistance were reversed by exercise in young subjects, while six genes (MSTN, CFHR1, PFKFB3, IL32, RGCC and NMRK2) were identified in old subjects, suggesting that those genes play potential roles in insulin resistance response to exercise. In addition, we observed that two insulin resistance-related genes, MSTN and IL32, were identified in muscle cells of both young and old subjects, indicating their important roles in the mechanisms behind the beneficial effects of exercise on humans with inherited insulin resistance. Several pathways were also identified, such as "collagen metabolic process", "focal adhesion" and "negative regulation of myoblast differentiation". Taken together, our findings provide novel markers in insulin resistant patients and exercise, and some valuable information for future functional studies on how exercise ameliorating insulin resistance.

Global Transcriptome and Deletome Profiles of Yeast Exposed to Transition Metals

PubMed Central

Jin, Yong Hwan; Dunlap, Paul E.; McBride, Sandra J.; Al-Refai, Hanan; Bushel, Pierre R.; Freedman, Jonathan H.

2008-01-01

A variety of pathologies are associated with exposure to supraphysiological concentrations of essential metals and to non-essential metals and metalloids. The molecular mechanisms linking metal exposure to human pathologies have not been clearly defined. To address these gaps in our understanding of the molecular biology of transition metals, the genomic effects of exposure to Group IB (copper, silver), IIB (zinc, cadmium, mercury), VIA (chromium), and VB (arsenic) elements on the yeast Saccharomyces cerevisiae were examined. Two comprehensive sets of metal-responsive genomic profiles were generated following exposure to equi-toxic concentrations of metal: one that provides information on the transcriptional changes associated with metal exposure (transcriptome), and a second that provides information on the relationship between the expression of ∼4,700 non-essential genes and sensitivity to metal exposure (deletome). Approximately 22% of the genome was affected by exposure to at least one metal. Principal component and cluster analyses suggest that the chemical properties of the metal are major determinants in defining the expression profile. Furthermore, cells may have developed common or convergent regulatory mechanisms to accommodate metal exposure. The transcriptome and deletome had 22 genes in common, however, comparison between Gene Ontology biological processes for the two gene sets revealed that metal stress adaptation and detoxification categories were commonly enriched. Analysis of the transcriptome and deletome identified several evolutionarily conserved, signal transduction pathways that may be involved in regulating the responses to metal exposure. In this study, we identified genes and cognate signaling pathways that respond to exposure to essential and non-essential metals. In addition, genes that are essential for survival in the presence of these metals were identified. This information will contribute to our understanding of the molecular mechanism by which organisms respond to metal stress, and could lead to an understanding of the connection between environmental stress and signal transduction pathways. PMID:18437200

Transcriptome Analysis of Capsicum Chlorosis Virus-Induced Hypersensitive Resistance Response in Bell Capsicum.

PubMed

Widana Gamage, Shirani M K; McGrath, Desmond J; Persley, Denis M; Dietzgen, Ralf G

2016-01-01

Capsicum chlorosis virus (CaCV) is an emerging pathogen of capsicum, tomato and peanut crops in Australia and South-East Asia. Commercial capsicum cultivars with CaCV resistance are not yet available, but CaCV resistance identified in Capsicum chinense is being introgressed into commercial Bell capsicum. However, our knowledge of the molecular mechanisms leading to the resistance response to CaCV infection is limited. Therefore, transcriptome and expression profiling data provide an important resource to better understand CaCV resistance mechanisms. We assembled capsicum transcriptomes and analysed gene expression using Illumina HiSeq platform combined with a tag-based digital gene expression system. Total RNA extracted from CaCV/mock inoculated CaCV resistant (R) and susceptible (S) capsicum at the time point when R line showed a strong hypersensitive response to CaCV infection was used in transcriptome assembly. Gene expression profiles of R and S capsicum in CaCV- and buffer-inoculated conditions were compared. None of the genes were differentially expressed (DE) between R and S cultivars when mock-inoculated, while 2484 genes were DE when inoculated with CaCV. Functional classification revealed that the most highly up-regulated DE genes in R capsicum included pathogenesis-related genes, cell death-associated genes, genes associated with hormone-mediated signalling pathways and genes encoding enzymes involved in synthesis of defense-related secondary metabolites. We selected 15 genes to confirm DE expression levels by real-time quantitative PCR. DE transcript profiling data provided comprehensive gene expression information to gain an understanding of the underlying CaCV resistance mechanisms. Further, we identified candidate CaCV resistance genes in the CaCV-resistant C. annuum x C. chinense breeding line. This knowledge will be useful in future for fine mapping of the CaCV resistance locus and potential genetic engineering of resistance into CaCV-susceptible crops.

Transcriptome Analysis of Capsicum Chlorosis Virus-Induced Hypersensitive Resistance Response in Bell Capsicum

PubMed Central

Widana Gamage, Shirani M. K.; McGrath, Desmond J.; Persley, Denis M.

2016-01-01

Background Capsicum chlorosis virus (CaCV) is an emerging pathogen of capsicum, tomato and peanut crops in Australia and South-East Asia. Commercial capsicum cultivars with CaCV resistance are not yet available, but CaCV resistance identified in Capsicum chinense is being introgressed into commercial Bell capsicum. However, our knowledge of the molecular mechanisms leading to the resistance response to CaCV infection is limited. Therefore, transcriptome and expression profiling data provide an important resource to better understand CaCV resistance mechanisms. Methodology/Principal Findings We assembled capsicum transcriptomes and analysed gene expression using Illumina HiSeq platform combined with a tag-based digital gene expression system. Total RNA extracted from CaCV/mock inoculated CaCV resistant (R) and susceptible (S) capsicum at the time point when R line showed a strong hypersensitive response to CaCV infection was used in transcriptome assembly. Gene expression profiles of R and S capsicum in CaCV- and buffer-inoculated conditions were compared. None of the genes were differentially expressed (DE) between R and S cultivars when mock-inoculated, while 2484 genes were DE when inoculated with CaCV. Functional classification revealed that the most highly up-regulated DE genes in R capsicum included pathogenesis-related genes, cell death-associated genes, genes associated with hormone-mediated signalling pathways and genes encoding enzymes involved in synthesis of defense-related secondary metabolites. We selected 15 genes to confirm DE expression levels by real-time quantitative PCR. Conclusion/Significance DE transcript profiling data provided comprehensive gene expression information to gain an understanding of the underlying CaCV resistance mechanisms. Further, we identified candidate CaCV resistance genes in the CaCV-resistant C. annuum x C. chinense breeding line. This knowledge will be useful in future for fine mapping of the CaCV resistance locus and potential genetic engineering of resistance into CaCV-susceptible crops. PMID:27398596

Loss of stomach, loss of appetite? Sequencing of the ballan wrasse (Labrus bergylta) genome and intestinal transcriptomic profiling illuminate the evolution of loss of stomach function in fish.

PubMed

Lie, Kai K; Tørresen, Ole K; Solbakken, Monica Hongrø; Rønnestad, Ivar; Tooming-Klunderud, Ave; Nederbragt, Alexander J; Jentoft, Sissel; Sæle, Øystein

2018-03-06

The ballan wrasse (Labrus bergylta) belongs to a large teleost family containing more than 600 species showing several unique evolutionary traits such as lack of stomach and hermaphroditism. Agastric fish are found throughout the teleost phylogeny, in quite diverse and unrelated lineages, indicating stomach loss has occurred independently multiple times in the course of evolution. By assembling the ballan wrasse genome and transcriptome we aimed to determine the genetic basis for its digestive system function and appetite regulation. Among other, this knowledge will aid the formulation of aquaculture diets that meet the nutritional needs of agastric species. Long and short read sequencing technologies were combined to generate a ballan wrasse genome of 805 Mbp. Analysis of the genome and transcriptome assemblies confirmed the absence of genes that code for proteins involved in gastric function. The gene coding for the appetite stimulating protein ghrelin was also absent in wrasse. Gene synteny mapping identified several appetite-controlling genes and their paralogs previously undescribed in fish. Transcriptome profiling along the length of the intestine found a declining expression gradient from the anterior to the posterior, and a distinct expression profile in the hind gut. We showed gene loss has occurred for all known genes related to stomach function in the ballan wrasse, while the remaining functions of the digestive tract appear intact. The results also show appetite control in ballan wrasse has undergone substantial changes. The loss of ghrelin suggests that other genes, such as motilin, may play a ghrelin like role. The wrasse genome offers novel insight in to the evolutionary traits of this large family. As the stomach plays a major role in protein digestion, the lack of genes related to stomach digestion in wrasse suggests it requires formulated diets with higher levels of readily digestible protein than those for gastric species.

A machine learning approach for the identification of key markers involved in brain development from single-cell transcriptomic data.

PubMed

Hu, Yongli; Hase, Takeshi; Li, Hui Peng; Prabhakar, Shyam; Kitano, Hiroaki; Ng, See Kiong; Ghosh, Samik; Wee, Lawrence Jin Kiat

2016-12-22

The ability to sequence the transcriptomes of single cells using single-cell RNA-seq sequencing technologies presents a shift in the scientific paradigm where scientists, now, are able to concurrently investigate the complex biology of a heterogeneous population of cells, one at a time. However, till date, there has not been a suitable computational methodology for the analysis of such intricate deluge of data, in particular techniques which will aid the identification of the unique transcriptomic profiles difference between the different cellular subtypes. In this paper, we describe the novel methodology for the analysis of single-cell RNA-seq data, obtained from neocortical cells and neural progenitor cells, using machine learning algorithms (Support Vector machine (SVM) and Random Forest (RF)). Thirty-eight key transcripts were identified, using the SVM-based recursive feature elimination (SVM-RFE) method of feature selection, to best differentiate developing neocortical cells from neural progenitor cells in the SVM and RF classifiers built. Also, these genes possessed a higher discriminative power (enhanced prediction accuracy) as compared commonly used statistical techniques or geneset-based approaches. Further downstream network reconstruction analysis was carried out to unravel hidden general regulatory networks where novel interactions could be further validated in web-lab experimentation and be useful candidates to be targeted for the treatment of neuronal developmental diseases. This novel approach reported for is able to identify transcripts, with reported neuronal involvement, which optimally differentiate neocortical cells and neural progenitor cells. It is believed to be extensible and applicable to other single-cell RNA-seq expression profiles like that of the study of the cancer progression and treatment within a highly heterogeneous tumour.

Determining the mode of action of anti-mycobacterial C17 diyne natural products using expression profiling: evidence for fatty acid biosynthesis inhibition.

PubMed

Li, Haoxin; Cowie, Andrew; Johnson, John A; Webster, Duncan; Martyniuk, Christopher J; Gray, Christopher A

2016-08-11

The treatment of microbial infections is becoming increasingly challenging because of limited therapeutic options and the growing number of pathogenic strains that are resistant to current antibiotics. There is an urgent need to identify molecules with novel modes of action to facilitate the development of new and more effective therapeutic agents. The anti-mycobacterial activity of the C17 diyne natural products falcarinol and panaxydol has been described previously; however, their mode of action remains largely undetermined in microbes. Gene expression profiling was therefore used to determine the transcriptomic response of Mycobacterium smegmatis upon treatment with falcarinol and panaxydol to better characterize the mode of action of these C17 diynes. Our analyses identified 704 and 907 transcripts that were differentially expressed in M. smegmatis after treatment with falcarinol and panaxydol respectively. Principal component analysis suggested that the C17 diynes exhibit a mode of action that is distinct to commonly used antimycobacterial drugs. Functional enrichment analysis and pathway enrichment analysis revealed that cell processes such as ectoine biosynthesis and cyclopropane-fatty-acyl-phospholipid synthesis were responsive to falcarinol and panaxydol treatment at the transcriptome level in M. smegmatis. The modes of action of the two C17 diynes were also predicted through Prediction of Activity Spectra of Substances (PASS). Based upon convergence of these three independent analyses, we hypothesize that the C17 diynes inhibit fatty acid biosynthesis, specifically phospholipid synthesis, in mycobacteria. Based on transcriptomic responses, it is suggested that the C17 diynes act differently than other anti-mycobacterial compounds in M. smegmatis, and do so by inhibiting phospholipid biosynthesis.

Sequencing and de novo analysis of the hemocytes transcriptome in Litopenaeus vannamei response to white spot syndrome virus infection.

PubMed

Xue, Shuxia; Liu, Yichen; Zhang, Yichen; Sun, Yan; Geng, Xuyun; Sun, Jinsheng

2013-01-01

White spot syndrome virus (WSSV) is a causative pathogen found in most shrimp farming areas of the world and causes large economic losses to the shrimp aquaculture. The mechanism underlying the molecular pathogenesis of the highly virulent WSSV remains unknown. To better understand the virus-host interactions at the molecular level, the transcriptome profiles in hemocytes of unchallenged and WSSV-challenged shrimp (Litopenaeus vannamei) were compared using a short-read deep sequencing method (Illumina). RNA-seq analysis generated more than 25.81 million clean pair end (PE) reads, which were assembled into 52,073 unigenes (mean size = 520 bp). Based on sequence similarity searches, 23,568 (45.3%) genes were identified, among which 6,562 and 7,822 unigenes were assigned to gene ontology (GO) categories and clusters of orthologous groups (COG), respectively. Searches in the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG) mapped 14,941 (63.4%) unigenes to 240 KEGG pathways. Among all the annotated unigenes, 1,179 were associated with immune-related genes. Digital gene expression (DGE) analysis revealed that the host transcriptome profile was slightly changed in the early infection (5 hours post injection) of the virus, while large transcriptional differences were identified in the late infection (48 hpi) of WSSV. The differentially expressed genes mainly involved in pattern recognition genes and some immune response factors. The results indicated that antiviral immune mechanisms were probably involved in the recognition of pathogen-associated molecular patterns. This study provided a global survey of host gene activities against virus infection in a non-model organism, pacific white shrimp. Results can contribute to the in-depth study of candidate genes in white shrimp, and help to improve the current understanding of host-pathogen interactions.

De novo transcriptome sequencing and digital gene expression analysis predict biosynthetic pathway of rhynchophylline and isorhynchophylline from Uncaria rhynchophylla, a non-model plant with potent anti-alzheimer's properties.

PubMed

Guo, Qianqian; Ma, Xiaojun; Wei, Shugen; Qiu, Deyou; Wilson, Iain W; Wu, Peng; Tang, Qi; Liu, Lijun; Dong, Shoukun; Zu, Wei

2014-08-12

The major medicinal alkaloids isolated from Uncaria rhynchophylla (gouteng in chinese) capsules are rhynchophylline (RIN) and isorhynchophylline (IRN). Extracts containing these terpene indole alkaloids (TIAs) can inhibit the formation and destabilize preformed fibrils of amyloid β protein (a pathological marker of Alzheimer's disease), and have been shown to improve the cognitive function of mice with Alzheimer-like symptoms. The biosynthetic pathways of RIN and IRN are largely unknown. In this study, RNA-sequencing of pooled Uncaria capsules RNA samples taken at three developmental stages that accumulate different amount of RIN and IRN was performed. More than 50 million high-quality reads from a cDNA library were generated and de novo assembled. Sequences for all of the known enzymes involved in TIAs synthesis were identified. Additionally, 193 cytochrome P450 (CYP450), 280 methyltransferase and 144 isomerase genes were identified, that are potential candidates for enzymes involved in RIN and IRN synthesis. Digital gene expression profile (DGE) analysis was performed on the three capsule developmental stages, and based on genes possessing expression profiles consistent with RIN and IRN levels; four CYP450s, three methyltransferases and three isomerases were identified as the candidates most likely to be involved in the later steps of RIN and IRN biosynthesis. A combination of de novo transcriptome assembly and DGE analysis was shown to be a powerful method for identifying genes encoding enzymes potentially involved in the biosynthesis of important secondary metabolites in a non-model plant. The transcriptome data from this study provides an important resource for understanding the formation of major bioactive constituents in the capsule extract from Uncaria, and provides information that may aid in metabolic engineering to increase yields of these important alkaloids.

The developmental transcriptome atlas of the spoon worm Urechis unicinctus (Echiurida: Annelida).

PubMed

Park, Chungoo; Han, Yong-Hee; Lee, Sung-Gwon; Ry, Kyoung-Bin; Oh, Jooseong; Kern, Elizabeth M A; Park, Joong-Ki; Cho, Sung-Jin

2018-03-01

Echiurida is one of the most intriguing major subgroups of annelida because, unlike most other annelids, echiurids lack metameric body segmentation as adults. For this reason, transcriptome analyses from various developmental stages of echiurid species can be of substantial value for understanding precise expression levels and the complex regulatory networks during early and larval development. A total of 914 million raw RNA-Seq reads were produced from 14 developmental stages of Urechis unicinctus and were de novo assembled into contigs spanning 63,928,225 bp with an N50 length of 2700 bp. The resulting comprehensive transcriptome database of the early developmental stages of U. unicinctus consists of 20,305 representative functional protein-coding transcripts. Approximately 66% of unigenes were assigned to superphylum-level taxa, including Lophotrochozoa (40%). The completeness of the transcriptome assembly was assessed using benchmarking universal single-copy orthologs; 75.7% of the single-copy orthologs were presented in our transcriptome database. We observed 3 distinct patterns of global transcriptome profiles from 14 developmental stages and identified 12,705 genes that showed dynamic regulation patterns during the differentiation and maturation of U. unicinctus cells. We present the first large-scale developmental transcriptome dataset of U. unicinctus and provide a general overview of the dynamics of global gene expression changes during its early developmental stages. The analysis of time-course gene expression data is a first step toward understanding the complex developmental gene regulatory networks in U. unicinctus and will furnish a valuable resource for analyzing the functions of gene repertoires in various developmental phases.

Next-generation sequencing (NGS) transcriptomes reveal association of multiple genes and pathways contributing to secondary metabolites accumulation in tuberous roots of Aconitum heterophyllum Wall.

PubMed

Pal, Tarun; Malhotra, Nikhil; Chanumolu, Sree Krishna; Chauhan, Rajinder Singh

2015-07-01

The transcriptomes of Aconitum heterophyllum were assembled and characterized for the first time to decipher molecular components contributing to biosynthesis and accumulation of metabolites in tuberous roots. Aconitum heterophyllum Wall., popularly known as Atis, is a high-value medicinal herb of North-Western Himalayas. No information exists as of today on genetic factors contributing to the biosynthesis of secondary metabolites accumulating in tuberous roots, thereby, limiting genetic interventions towards genetic improvement of A. heterophyllum. Illumina paired-end sequencing followed by de novo assembly yielded 75,548 transcripts for root transcriptome and 39,100 transcripts for shoot transcriptome with minimum length of 200 bp. Biological role analysis of root versus shoot transcriptomes assigned 27,596 and 16,604 root transcripts; 12,340 and 9398 shoot transcripts into gene ontology and clusters of orthologous group, respectively. KEGG pathway mapping assigned 37 and 31 transcripts onto starch-sucrose metabolism while 329 and 341 KEGG orthologies associated with transcripts were found to be involved in biosynthesis of various secondary metabolites for root and shoot transcriptomes, respectively. In silico expression profiling of the mevalonate/2-C-methyl-D-erythritol 4-phosphate (non-mevalonate) pathway genes for aconites biosynthesis revealed 4 genes HMGR (3-hydroxy-3-methylglutaryl-CoA reductase), MVK (mevalonate kinase), MVDD (mevalonate diphosphate decarboxylase) and HDS (1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate synthase) with higher expression in root transcriptome compared to shoot transcriptome suggesting their key role in biosynthesis of aconite alkaloids. Five genes, GMPase (geranyl diphosphate mannose pyrophosphorylase), SHAGGY, RBX1 (RING-box protein 1), SRF receptor kinases and β-amylase, implicated in tuberous root formation in other plant species showed higher levels of expression in tuberous roots compared to shoots. A total of 15,487 transcription factors belonging to bHLH, MYB, bZIP families and 399 ABC transporters which regulate biosynthesis and accumulation of bioactive compounds were identified in root and shoot transcriptomes. The expression of 5 ABC transporters involved in tuberous root development was validated by quantitative PCR analysis. Network connectivity diagrams were drawn for starch-sucrose metabolism and isoquinoline alkaloid biosynthesis associated with tuberous root growth and secondary metabolism, respectively, in root transcriptome of A. heterophyllum. The current endeavor will be of practical importance in planning a suitable genetic intervention strategy for the improvement of A. heterophyllum.

Profiling the venom gland transcriptomes of Costa Rican snakes by 454 pyrosequencing

PubMed Central

2011-01-01

Background A long term research goal of venomics, of applied importance for improving current antivenom therapy, but also for drug discovery, is to understand the pharmacological potential of venoms. Individually or combined, proteomic and transcriptomic studies have demonstrated their feasibility to explore in depth the molecular diversity of venoms. In the absence of genome sequence, transcriptomes represent also valuable searchable databases for proteomic projects. Results The venom gland transcriptomes of 8 Costa Rican taxa from 5 genera (Crotalus, Bothrops, Atropoides, Cerrophidion, and Bothriechis) of pitvipers were investigated using high-throughput 454 pyrosequencing. 100,394 out of 330,010 masked reads produced significant hits in the available databases. 5.165,220 nucleotides (8.27%) were masked by RepeatMasker, the vast majority of which corresponding to class I (retroelements) and class II (DNA transposons) mobile elements. BLAST hits included 79,991 matches to entries of the taxonomic suborder Serpentes, of which 62,433 displayed similarity to documented venom proteins. Strong discrepancies between the transcriptome-computed and the proteome-gathered toxin compositions were obvious at first sight. Although the reasons underlaying this discrepancy are elusive, since no clear trend within or between species is apparent, the data indicate that individual mRNA species may be translationally controlled in a species-dependent manner. The minimum number of genes from each toxin family transcribed into the venom gland transcriptome of each species was calculated from multiple alignments of reads matched to a full-length reference sequence of each toxin family. Reads encoding ORF regions of Kazal-type inhibitor-like proteins were uniquely found in Bothriechis schlegelii and B. lateralis transcriptomes, suggesting a genus-specific recruitment event during the early-Middle Miocene. A transcriptome-based cladogram supports the large divergence between A. mexicanus and A. picadoi, and a closer kinship between A. mexicanus and C. godmani. Conclusions Our comparative next-generation sequencing (NGS) analysis reveals taxon-specific trends governing the formulation of the venom arsenal. Knowledge of the venom proteome provides hints on the translation efficiency of toxin-coding transcripts, contributing thereby to a more accurate interpretation of the transcriptome. The application of NGS to the analysis of snake venom transcriptomes, may represent the tool for opening the door to systems venomics. PMID:21605378

Transcriptomic events associated with internal browning of apple during postharvest storage

USDA-ARS?s Scientific Manuscript database

Although apple (Malus x domestica) is of economic importance, little information is available about the transriptomic profiling of postharvest disorders, particularly internal browning. This study aimed to explore changes in the apple transcriptome associated with development of flesh browning durin...

The transcriptome landscape of early maize meiosis

USDA-ARS?s Scientific Manuscript database

Meiosis, particularly meiotic recombination, is a major factor affecting yield and breeding of plants. To gain insight into the transcriptome landscape during early initiation steps of meiotic recombination, we profiled early prophase I meiocytes from maize using RNA-seq. Our analyses of genes prefe...

Picking Cell Lines for High-Throughput Transcriptomic Toxicity Screening (SOT)

EPA Science Inventory

High throughput, whole genome transcriptomic profiling is a promising approach to comprehensively evaluate chemicals for potential biological effects. To be useful for in vitro toxicity screening, gene expression must be quantified in a set of representative cell types that captu...

Transcriptome Analysis and Discovery of Genes Involved in Immune Pathways from Coelomocytes of Sea Cucumber (Apostichopus japonicus) after Vibrio splendidus Challenge.

PubMed

Gao, Qiong; Liao, Meijie; Wang, Yingeng; Li, Bin; Zhang, Zheng; Rong, Xiaojun; Chen, Guiping; Wang, Lan

2015-07-17

Vibrio splendidus is identified as one of the major pathogenic factors for the skin ulceration syndrome in sea cucumber (Apostichopus japonicus), which has vastly limited the development of the sea cucumber culture industry. In order to screen the immune genes involving Vibrio splendidus challenge in sea cucumber and explore the molecular mechanism of this process, the related transcriptome and gene expression profiling of resistant and susceptible biotypes of sea cucumber with Vibrio splendidus challenge were collected for analysis. A total of 319,455,942 trimmed reads were obtained, which were assembled into 186,658 contigs. After that, 89,891 representative contigs (without isoform) were clustered. The analysis of the gene expression profiling identified 358 differentially expression genes (DEGs) in the bacterial-resistant group, and 102 DEGs in the bacterial-susceptible group, compared with that in control group. According to the reported references and annotation information from BLAST, GO and KEGG, 30 putative bacterial-resistant genes and 19 putative bacterial-susceptible genes were identified from DEGs. The qRT-PCR results were consistent with the RNA-Seq results. Furthermore, many DGEs were involved in immune signaling related pathways, such as Endocytosis, Lysosome, MAPK, Chemokine and the ERBB signaling pathway.

Transcriptome Analysis and Discovery of Genes Involved in Immune Pathways from Coelomocytes of Sea Cucumber (Apostichopus japonicus) after Vibrio splendidus Challenge

PubMed Central

Gao, Qiong; Liao, Meijie; Wang, Yingeng; Li, Bin; Zhang, Zheng; Rong, Xiaojun; Chen, Guiping; Wang, Lan

2015-01-01

Vibrio splendidus is identified as one of the major pathogenic factors for the skin ulceration syndrome in sea cucumber (Apostichopus japonicus), which has vastly limited the development of the sea cucumber culture industry. In order to screen the immune genes involving Vibrio splendidus challenge in sea cucumber and explore the molecular mechanism of this process, the related transcriptome and gene expression profiling of resistant and susceptible biotypes of sea cucumber with Vibrio splendidus challenge were collected for analysis. A total of 319,455,942 trimmed reads were obtained, which were assembled into 186,658 contigs. After that, 89,891 representative contigs (without isoform) were clustered. The analysis of the gene expression profiling identified 358 differentially expression genes (DEGs) in the bacterial-resistant group, and 102 DEGs in the bacterial-susceptible group, compared with that in control group. According to the reported references and annotation information from BLAST, GO and KEGG, 30 putative bacterial-resistant genes and 19 putative bacterial-susceptible genes were identified from DEGs. The qRT-PCR results were consistent with the RNA-Seq results. Furthermore, many DGEs were involved in immune signaling related pathways, such as Endocytosis, Lysosome, MAPK, Chemokine and the ERBB signaling pathway. PMID:26193268

Ovary transcriptome profiling via artificial intelligence reveals a transcriptomic fingerprint predicting egg quality in striped bass, Morone saxatilis.

PubMed

Chapman, Robert W; Reading, Benjamin J; Sullivan, Craig V

2014-01-01

Inherited gene transcripts deposited in oocytes direct early embryonic development in all vertebrates, but transcript profiles indicative of embryo developmental competence have not previously been identified. We employed artificial intelligence to model profiles of maternal ovary gene expression and their relationship to egg quality, evaluated as production of viable mid-blastula stage embryos, in the striped bass (Morone saxatilis), a farmed species with serious egg quality problems. In models developed using artificial neural networks (ANNs) and supervised machine learning, collective changes in the expression of a limited suite of genes (233) representing <2% of the queried ovary transcriptome explained >90% of the eventual variance in embryo survival. Egg quality related to minor changes in gene expression (<0.2-fold), with most individual transcripts making a small contribution (<1%) to the overall prediction of egg quality. These findings indicate that the predictive power of the transcriptome as regards egg quality resides not in levels of individual genes, but rather in the collective, coordinated expression of a suite of transcripts constituting a transcriptomic "fingerprint". Correlation analyses of the corresponding candidate genes indicated that dysfunction of the ubiquitin-26S proteasome, COP9 signalosome, and subsequent control of the cell cycle engenders embryonic developmental incompetence. The affected gene networks are centrally involved in regulation of early development in all vertebrates, including humans. By assessing collective levels of the relevant ovarian transcripts via ANNs we were able, for the first time in any vertebrate, to accurately predict the subsequent embryo developmental potential of eggs from individual females. Our results show that the transcriptomic fingerprint evidencing developmental dysfunction is highly predictive of, and therefore likely to regulate, egg quality, a biologically complex trait crucial to reproductive fitness.

Ovary Transcriptome Profiling via Artificial Intelligence Reveals a Transcriptomic Fingerprint Predicting Egg Quality in Striped Bass, Morone saxatilis

PubMed Central

2014-01-01

Inherited gene transcripts deposited in oocytes direct early embryonic development in all vertebrates, but transcript profiles indicative of embryo developmental competence have not previously been identified. We employed artificial intelligence to model profiles of maternal ovary gene expression and their relationship to egg quality, evaluated as production of viable mid-blastula stage embryos, in the striped bass (Morone saxatilis), a farmed species with serious egg quality problems. In models developed using artificial neural networks (ANNs) and supervised machine learning, collective changes in the expression of a limited suite of genes (233) representing <2% of the queried ovary transcriptome explained >90% of the eventual variance in embryo survival. Egg quality related to minor changes in gene expression (<0.2-fold), with most individual transcripts making a small contribution (<1%) to the overall prediction of egg quality. These findings indicate that the predictive power of the transcriptome as regards egg quality resides not in levels of individual genes, but rather in the collective, coordinated expression of a suite of transcripts constituting a transcriptomic “fingerprint”. Correlation analyses of the corresponding candidate genes indicated that dysfunction of the ubiquitin-26S proteasome, COP9 signalosome, and subsequent control of the cell cycle engenders embryonic developmental incompetence. The affected gene networks are centrally involved in regulation of early development in all vertebrates, including humans. By assessing collective levels of the relevant ovarian transcripts via ANNs we were able, for the first time in any vertebrate, to accurately predict the subsequent embryo developmental potential of eggs from individual females. Our results show that the transcriptomic fingerprint evidencing developmental dysfunction is highly predictive of, and therefore likely to regulate, egg quality, a biologically complex trait crucial to reproductive fitness. PMID:24820964

Selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) for direct, versatile and accurate RNA structure analysis.

PubMed

Smola, Matthew J; Rice, Greggory M; Busan, Steven; Siegfried, Nathan A; Weeks, Kevin M

2015-11-01

Selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) chemistries exploit small electrophilic reagents that react with 2'-hydroxyl groups to interrogate RNA structure at single-nucleotide resolution. Mutational profiling (MaP) identifies modified residues by using reverse transcriptase to misread a SHAPE-modified nucleotide and then counting the resulting mutations by massively parallel sequencing. The SHAPE-MaP approach measures the structure of large and transcriptome-wide systems as accurately as can be done for simple model RNAs. This protocol describes the experimental steps, implemented over 3 d, that are required to perform SHAPE probing and to construct multiplexed SHAPE-MaP libraries suitable for deep sequencing. Automated processing of MaP sequencing data is accomplished using two software packages. ShapeMapper converts raw sequencing files into mutational profiles, creates SHAPE reactivity plots and provides useful troubleshooting information. SuperFold uses these data to model RNA secondary structures, identify regions with well-defined structures and visualize probable and alternative helices, often in under 1 d. SHAPE-MaP can be used to make nucleotide-resolution biophysical measurements of individual RNA motifs, rare components of complex RNA ensembles and entire transcriptomes.

Transcriptome profiling analysis of Vibrio vulnificus during human infection.

PubMed

Bisharat, Naiel; Bronstein, Michal; Korner, Mira; Schnitzer, Temima; Koton, Yael

2013-09-01

Vibrio vulnificus is a waterborne pathogen that was responsible for an outbreak of severe soft-tissue infections among fish farmers and fish consumers in Israel. Several factors have been shown to be associated with virulence. However, the transcriptome profile of the pathogen during human infection has not been determined yet. We compared the transcriptome profile, using RNA sequencing, of a human-pathogenic strain harvested directly from tissue of a patient suffering from severe soft-tissue infection with necrotizing fasciitis, with the same strain and three other environmental strains grown in vitro. The five sequenced libraries were aligned to the reference genomes of V. vulnificus strains CMCP6 and YJ016. Approximately 47.8 to 62.3 million paired-end raw reads were generated from the five runs. Nearly 84 % of the genome was covered by reads from at least one of the five runs, suggesting that nearly 16 % of the genome is not transcribed or is transcribed at low levels. We identified 123 genes that were differentially expressed during the acute phase of infection. Sixty-three genes were mapped to the large chromosome, 47 genes mapped to the small chromosome and 13 genes mapped to the YJ016 plasmid. The 123 genes fell into a variety of functional categories including transcription, signal transduction, cell motility, carbohydrate metabolism, intracellular trafficking and cell envelope biogenesis. Among the genes differentially expressed during human infection we identified genes encoding bacterial toxin (RtxA1) and genes involved in flagellar components, Flp-coding region, GGDEF family protein, iron acquisition system and sialic acid metabolism.

Integrated Systems Biology Analysis of Transcriptomes Reveals Candidate Genes for Acidity Control in Developing Fruits of Sweet Orange (Citrus sinensis L. Osbeck).

PubMed

Huang, Dingquan; Zhao, Yihong; Cao, Minghao; Qiao, Liang; Zheng, Zhi-Liang

2016-01-01

Organic acids, such as citrate and malate, are important contributors for the sensory traits of fleshy fruits. Although their biosynthesis has been illustrated, regulatory mechanisms of acid accumulation remain to be dissected. To provide transcriptional architecture and identify candidate genes for citrate accumulation in fruits, we have selected for transcriptome analysis four varieties of sweet orange (Citrus sinensis L. Osbeck) with varying fruit acidity, Succari (acidless), Bingtang (low acid), and Newhall and Xinhui (normal acid). Fruits of these varieties at 45 days post anthesis (DPA), which corresponds to Stage I (cell division), had similar acidity, but they displayed differential acid accumulation at 142 DPA (Stage II, cell expansion). Transcriptomes of fruits at 45 and 142 DPA were profiled using RNA sequencing and analyzed with three different algorithms (Pearson correlation, gene coexpression network and surrogate variable analysis). Our network analysis shows that the acid-correlated genes belong to three distinct network modules. Several of these candidate fruit acidity genes encode regulatory proteins involved in transport (such as AHA10), degradation (such as APD2) and transcription (such as AIL6) and act as hubs in the citrate accumulation gene networks. Taken together, our integrated systems biology analysis has provided new insights into the fruit citrate accumulation gene network and led to the identification of candidate genes likely associated with the fruit acidity control.

Integrated Systems Biology Analysis of Transcriptomes Reveals Candidate Genes for Acidity Control in Developing Fruits of Sweet Orange (Citrus sinensis L. Osbeck)

PubMed Central

Huang, Dingquan; Zhao, Yihong; Cao, Minghao; Qiao, Liang; Zheng, Zhi-Liang

2016-01-01

Organic acids, such as citrate and malate, are important contributors for the sensory traits of fleshy fruits. Although their biosynthesis has been illustrated, regulatory mechanisms of acid accumulation remain to be dissected. To provide transcriptional architecture and identify candidate genes for citrate accumulation in fruits, we have selected for transcriptome analysis four varieties of sweet orange (Citrus sinensis L. Osbeck) with varying fruit acidity, Succari (acidless), Bingtang (low acid), and Newhall and Xinhui (normal acid). Fruits of these varieties at 45 days post anthesis (DPA), which corresponds to Stage I (cell division), had similar acidity, but they displayed differential acid accumulation at 142 DPA (Stage II, cell expansion). Transcriptomes of fruits at 45 and 142 DPA were profiled using RNA sequencing and analyzed with three different algorithms (Pearson correlation, gene coexpression network and surrogate variable analysis). Our network analysis shows that the acid-correlated genes belong to three distinct network modules. Several of these candidate fruit acidity genes encode regulatory proteins involved in transport (such as AHA10), degradation (such as APD2) and transcription (such as AIL6) and act as hubs in the citrate accumulation gene networks. Taken together, our integrated systems biology analysis has provided new insights into the fruit citrate accumulation gene network and led to the identification of candidate genes likely associated with the fruit acidity control. PMID:27092171

Concurrent Host-Pathogen Transcriptional Responses in a Clostridium perfringens Murine Myonecrosis Infection

PubMed Central

2018-01-01

ABSTRACT To obtain an insight into host-pathogen interactions in clostridial myonecrosis, we carried out comparative transcriptome analysis of both the bacterium and the host in a murine Clostridium perfringens infection model, which is the first time that such an investigation has been conducted. Analysis of the host transcriptome from infected muscle tissues indicated that many genes were upregulated compared to the results seen with mock-infected mice. These genes were enriched for host defense pathways, including Toll-like receptor (TLR) and Nod-like receptor (NLR) signaling components. Real-time PCR confirmed that host TLR2 and NLRP3 inflammasome genes were induced in response to C. perfringens infection. Comparison of the transcriptome of C. perfringens cells from the infected tissues with that from broth cultures showed that host selective pressure induced a global change in C. perfringens gene expression. A total of 33% (923) of C. perfringens genes were differentially regulated, including 10 potential virulence genes that were upregulated relative to their expression in vitro. These genes encoded putative proteins that may be involved in the synthesis of cell wall-associated macromolecules, in adhesion to host cells, or in protection from host cationic antimicrobial peptides. This report presents the first successful expression profiling of coregulated transcriptomes of bacterial and host genes during a clostridial myonecrosis infection and provides new insights into disease pathogenesis and host-pathogen interactions. PMID:29588405

Doubled Haploid ‘CUDH2107’ as a Reference for Bulb Onion (Allium cepa L.) Research: Development of a Transcriptome Catalogue and Identification of Transcripts Associated with Male Fertility

PubMed Central

Khosa, Jiffinvir S.; Lee, Robyn; Bräuning, Sophia; Lord, Janice; Pither-Joyce, Meeghan; McCallum, John; Macknight, Richard C.

2016-01-01

Researchers working on model plants have derived great benefit from developing genomic and genetic resources using ‘reference’ genotypes. Onion has a large and highly heterozygous genome making the sharing of germplasm and analysis of sequencing data complicated. To simplify the discovery and analysis of genes underlying important onion traits, we are promoting the use of the homozygous double haploid line ‘CUDH2107’ by the onion research community. In the present investigation, we performed transcriptome sequencing on vegetative and reproductive tissues of CUDH2107 to develop a multi-organ reference transcriptome catalogue. A total of 396 million 100 base pair paired reads was assembled using the Trinity pipeline, resulting in 271,665 transcript contigs. This dataset was analysed for gene ontology and transcripts were classified on the basis of putative biological processes, molecular function and cellular localization. Significant differences were observed in transcript expression profiles between different tissues. To demonstrate the utility of our CUDH2107 transcriptome catalogue for understanding the genetic and molecular basis of various traits, we identified orthologues of rice genes involved in male fertility and flower development. These genes provide an excellent starting point for studying the molecular regulation, and the engineering of reproductive traits. PMID:27861615

Transcriptome profiling in Arabidopsis inflorescence stems grown under hypergravity in terms of cell walls and plant hormones

NASA Astrophysics Data System (ADS)

Tamaoki, D.; Karahara, I.; Nishiuchi, T.; De Oliveira, S.; Schreiber, L.; Wakasugi, T.; Yamada, K.; Yamaguchi, K.; Kamisaka, S.

2009-07-01

Land plants rely on lignified secondary cell walls in supporting their body weight on the Earth. Although gravity influences the formation of the secondary cell walls, the regulatory mechanism of their formation by gravity is not yet understood. We carried out a comprehensive analysis of gene expression in inflorescence stems of Arabidopsis thaliana L. using microarray (22 K) to identify genes whose expression is modulated under hypergravity condition (300 g). Total RNA was isolated from the basal region of inflorescence stems of plants grown for 24 h at 300 g or 1 g. Microarray analysis showed that hypergravity up-regulated the expression of 403 genes to more than 2-fold. Hypergravity up-regulated the genes responsible for the biosynthesis or modification of cell wall components such as lignin, xyloglucan, pectin and structural proteins. In addition, hypergravity altered the expression of genes related to the biosynthesis of plant hormones such as auxin and ethylene and that of genes encoding hormone-responsive proteins. Our transcriptome profiling indicates that hypergravity influences the formation of secondary cell walls by modulating the pattern of gene expression, and that auxin and/or ethylene play an important role in signaling hypergravity stimulus.

Single-Cell Sequencing for Precise Cancer Research: Progress and Prospects.

PubMed

Zhang, Xiaoyan; Marjani, Sadie L; Hu, Zhaoyang; Weissman, Sherman M; Pan, Xinghua; Wu, Shixiu

2016-03-15

Advances in genomic technology have enabled the faithful detection and measurement of mutations and the gene expression profile of cancer cells at the single-cell level. Recently, several single-cell sequencing methods have been developed that permit the comprehensive and precise analysis of the cancer-cell genome, transcriptome, and epigenome. The use of these methods to analyze cancer cells has led to a series of unanticipated discoveries, such as the high heterogeneity and stochastic changes in cancer-cell populations, the new driver mutations and the complicated clonal evolution mechanisms, and the novel identification of biomarkers of variant tumors. These methods and the knowledge gained from their utilization could potentially improve the early detection and monitoring of rare cancer cells, such as circulating tumor cells and disseminated tumor cells, and promote the development of personalized and highly precise cancer therapy. Here, we discuss the current methods for single cancer-cell sequencing, with a strong focus on those practically used or potentially valuable in cancer research, including single-cell isolation, whole genome and transcriptome amplification, epigenome profiling, multi-dimensional sequencing, and next-generation sequencing and analysis. We also examine the current applications, challenges, and prospects of single cancer-cell sequencing. ©2016 American Association for Cancer Research.

Comparative Transcriptome Analysis of Genes Involved in Anthocyanin Biosynthesis in the Red and Yellow Fruits of Sweet Cherry (Prunus avium L.)

PubMed Central

Wei, Hairong; Chen, Xin; Zong, Xiaojuan; Shu, Huairui; Gao, Dongsheng; Liu, Qingzhong

2015-01-01

Background Fruit color is one of the most important economic traits of the sweet cherry (Prunus avium L.). The red coloration of sweet cherry fruit is mainly attributed to anthocyanins. However, limited information is available regarding the molecular mechanisms underlying anthocyanin biosynthesis and its regulation in sweet cherry. Methodology/Principal Findings In this study, a reference transcriptome of P. avium L. was sequenced and annotated to identify the transcriptional determinants of fruit color. Normalized cDNA libraries from red and yellow fruits were sequenced using the next-generation Illumina/Solexa sequencing platform and de novo assembly. Over 66 million high-quality reads were assembled into 43,128 unigenes using a combined assembly strategy. Then a total of 22,452 unigenes were compared to public databases using homology searches, and 20,095 of these unigenes were annotated in the Nr protein database. Furthermore, transcriptome differences between the four stages of fruit ripening were analyzed using Illumina digital gene expression (DGE) profiling. Biological pathway analysis revealed that 72 unigenes were involved in anthocyanin biosynthesis. The expression patterns of unigenes encoding phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavanone 3’-hydroxylase (F3’H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS) and UDP glucose: flavonol 3-O-glucosyltransferase (UFGT) during fruit ripening differed between red and yellow fruit. In addition, we identified some transcription factor families (such as MYB, bHLH and WD40) that may control anthocyanin biosynthesis. We confirmed the altered expression levels of eighteen unigenes that encode anthocyanin biosynthetic enzymes and transcription factors using quantitative real-time PCR (qRT-PCR). Conclusions/Significance The obtained sweet cherry transcriptome and DGE profiling data provide comprehensive gene expression information that lends insights into the molecular mechanisms underlying anthocyanin biosynthesis. These results will provide a platform for further functional genomic research on this fruit crop. PMID:25799516

Low Temperature and Short-Term High-CO2 Treatment in Postharvest Storage of Table Grapes at Two Maturity Stages: Effects on Transcriptome Profiling

PubMed Central

Rosales, Raquel; Romero, Irene; Fernandez-Caballero, Carlos; Escribano, M. Isabel; Merodio, Carmen; Sanchez-Ballesta, M. Teresa

2016-01-01

Table grapes (Vitis vinifera cv. Cardinal) are highly perishable and their quality deteriorates during postharvest storage at low temperature mainly because of sensitivity to fungal decay and senescence of rachis. The application of a 3-day CO2 treatment (20 kPa CO2 + 20 kPa O2 + 60 kPa N2) at 0°C reduced total decay and retained fruit quality in early and late-harvested table grapes during postharvest storage. In order to study the transcriptional responsiveness of table grapes to low temperature and high CO2 levels in the first stage of storage and how the maturity stage affect these changes, we have performed a comparative large-scale transcriptional analysis using the custom-made GrapeGen GeneChip®. In the first stage of storage, low temperature led to a significantly intense change in grape skin transcriptome irrespective of fruit maturity, although there were different changes within each stage. In the case of CO2 treated samples, in comparison to fruit at time zero, only slight differences were observed. Functional enrichment analysis revealed that major modifications in the transcriptome profile of early- and late-harvested grapes stored at 0°C are linked to biotic and abiotic stress-responsive terms. However, in both cases there is a specific reprogramming of the transcriptome during the first stage of storage at 0°C in order to withstand the cold stress. Thus, genes involved in gluconeogenesis, photosynthesis, mRNA translation and lipid transport were up-regulated in the case of early-harvested grapes, and genes related to protein folding stability and intracellular membrane trafficking in late-harvested grapes. The beneficial effect of high CO2 treatment maintaining table grape quality seems to be an active process requiring the induction of several transcription factors and kinases in early-harvested grapes, and the activation of processes associated to the maintenance of energy in late-harvested grapes. PMID:27468290

Meta-Analyses of Dehalococcoides mccartyi Strain 195 Transcriptomic Profiles Identify a Respiration Rate-Related Gene Expression Transition Point and Interoperon Recruitment of a Key Oxidoreductase Subunit

PubMed Central

Mansfeldt, Cresten B.; Rowe, Annette R.; Heavner, Gretchen L. W.; Zinder, Stephen H.

2014-01-01

A cDNA-microarray was designed and used to monitor the transcriptomic profile of Dehalococcoides mccartyi strain 195 (in a mixed community) respiring various chlorinated organics, including chloroethenes and 2,3-dichlorophenol. The cultures were continuously fed in order to establish steady-state respiration rates and substrate levels. The organization of array data into a clustered heat map revealed two major experimental partitions. This partitioning in the data set was further explored through principal component analysis. The first two principal components separated the experiments into those with slow (1.6 ± 0.6 μM Cl−/h)- and fast (22.9 ± 9.6 μM Cl−/h)-respiring cultures. Additionally, the transcripts with the highest loadings in these principal components were identified, suggesting that those transcripts were responsible for the partitioning of the experiments. By analyzing the transcriptomes (n = 53) across experiments, relationships among transcripts were identified, and hypotheses about the relationships between electron transport chain members were proposed. One hypothesis, that the hydrogenases Hup and Hym and the formate dehydrogenase-like oxidoreductase (DET0186-DET0187) form a complex (as displayed by their tight clustering in the heat map analysis), was explored using a nondenaturing protein separation technique combined with proteomic sequencing. Although these proteins did not migrate as a single complex, DET0112 (an FdhB-like protein encoded in the Hup operon) was found to comigrate with DET0187 rather than with the catalytic Hup subunit DET0110. On closer inspection of the genome annotations of all Dehalococcoides strains, the DET0185-to-DET0187 operon was found to lack a key subunit, an FdhB-like protein. Therefore, on the basis of the transcriptomic, genomic, and proteomic evidence, the place of the missing subunit in the DET0185-to-DET0187 operon is likely filled by recruiting a subunit expressed from the Hup operon (DET0112). PMID:25063656

Transcriptomic analysis of the red seaweed Laurencia dendroidea (Florideophyceae, Rhodophyta) and its microbiome.

PubMed

de Oliveira, Louisi Souza; Gregoracci, Gustavo Bueno; Silva, Genivaldo Gueiros Zacarias; Salgado, Leonardo Tavares; Filho, Gilberto Amado; Alves-Ferreira, Marcio; Pereira, Renato Crespo; Thompson, Fabiano L

2012-09-17

Seaweeds of the Laurencia genus have a broad geographic distribution and are largely recognized as important sources of secondary metabolites, mainly halogenated compounds exhibiting diverse potential pharmacological activities and relevant ecological role as anti-epibiosis. Host-microbe interaction is a driving force for co-evolution in the marine environment, but molecular studies of seaweed-associated microbial communities are still rare. Despite the large amount of research describing the chemical compositions of Laurencia species, the genetic knowledge regarding this genus is currently restricted to taxonomic markers and general genome features. In this work we analyze the transcriptomic profile of L. dendroidea J. Agardh, unveil the genes involved on the biosynthesis of terpenoid compounds in this seaweed and explore the interactions between this host and its associated microbiome. A total of 6 transcriptomes were obtained from specimens of L. dendroidea sampled in three different coastal locations of the Rio de Janeiro state. Functional annotations revealed predominantly basic cellular metabolic pathways. Bacteria was the dominant active group in the microbiome of L. dendroidea, standing out nitrogen fixing Cyanobacteria and aerobic heterotrophic Proteobacteria. The analysis of the relative contribution of each domain highlighted bacterial features related to glycolysis, lipid and polysaccharide breakdown, and also recognition of seaweed surface and establishment of biofilm. Eukaryotic transcripts, on the other hand, were associated with photosynthesis, synthesis of carbohydrate reserves, and defense mechanisms, including the biosynthesis of terpenoids through the mevalonate-independent pathway. This work describes the first transcriptomic profile of the red seaweed L. dendroidea, increasing the knowledge about ESTs from the Florideophyceae algal class. Our data suggest an important role for L. dendroidea in the primary production of the holobiont and the role of Bacteria as consumers of organic matter and possibly also as nitrogen source. Furthermore, this seaweed expressed sequences related to terpene biosynthesis, including the complete mevalonate-independent pathway, which offers new possibilities for biotechnological applications using secondary metabolites from L. dendroidea.

De novo Transcriptome Analysis of Miscanthus lutarioriparius Identifies Candidate Genes in Rhizome Development

PubMed Central

Hu, Ruibo; Yu, Changjiang; Wang, Xiaoyu; Jia, Chunlin; Pei, Shengqiang; He, Kang; He, Guo; Kong, Yingzhen; Zhou, Gongke

2017-01-01

HIGHLIGHT De novo transcriptome profiling of five tissues reveals candidate genes putatively involved in rhizome development in M. lutarioriparius. Miscanthus lutarioriparius is a promising lignocellulosic feedstock for second-generation bioethanol production. However, the genomic resource for this species is relatively limited thus hampers our understanding of the molecular mechanisms underlying many important biological processes. In this study, we performed the first de novo transcriptome analysis of five tissues (leaf, stem, root, lateral bud and rhizome bud) of M. lutarioriparius with an emphasis to identify putative genes involved in rhizome development. Approximately 66 gigabase (GB) paired-end clean reads were obtained and assembled into 169,064 unigenes with an average length of 759 bp. Among these unigenes, 103,899 (61.5%) were annotated in seven public protein databases. Differential gene expression profiling analysis revealed that 4,609, 3,188, 1,679, 1,218, and 1,077 genes were predominantly expressed in root, leaf, stem, lateral bud, and rhizome bud, respectively. Their expression patterns were further classified into 12 distinct clusters. Pathway enrichment analysis revealed that genes predominantly expressed in rhizome bud were mainly involved in primary metabolism and hormone signaling and transduction pathways. Noteworthy, 19 transcription factors (TFs) and 16 hormone signaling pathway-related genes were identified to be predominantly expressed in rhizome bud compared with the other tissues, suggesting putative roles in rhizome formation and development. In addition, a predictive regulatory network was constructed between four TFs and six auxin and abscisic acid (ABA) -related genes. Furthermore, the expression of 24 rhizome-specific genes was further validated by quantitative real-time RT-PCR (qRT-PCR) analysis. Taken together, this study provide a global portrait of gene expression across five different tissues and reveal preliminary insights into rhizome growth and development. The data presented will contribute to our understanding of the molecular mechanisms underlying rhizome development in M. lutarioriparius and remarkably enrich the genomic resources of Miscanthus. PMID:28446913

Comparative transcriptome analysis of papilla and skin in the sea cucumber, Apostichopus japonicus.

PubMed

Zhou, Xiaoxu; Cui, Jun; Liu, Shikai; Kong, Derong; Sun, He; Gu, Chenlei; Wang, Hongdi; Qiu, Xuemei; Chang, Yaqing; Liu, Zhanjiang; Wang, Xiuli

2016-01-01

Papilla and skin are two important organs of the sea cucumber. Both tissues have ectodermic origin, but they are morphologically and functionally very different. In the present study, we performed comparative transcriptome analysis of the papilla and skin from the sea cucumber (Apostichopus japonicus) in order to identify and characterize gene expression profiles by using RNA-Seq technology. We generated 30.6 and 36.4 million clean reads from the papilla and skin and de novo assembled in 156,501 transcripts. The Gene Ontology (GO) analysis indicated that cell part, metabolic process and catalytic activity were the most abundant GO category in cell component, biological process and molecular funcation, respectively. Comparative transcriptome analysis between the papilla and skin allowed the identification of 1,059 differentially expressed genes, of which 739 genes were expressed at higher levels in papilla, while 320 were expressed at higher levels in skin. In addition, 236 differentially expressed unigenes were not annotated with any database, 160 of which were apparently expressed at higher levels in papilla, 76 were expressed at higher levels in skin. We identified a total of 288 papilla-specific genes, 171 skin-specific genes and 600 co-expressed genes. Also, 40 genes in papilla-specific were not annotated with any database, 2 in skin-specific. Development-related genes were also enriched, such as fibroblast growth factor, transforming growth factor-β, collagen-α2 and Integrin-α2, which may be related to the formation of the papilla and skin in sea cucumber. Further pathway analysis identified ten KEGG pathways that were differently enriched between the papilla and skin. The findings on expression profiles between two key organs of the sea cucumber should be valuable to reveal molecular mechanisms involved in the development of organs that are related but with morphological differences in the sea cucumber.

Pairwise comparisons of ten porcine tissues identify differential transcriptional regulation at the gene, isoform, promoter and transcription start site level

DOE Office of Scientific and Technical Information (OSTI.GOV)

Farajzadeh, Leila; Hornshøj, Henrik; Momeni, Jamal

Highlights: •Transcriptome sequencing yielded 223 mill porcine RNA-seq reads, and 59,000 transcribed locations. •Establishment of unique transcription profiles for ten porcine tissues including four brain tissues. •Comparison of transcription profiles at gene, isoform, promoter and transcription start site level. •Highlights a high level of regulation of neuro-related genes at both gene, isoform, and TSS level. •Our results emphasize the pig as a valuable animal model with respect to human biological issues. -- Abstract: The transcriptome is the absolute set of transcripts in a tissue or cell at the time of sampling. In this study RNA-Seq is employed to enable themore » differential analysis of the transcriptome profile for ten porcine tissues in order to evaluate differences between the tissues at the gene and isoform expression level, together with an analysis of variation in transcription start sites, promoter usage, and splicing. Totally, 223 million RNA fragments were sequenced leading to the identification of 59,930 transcribed gene locations and 290,936 transcript variants using Cufflinks with similarity to approximately 13,899 annotated human genes. Pairwise analysis of tissues for differential expression at the gene level showed that the smallest differences were between tissues originating from the porcine brain. Interestingly, the relative level of differential expression at the isoform level did generally not vary between tissue contrasts. Furthermore, analysis of differential promoter usage between tissues, revealed a proportionally higher variation between cerebellum (CBE) versus frontal cortex and cerebellum versus hypothalamus (HYP) than in the remaining comparisons. In addition, the comparison of differential transcription start sites showed that the number of these sites is generally increased in comparisons including hypothalamus in contrast to other pairwise assessments. A comprehensive analysis of one of the tissue contrasts, i.e. cerebellum versus heart for differential variation at the gene, isoform, and transcription start site (TSS), and promoter level showed that several of the genes differed at all four levels. Interestingly, these genes were mainly annotated to the “electron transport chain” and neuronal differentiation, emphasizing that “tissue important” genes are regulated at several levels. Furthermore, our analysis shows that the “across tissue approach” has a promising potential when screening for possible explanations for variations, such as those observed at the gene expression levels.« less

Transcriptomic meta-analysis identifies gene expression characteristics in various samples of HIV-infected patients with nonprogressive disease.

PubMed

Zhang, Le-Le; Zhang, Zi-Ning; Wu, Xian; Jiang, Yong-Jun; Fu, Ya-Jing; Shang, Hong

2017-09-12

A small proportion of HIV-infected patients remain clinically and/or immunologically stable for years, including elite controllers (ECs) who have undetectable viremia (<50 copies/ml) and long-term nonprogressors (LTNPs) who maintain normal CD4 + T cell counts for prolonged periods (>10 years). However, the mechanism of nonprogression needs to be further resolved. In this study, a transcriptome meta-analysis was performed on nonprogressor and progressor microarray data to identify differential transcriptome pathways and potential biomarkers. Using the INMEX (integrative meta-analysis of expression data) program, we performed the meta-analysis to identify consistently differentially expressed genes (DEGs) in nonprogressors and further performed functional interpretation (gene ontology analysis and pathway analysis) of the DEGs identified in the meta-analysis. Five microarray datasets (81 cases and 98 controls in total), including whole blood, CD4 + and CD8 + T cells, were collected for meta-analysis. We determined that nonprogressors have reduced expression of important interferon-stimulated genes (ISGs), CD38, lymphocyte activation gene 3 (LAG-3) in whole blood, CD4 + and CD8 + T cells. Gene ontology (GO) analysis showed a significant enrichment in DEGs that function in the type I interferon signaling pathway. Upregulated pathways, including the PI3K-Akt signaling pathway in whole blood, cytokine-cytokine receptor interaction in CD4 + T cells and the MAPK signaling pathway in CD8 + T cells, were identified in nonprogressors compared with progressors. In each metabolic functional category, the number of downregulated DEGs was more than the upregulated DEGs, and almost all genes were downregulated DEGs in the oxidative phosphorylation (OXPHOS) and tricarboxylic acid (TCA) cycle in the three types of samples. Our transcriptomic meta-analysis provides a comprehensive evaluation of the gene expression profiles in major blood types of nonprogressors, providing new insights in the understanding of HIV pathogenesis and developing strategies to delay HIV disease progression.

Molecular characteristics of the KCNJ5 mutated aldosterone-producing adenomas.

PubMed

Murakami, Masanori; Yoshimoto, Takanobu; Nakabayashi, Kazuhiko; Nakano, Yujiro; Fukaishi, Takahiro; Tsuchiya, Kyoichiro; Minami, Isao; Bouchi, Ryotaro; Okamura, Kohji; Fujii, Yasuhisa; Hashimoto, Koshi; Hata, Ken-Ichiro; Kihara, Kazunori; Ogawa, Yoshihiro

2017-10-01

The pathophysiology of aldosterone-producing adenomas (APAs) has been investigated via genetic approaches and the pathogenic significance of a series of somatic mutations, including KCNJ5 , has been uncovered. However, how the mutational status of an APA is associated with its molecular characteristics, including its transcriptome and methylome, has not been fully understood. This study was undertaken to explore the molecular characteristics of APAs, specifically focusing on APAs with KCNJ5 mutations as opposed to those without KCNJ5 mutations, by comparing their transcriptome and methylome status. Cortisol-producing adenomas (CPAs) were used as reference. We conducted transcriptome and methylome analyses of 29 APAs with KCNJ5 mutations, 8 APAs without KCNJ5 mutations and 5 CPAs. Genome-wide gene expression and CpG methylation profiles were obtained from RNA and DNA samples extracted from these 42 adrenal tumors. Cluster analysis of the transcriptome and methylome revealed molecular heterogeneity in APAs depending on their mutational status. DNA hypomethylation and gene expression changes in Wnt signaling and inflammatory response pathways were characteristic of APAs with KCNJ5 mutations. Comparisons between transcriptome data from our APAs and that from normal adrenal cortex obtained from the Gene Expression Omnibus suggested similarities between APAs with KCNJ5 mutations and zona glomerulosa. The present study, which is based on transcriptome and methylome analyses, indicates the molecular heterogeneity of APAs depends on their mutational status. Here, we report the unique characteristics of APAs with KCNJ5 mutations. © 2017 Society for Endocrinology.

Lathyrus sativus transcriptome resistance response to Ascochyta lathyri investigated by deepSuperSAGE analysis

PubMed Central

Almeida, Nuno F.; Krezdorn, Nicolas; Rotter, Björn; Winter, Peter; Rubiales, Diego; Vaz Patto, Maria C.

2015-01-01

Lathyrus sativus (grass pea) is a temperate grain legume crop with a great potential for expansion in dry areas or zones that are becoming more drought-prone. It is also recognized as a potential source of resistance to several important diseases in legumes, such as ascochyta blight. Nevertheless, the lack of detailed genomic and/or transcriptomic information hampers further exploitation of grass pea resistance-related genes in precision breeding. To elucidate the pathways differentially regulated during ascochyta-grass pea interaction and to identify resistance candidate genes, we compared the early response of the leaf gene expression profile of a resistant L. sativus genotype to Ascochyta lathyri infection with a non-inoculated control sample from the same genotype employing deepSuperSAGE. This analysis generated 14.387 UniTags of which 95.7% mapped to a reference grass pea/rust interaction transcriptome. From the total mapped UniTags, 738 were significantly differentially expressed between control and inoculated leaves. The results indicate that several gene classes acting in different phases of the plant/pathogen interaction are involved in the L. sativus response to A. lathyri infection. Most notably a clear up-regulation of defense-related genes involved in and/or regulated by the ethylene pathway was observed. There was also evidence of alterations in cell wall metabolism indicated by overexpression of cellulose synthase and lignin biosynthesis genes. This first genome-wide overview of the gene expression profile of the L. sativus response to ascochyta infection delivered a valuable set of candidate resistance genes for future use in precision breeding. PMID:25852725

Gene Expression Profiling of Development and Anthocyanin Accumulation in Kiwifruit (Actinidia chinensis) Based on Transcriptome Sequencing

PubMed Central

Zeng, Shaohua; Xiao, Gong; Wang, Gan; Wang, Ying; Peng, Ming; Huang, Hongwen

2015-01-01

Red-fleshed kiwifruit (Actinidia chinensis Planch. ‘Hongyang’) is a promising commercial cultivar due to its nutritious value and unique flesh color, derived from vitamin C and anthocyanins. In this study, we obtained transcriptome data of ‘Hongyang’ from seven developmental stages using Illumina sequencing. We mapped 39–54 million reads to the recently sequenced kiwifruit genome and other databases to define gene structure, to analyze alternative splicing, and to quantify gene transcript abundance at different developmental stages. The transcript profiles throughout red kiwifruit development were constructed and analyzed, with a focus on the biosynthesis and metabolism of compounds such as phytohormones, sugars, starch and L-ascorbic acid, which are indispensable for the development and formation of quality fruit. Candidate genes for these pathways were identified through MapMan and phylogenetic analysis. The transcript levels of genes involved in sucrose and starch metabolism were consistent with the change in soluble sugar and starch content throughout kiwifruit development. The metabolism of L-ascorbic acid was very active, primarily through the L-galactose pathway. The genes responsible for the accumulation of anthocyanin in red kiwifruit were identified, and their expression levels were investigated during kiwifruit development. This survey of gene expression during kiwifruit development paves the way for further investigation of the development of this uniquely colored and nutritious fruit and reveals which factors are needed for high quality fruit formation. This transcriptome data and its analysis will be useful for improving kiwifruit genome annotation, for basic fruit molecular biology research, and for kiwifruit breeding and improvement. PMID:26301713

RNA-Seq Analysis of the Expression of Genes Encoding Cell Wall Degrading Enzymes during Infection of Lupin (Lupinus angustifolius) by Phytophthora parasitica

PubMed Central

Blackman, Leila M.; Cullerne, Darren P.; Torreña, Pernelyn; Taylor, Jen; Hardham, Adrienne R.

2015-01-01

RNA-Seq analysis has shown that over 60% (12,962) of the predicted transcripts in the Phytophthora parasitica genome are expressed during the first 60 h of lupin root infection. The infection transcriptomes included 278 of the 431 genes encoding P. parasitica cell wall degrading enzymes. The transcriptome data provide strong evidence of global transcriptional cascades of genes whose encoded proteins target the main categories of plant cell wall components. A major cohort of pectinases is predominantly expressed early but as infection progresses, the transcriptome becomes increasingly dominated by transcripts encoding cellulases, hemicellulases, β-1,3-glucanases and glycoproteins. The most highly expressed P. parasitica carbohydrate active enzyme gene contains two CBM1 cellulose binding modules and no catalytic domains. The top 200 differentially expressed genes include β-1,4-glucosidases, β-1,4-glucanases, β-1,4-galactanases, a β-1,3-glucanase, an α-1,4-polygalacturonase, a pectin deacetylase and a pectin methylesterase. Detailed analysis of gene expression profiles provides clues as to the order in which linkages within the complex carbohydrates may come under attack. The gene expression profiles suggest that (i) demethylation of pectic homogalacturonan occurs before its deacetylation; (ii) cleavage of the backbone of pectic rhamnogalacturonan I precedes digestion of its side chains; (iii) early attack on cellulose microfibrils by non-catalytic cellulose-binding proteins and enzymes with auxiliary activities may facilitate subsequent attack by glycosyl hydrolases and enzymes containing CBM1 cellulose-binding modules; (iv) terminal hemicellulose backbone residues are targeted after extensive internal backbone cleavage has occurred; and (v) the carbohydrate chains on glycoproteins are degraded late in infection. A notable feature of the P. parasitica infection transcriptome is the high level of transcription of genes encoding enzymes that degrade β-1,3-glucanases during middle and late stages of infection. The results suggest that high levels of β-1,3-glucanases may effectively degrade callose as it is produced by the plant during the defence response. PMID:26332397

RNA-Seq Analysis of the Expression of Genes Encoding Cell Wall Degrading Enzymes during Infection of Lupin (Lupinus angustifolius) by Phytophthora parasitica.

PubMed

Blackman, Leila M; Cullerne, Darren P; Torreña, Pernelyn; Taylor, Jen; Hardham, Adrienne R

2015-01-01

RNA-Seq analysis has shown that over 60% (12,962) of the predicted transcripts in the Phytophthora parasitica genome are expressed during the first 60 h of lupin root infection. The infection transcriptomes included 278 of the 431 genes encoding P. parasitica cell wall degrading enzymes. The transcriptome data provide strong evidence of global transcriptional cascades of genes whose encoded proteins target the main categories of plant cell wall components. A major cohort of pectinases is predominantly expressed early but as infection progresses, the transcriptome becomes increasingly dominated by transcripts encoding cellulases, hemicellulases, β-1,3-glucanases and glycoproteins. The most highly expressed P. parasitica carbohydrate active enzyme gene contains two CBM1 cellulose binding modules and no catalytic domains. The top 200 differentially expressed genes include β-1,4-glucosidases, β-1,4-glucanases, β-1,4-galactanases, a β-1,3-glucanase, an α-1,4-polygalacturonase, a pectin deacetylase and a pectin methylesterase. Detailed analysis of gene expression profiles provides clues as to the order in which linkages within the complex carbohydrates may come under attack. The gene expression profiles suggest that (i) demethylation of pectic homogalacturonan occurs before its deacetylation; (ii) cleavage of the backbone of pectic rhamnogalacturonan I precedes digestion of its side chains; (iii) early attack on cellulose microfibrils by non-catalytic cellulose-binding proteins and enzymes with auxiliary activities may facilitate subsequent attack by glycosyl hydrolases and enzymes containing CBM1 cellulose-binding modules; (iv) terminal hemicellulose backbone residues are targeted after extensive internal backbone cleavage has occurred; and (v) the carbohydrate chains on glycoproteins are degraded late in infection. A notable feature of the P. parasitica infection transcriptome is the high level of transcription of genes encoding enzymes that degrade β-1,3-glucanases during middle and late stages of infection. The results suggest that high levels of β-1,3-glucanases may effectively degrade callose as it is produced by the plant during the defence response.

Revealing the beneficial effect of protease supplementation to high gravity beer fermentations using "-omics" techniques

PubMed Central

2011-01-01

Background Addition of sugar syrups to the basic wort is a popular technique to achieve higher gravity in beer fermentations, but it results in dilution of the free amino nitrogen (FAN) content in the medium. The multicomponent protease enzyme Flavourzyme has beneficial effect on the brewer's yeast fermentation performance during high gravity fermentations as it increases the initial FAN value and results in higher FAN uptake, higher specific growth rate, higher ethanol yield and improved flavour profile. Results In the present study, transcriptome and metabolome analysis were used to elucidate the effect on the addition of the multicomponent protease enzyme Flavourzyme and its influence on the metabolism of the brewer's yeast strain Weihenstephan 34/70. The study underlines the importance of sufficient nitrogen availability during the course of beer fermentation. The applied metabolome and transcriptome analysis allowed mapping the effect of the wort sugar composition on the nitrogen uptake. Conclusion Both the transcriptome and the metabolome analysis revealed that there is a significantly higher impact of protease addition for maltose syrup supplemented fermentations, while addition of glucose syrup to increase the gravity in the wort resulted in increased glucose repression that lead to inhibition of amino acid uptake and hereby inhibited the effect of the protease addition. PMID:21513553

Transcriptome analysis of thermogenic Arum concinnatum reveals the molecular components of floral scent production

PubMed Central

Onda, Yoshihiko; Mochida, Keiichi; Yoshida, Takuhiro; Sakurai, Tetsuya; Seymour, Roger S.; Umekawa, Yui; Pirintsos, Stergios Arg; Shinozaki, Kazuo; Ito, Kikukatsu

2015-01-01

Several plant species can generate enough heat to increase their internal floral temperature above ambient temperature. Among thermogenic plants, Arum concinnatum shows the highest respiration activity during thermogenesis. However, an overall understanding of the genes related to plant thermogenesis has not yet been achieved. In this study, we performed de novo transcriptome analysis of flower organs in A. concinnatum. The de novo transcriptome assembly represented, in total, 158,490 non-redundant transcripts, and 53,315 of those showed significant homology with known genes. To explore genes associated with thermogenesis, we filtered 1266 transcripts that showed a significant correlation between expression pattern and the temperature trend of each sample. We confirmed five putative alternative oxidase transcripts were included in filtered transcripts as expected. An enrichment analysis of the Gene Ontology terms for the filtered transcripts suggested over-representation of genes involved in 1-deoxy-d-xylulose-5-phosphate synthase (DXS) activity. The expression profiles of DXS transcripts in the methyl-d-erythritol 4-phosphate (MEP) pathway were significantly correlated with thermogenic levels. Our results suggest that the MEP pathway is the main biosynthesis route for producing scent monoterpenes. To our knowledge, this is the first report describing the candidate pathway and the key enzyme for floral scent production in thermogenic plants. PMID:25736477

Morphological Characters and Transcriptome Profiles Associated with Black Skin and Red Skin in Crimson Snapper (Lutjanus erythropterus)

PubMed Central

Zhang, Yan-Ping; Wang, Zhong-Duo; Guo, Yu-Song; Liu, Li; Yu, Juan; Zhang, Shun; Liu, Shao-Jun; Liu, Chu-Wu

2015-01-01

In this study, morphology observation and illumina sequencing were performed on two different coloration skins of crimson snapper (Lutjanus erythropterus), the black zone and the red zone. Three types of chromatophores, melanophores, iridophores and xanthophores, were organized in the skins. The main differences between the two colorations were in the amount and distribution of the three chromatophores. After comparing the two transcriptomes, 9200 unigenes with significantly different expressions (ratio change ≥ 2 and q-value ≤ 0.05) were found, of which 5972 were up-regulated in black skin and 3228 were up-regulated in red skin. Through the function annotation, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the differentially transcribed genes, we excavated a number of uncharacterized candidate pigment genes as well as found the conserved genes affecting pigmentation in crimson snapper. The patterns of expression of 14 pigment genes were confirmed by the Quantitative real-time PCR analysis between the two color skins. Overall, this study shows a global survey of the morphological characters and transcriptome analysis of the different coloration skins in crimson snapper, and provides valuable cellular and genetic information to uncover the mechanism of the formation of pigment patterns in snappers. PMID:26569232

Morphological Characters and Transcriptome Profiles Associated with Black Skin and Red Skin in Crimson Snapper (Lutjanus erythropterus).

PubMed

Zhang, Yan-Ping; Wang, Zhong-Duo; Guo, Yu-Song; Liu, Li; Yu, Juan; Zhang, Shun; Liu, Shao-Jun; Liu, Chu-Wu

2015-11-12

In this study, morphology observation and illumina sequencing were performed on two different coloration skins of crimson snapper (Lutjanus erythropterus), the black zone and the red zone. Three types of chromatophores, melanophores, iridophores and xanthophores, were organized in the skins. The main differences between the two colorations were in the amount and distribution of the three chromatophores. After comparing the two transcriptomes, 9200 unigenes with significantly different expressions (ratio change ≥ 2 and q-value ≤ 0.05) were found, of which 5972 were up-regulated in black skin and 3228 were up-regulated in red skin. Through the function annotation, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the differentially transcribed genes, we excavated a number of uncharacterized candidate pigment genes as well as found the conserved genes affecting pigmentation in crimson snapper. The patterns of expression of 14 pigment genes were confirmed by the Quantitative real-time PCR analysis between the two color skins. Overall, this study shows a global survey of the morphological characters and transcriptome analysis of the different coloration skins in crimson snapper, and provides valuable cellular and genetic information to uncover the mechanism of the formation of pigment patterns in snappers.

De Novo Transcriptome Sequencing of Olea europaea L. to Identify Genes Involved in the Development of the Pollen Tube.

PubMed

Iaria, Domenico; Chiappetta, Adriana; Muzzalupo, Innocenzo

2016-01-01

In olive (Olea europaea L.), the processes controlling self-incompatibility are still unclear and the molecular basis underlying this process are still not fully characterized. In order to determine compatibility relationships, using next-generation sequencing techniques and a de novo transcriptome assembly strategy, we show that pollen tubes from different olive plants, grown in vitro in a medium containing its own pistil and in combination pollen/pistil from self-sterile and self-fertile cultivars, have a distinct gene expression profile and many of the differentially expressed sequences between the samples fall within gene families involved in the development of the pollen tube, such as lipase, carboxylesterase, pectinesterase, pectin methylesterase, and callose synthase. Moreover, different genes involved in signal transduction, transcription, and growth are overrepresented. The analysis also allowed us to identify members in actin and actin depolymerization factor and fibrin gene family and member of the Ca(2+) binding gene family related to the development and polarization of pollen apical tip. The whole transcriptomic analysis, through the identification of the differentially expressed transcripts set and an extended functional annotation analysis, will lead to a better understanding of the mechanisms of pollen germination and pollen tube growth in the olive.

Transcriptome-wide N 6 -methyladenosine methylome profiling of porcine muscle and adipose tissues reveals a potential mechanism for transcriptional regulation and differential methylation pattern.

PubMed

Tao, Xuelian; Chen, Jianning; Jiang, Yanzhi; Wei, Yingying; Chen, Yan; Xu, Huaming; Zhu, Li; Tang, Guoqing; Li, Mingzhou; Jiang, Anan; Shuai, Surong; Bai, Lin; Liu, Haifeng; Ma, Jideng; Jin, Long; Wen, Anxiang; Wang, Qin; Zhu, Guangxiang; Xie, Meng; Wu, Jiayun; He, Tao; Huang, Chunyu; Gao, Xiang; Li, Xuewei

2017-04-28

N 6 -methyladenosine (m 6 A) is the most prevalent internal form of modification in messenger RNA in higher eukaryotes and potential regulatory functions of reversible m 6 A methylation on mRNA have been revealed by mapping of m 6 A methylomes in several species. m 6 A modification in active gene regulation manifests itself as altered methylation profiles in a tissue-specific manner or in response to changing cellular or species living environment. However, up to date, there has no data on m 6 A porcine transcriptome-wide map and its potential biological roles in adipose deposition and muscle growth. In this work, we used methylated RNA immunoprecipitation with next-generation sequencing (MeRIP-Seq) technique to acquire the first ever m 6 A porcine transcriptome-wide map. Transcriptomes of muscle and adipose tissues from three different pig breeds, the wild boar, Landrace, and Rongchang pig, were used to generate these maps. Our findings show that there were 5,872 and 2,826 m 6 A peaks respectively, in the porcine muscle and adipose tissue transcriptomes. Stop codons, 3'-untranslated regions, and coding regions were found to be mainly enriched for m 6 A peaks. Gene ontology analysis revealed that common m 6 A peaks in nuclear genes are associated with transcriptional factors, suggestive of a relationship between m 6 A mRNA methylation and nuclear genome transcription. Some genes showed tissue- and breed-differential methylation, and have novel biological functions. We also found a relationship between the m 6 A methylation extent and the transcript level, suggesting a regulatory role for m 6 A in gene expression. This comprehensive map provides a solid basis for the determination of potential functional roles for RNA m 6 A modification in adipose deposition and muscle growth.

Probing the Reproducibility of Leaf Growth and Molecular Phenotypes: A Comparison of Three Arabidopsis Accessions Cultivated in Ten Laboratories1[W

PubMed Central

Massonnet, Catherine; Vile, Denis; Fabre, Juliette; Hannah, Matthew A.; Caldana, Camila; Lisec, Jan; Beemster, Gerrit T.S.; Meyer, Rhonda C.; Messerli, Gaëlle; Gronlund, Jesper T.; Perkovic, Josip; Wigmore, Emma; May, Sean; Bevan, Michael W.; Meyer, Christian; Rubio-Díaz, Silvia; Weigel, Detlef; Micol, José Luis; Buchanan-Wollaston, Vicky; Fiorani, Fabio; Walsh, Sean; Rinn, Bernd; Gruissem, Wilhelm; Hilson, Pierre; Hennig, Lars; Willmitzer, Lothar; Granier, Christine

2010-01-01

A major goal of the life sciences is to understand how molecular processes control phenotypes. Because understanding biological systems relies on the work of multiple laboratories, biologists implicitly assume that organisms with the same genotype will display similar phenotypes when grown in comparable conditions. We investigated to what extent this holds true for leaf growth variables and metabolite and transcriptome profiles of three Arabidopsis (Arabidopsis thaliana) genotypes grown in 10 laboratories using a standardized and detailed protocol. A core group of four laboratories generated similar leaf growth phenotypes, demonstrating that standardization is possible. But some laboratories presented significant differences in some leaf growth variables, sometimes changing the genotype ranking. Metabolite profiles derived from the same leaf displayed a strong genotype × environment (laboratory) component. Genotypes could be separated on the basis of their metabolic signature, but only when the analysis was limited to samples derived from one laboratory. Transcriptome data revealed considerable plant-to-plant variation, but the standardization ensured that interlaboratory variation was not considerably larger than intralaboratory variation. The different impacts of the standardization on phenotypes and molecular profiles could result from differences of temporal scale between processes involved at these organizational levels. Our findings underscore the challenge of describing, monitoring, and precisely controlling environmental conditions but also demonstrate that dedicated efforts can result in reproducible data across multiple laboratories. Finally, our comparative analysis revealed that small variations in growing conditions (light quality principally) and handling of plants can account for significant differences in phenotypes and molecular profiles obtained in independent laboratories. PMID:20200072

Digital transcriptome profiling of normal and glioblastoma-derived neural stem cells identifies genes associated with patient survival

PubMed Central

2012-01-01

Background Glioblastoma multiforme, the most common type of primary brain tumor in adults, is driven by cells with neural stem (NS) cell characteristics. Using derivation methods developed for NS cells, it is possible to expand tumorigenic stem cells continuously in vitro. Although these glioblastoma-derived neural stem (GNS) cells are highly similar to normal NS cells, they harbor mutations typical of gliomas and initiate authentic tumors following orthotopic xenotransplantation. Here, we analyzed GNS and NS cell transcriptomes to identify gene expression alterations underlying the disease phenotype. Methods Sensitive measurements of gene expression were obtained by high-throughput sequencing of transcript tags (Tag-seq) on adherent GNS cell lines from three glioblastoma cases and two normal NS cell lines. Validation by quantitative real-time PCR was performed on 82 differentially expressed genes across a panel of 16 GNS and 6 NS cell lines. The molecular basis and prognostic relevance of expression differences were investigated by genetic characterization of GNS cells and comparison with public data for 867 glioma biopsies. Results Transcriptome analysis revealed major differences correlated with glioma histological grade, and identified misregulated genes of known significance in glioblastoma as well as novel candidates, including genes associated with other malignancies or glioma-related pathways. This analysis further detected several long non-coding RNAs with expression profiles similar to neighboring genes implicated in cancer. Quantitative PCR validation showed excellent agreement with Tag-seq data (median Pearson r = 0.91) and discerned a gene set robustly distinguishing GNS from NS cells across the 22 lines. These expression alterations include oncogene and tumor suppressor changes not detected by microarray profiling of tumor tissue samples, and facilitated the identification of a GNS expression signature strongly associated with patient survival (P = 1e-6, Cox model). Conclusions These results support the utility of GNS cell cultures as a model system for studying the molecular processes driving glioblastoma and the use of NS cells as reference controls. The association between a GNS expression signature and survival is consistent with the hypothesis that a cancer stem cell component drives tumor growth. We anticipate that analysis of normal and malignant stem cells will be an important complement to large-scale profiling of primary tumors. PMID:23046790

Transcriptomic analysis illuminates genes involved in chlorophyll synthesis after nitrogen starvation in Acaryochloris sp. CCMEE 5410.

PubMed

Yoneda, Aki; Wittmann, Bruce J; King, Jeremy D; Blankenship, Robert E; Dantas, Gautam

2016-08-01

Acaryochloris species are a genus of cyanobacteria that utilize chlorophyll (chl) d as their primary chlorophyll molecule during oxygenic photosynthesis. Chl d allows Acaryochloris to harvest red-shifted light, which gives them the ability to live in filtered light environments that are depleted in visible light. Although genomes of multiple Acaryochloris species have been sequenced, their analysis has not revealed how chl d is synthesized. Here, we demonstrate that Acaryochloris sp. CCMEE 5410 cells undergo chlorosis by nitrogen depletion and exhibit robust regeneration of chl d by nitrogen repletion. We performed a time course RNA-Seq experiment to quantify global transcriptomic changes during chlorophyll recovery. We observed upregulation of numerous known chl biosynthesis genes and also identified an oxygenase gene with a similar transcriptional profile as these chl biosynthesis genes, suggesting its possible involvement in chl d biosynthesis. Moreover, our data suggest that multiple prochlorophyte chlorophyll-binding homologs are important during chlorophyll recovery, and light-independent chl synthesis genes are more dominant than the light-dependent gene at the transcription level. Transcriptomic characterization of this organism provides crucial clues toward mechanistic elucidation of chl d biosynthesis.

Unravelling molecular mechanisms from floral initiation to lipid biosynthesis in a promising biofuel tree species, Pongamia pinnata using transcriptome analysis

PubMed Central

Sreeharsha, Rachapudi V.; Mudalkar, Shalini; Singha, Kambam T.; Reddy, Attipalli R.

2016-01-01

Pongamia pinnata (L.) (Fabaceae) is a promising biofuel tree species which is underexploited in the areas of both fundamental and applied research, due to the lack of information either on transcriptome or genomic data. To investigate the possible metabolic pathways, we performed whole transcriptome analysis of Pongamia through Illumina NextSeq platform and generated 2.8 GB of paired end sequence reads. The de novo assembly of raw reads generated 40,000 contigs and 35,000 transcripts, representing leaf, flower and seed unigenes. Spatial and temporal expression profiles of photoperiod and floral homeotic genes in Pongamia, identified GIGANTEA (GI) - CONSTANS (CO) - FLOWERING LOCUS T (FT) as active signal cascade for floral initiation. Four prominent stages of seed development were selected in a high yielding Pongamia accession (TOIL 1) to follow the temporal expression patterns of key fatty acid biosynthetic genes involved in lipid biosynthesis and accumulation. Our results provide insights into an array of molecular events from flowering to seed maturity in Pongamia which will provide substantial basis for modulation of fatty acid composition and enhancing oil yields which should serve as a potential feedstock for biofuel production. PMID:27677333

Spatial transcriptomic analysis of cryosectioned tissue samples with Geo-seq.

PubMed

Chen, Jun; Suo, Shengbao; Tam, Patrick Pl; Han, Jing-Dong J; Peng, Guangdun; Jing, Naihe

2017-03-01

Conventional gene expression studies analyze multiple cells simultaneously or single cells, for which the exact in vivo or in situ position is unknown. Although cellular heterogeneity can be discerned when analyzing single cells, any spatially defined attributes that underpin the heterogeneous nature of the cells cannot be identified. Here, we describe how to use Geo-seq, a method that combines laser capture microdissection (LCM) and single-cell RNA-seq technology. The combination of these two methods enables the elucidation of cellular heterogeneity and spatial variance simultaneously. The Geo-seq protocol allows the profiling of transcriptome information from only a small number cells and retains their native spatial information. This protocol has wide potential applications to address biological and pathological questions of cellular properties such as prospective cell fates, biological function and the gene regulatory network. Geo-seq has been applied to investigate the spatial transcriptome of mouse early embryo, mouse brain, and pathological liver and sperm tissues. The entire protocol from tissue collection and microdissection to sequencing requires ∼5 d, Data analysis takes another 1 or 2 weeks, depending on the amount of data and the speed of the processor.

Transcriptional landscapes of Axolotl (Ambystoma mexicanum).

PubMed

Caballero-Pérez, Juan; Espinal-Centeno, Annie; Falcon, Francisco; García-Ortega, Luis F; Curiel-Quesada, Everardo; Cruz-Hernández, Andrés; Bako, Laszlo; Chen, Xuemei; Martínez, Octavio; Alberto Arteaga-Vázquez, Mario; Herrera-Estrella, Luis; Cruz-Ramírez, Alfredo

2018-01-15

The axolotl (Ambystoma mexicanum) is the vertebrate model system with the highest regeneration capacity. Experimental tools established over the past 100 years have been fundamental to start unraveling the cellular and molecular basis of tissue and limb regeneration. In the absence of a reference genome for the Axolotl, transcriptomic analysis become fundamental to understand the genetic basis of regeneration. Here we present one of the most diverse transcriptomic data sets for Axolotl by profiling coding and non-coding RNAs from diverse tissues. We reconstructed a population of 115,906 putative protein coding mRNAs as full ORFs (including isoforms). We also identified 352 conserved miRNAs and 297 novel putative mature miRNAs. Systematic enrichment analysis of gene expression allowed us to identify tissue-specific protein-coding transcripts. We also found putative novel and conserved microRNAs which potentially target mRNAs which are reported as important disease candidates in heart and liver. Copyright © 2017 Elsevier Inc. All rights reserved.

Convergence in probiotic Lactobacillus gut-adaptive responses in humans and mice.

PubMed

Marco, Maria L; de Vries, Maaike C; Wels, Michiel; Molenaar, Douwe; Mangell, Peter; Ahrne, Siv; de Vos, Willem M; Vaughan, Elaine E; Kleerebezem, Michiel

2010-11-01

Probiotic bacteria provide unique opportunities to study the global responses and molecular mechanisms underlying the effects of gut-associated microorganisms in the human digestive tract. In this study, we show by comparative transcriptome analysis using DNA microarrays that the established probiotic Lactobacillus plantarum 299v specifically adapts its metabolic capacity in the human intestine for carbohydrate acquisition and expression of exopolysaccharide and proteinaceous cell surface compounds. This report constitutes the first application of global gene expression profiling of a commensal microorganism in the human gut. A core L. plantarum transcriptome expressed in the mammalian intestine was also determined through comparisons of L. plantarum 299v activities in humans to those found for L. plantarum WCFS1 in germ-free mice. These results identify the niche-specific adaptations of a dietary microorganism to the intestinal ecosystem and provide novel targets for molecular analysis of microbial-host interactions which affect human health.

Integrative Analysis of Longitudinal Metabolomics Data from a Personal Multi-Omics Profile

PubMed Central

Stanberry, Larissa; Mias, George I.; Haynes, Winston; Higdon, Roger; Snyder, Michael; Kolker, Eugene

2013-01-01

The integrative personal omics profile (iPOP) is a pioneering study that combines genomics, transcriptomics, proteomics, metabolomics and autoantibody profiles from a single individual over a 14-month period. The observation period includes two episodes of viral infection: a human rhinovirus and a respiratory syncytial virus. The profile studies give an informative snapshot into the biological functioning of an organism. We hypothesize that pathway expression levels are associated with disease status. To test this hypothesis, we use biological pathways to integrate metabolomics and proteomics iPOP data. The approach computes the pathways’ differential expression levels at each time point, while taking into account the pathway structure and the longitudinal design. The resulting pathway levels show strong association with the disease status. Further, we identify temporal patterns in metabolite expression levels. The changes in metabolite expression levels also appear to be consistent with the disease status. The results of the integrative analysis suggest that changes in biological pathways may be used to predict and monitor the disease. The iPOP experimental design, data acquisition and analysis issues are discussed within the broader context of personal profiling. PMID:24958148

Transcriptome assembly and expression profiling of molecular responses to cadmium toxicity in hepatopancreas of the freshwater crab Sinopotamon henanense

NASA Astrophysics Data System (ADS)

Sun, Min; Ting Li, Yi; Liu, Yang; Chin Lee, Shao; Wang, Lan

2016-01-01

Cadmium (Cd) pollution is a serious global problem, which causes irreversible toxic effects on animals. Freshwater crab, Sinopotamon henanense, is a useful environmental indicator since it is widely distributed in benthic habitats whereby it tends to accumulate Cd and other toxicants. However, its molecular responses to Cd toxicity remain unclear. In this study, we performed transcriptome sequencing and gene expression analyses of its hepatopancreas with and without Cd treatments. A total of 7.78 G clean reads were obtained from the pooled samples, and 68,648 unigenes with an average size of 622 bp were assembled, in which 5,436 were metabolism-associated and 2,728 were stimulus response-associated that include 380 immunity-related unigenes. Expression profile analysis demonstrated that most genes involved in macromolecular metabolism, oxidative phosphorylation, detoxification and anti-oxidant defense were up-regulated by Cd exposure, whereas immunity-related genes were down-regulated, except the genes involved in phagocytosis were up-regulated. The current data indicate that Cd exposure alters gene expressions in a concentration-dependent manner. Therefore, our results provide the first comprehensive S.henanense transcriptome dataset, which is useful for biological and ecotoxicological studies on this crab and its related species at molecular level, and some key Cd-responsive genes may provide candidate biomarkers for monitoring aquatic pollution by heavy metals.

Dynamics in the tomato root transcriptome on infection with the potato cyst nematode Globodera rostochiensis.

PubMed

Swiecicka, Magdalena; Filipecki, Marcin; Lont, Dieuwertje; Van Vliet, Joke; Qin, Ling; Goverse, Aska; Bakker, Jaap; Helder, Johannes

2009-07-01

Plant parasitic nematodes infect roots and trigger the formation of specialized feeding sites by substantial reprogramming of the developmental process of root cells. In this article, we describe the dynamic changes in the tomato root transcriptome during early interactions with the potato cyst nematode Globodera rostochiensis. Using amplified fragment length polymorphism-based mRNA fingerprinting (cDNA-AFLP), we monitored 17 600 transcript-derived fragments (TDFs) in infected and uninfected tomato roots, 1-14 days after inoculation with nematode larvae. Six hundred and twenty-four TDFs (3.5%) showed significant differential expression on nematode infection. We employed GenEST, a computer program which links gene expression profiles generated by cDNA-AFLP and databases of cDNA sequences, to identify 135 tomato sequences. These sequences were grouped into eight functional categories based on the presence of genes involved in hormone regulation, plant pathogen defence response, cell cycle and cytoskeleton regulation, cell wall modification, cellular signalling, transcriptional regulation, primary metabolism and allocation. The presence of unclassified genes was also taken into consideration. This article describes the responsiveness of numerous tomato genes hitherto uncharacterized during infection with endoparasitic cyst nematodes. The analysis of transcriptome profiles allowed the sequential order of expression to be dissected for many groups of genes and the genes to be connected with the biological processes involved in compatible interactions between the plant and nematode.

Single cell transcriptomic analysis of prostate cancer cells.

PubMed

Welty, Christopher J; Coleman, Ilsa; Coleman, Roger; Lakely, Bryce; Xia, Jing; Chen, Shu; Gulati, Roman; Larson, Sandy R; Lange, Paul H; Montgomery, Bruce; Nelson, Peter S; Vessella, Robert L; Morrissey, Colm

2013-02-16

The ability to interrogate circulating tumor cells (CTC) and disseminated tumor cells (DTC) is restricted by the small number detected and isolated (typically <10). To determine if a commercially available technology could provide a transcriptomic profile of a single prostate cancer (PCa) cell, we clonally selected and cultured a single passage of cell cycle synchronized C4-2B PCa cells. Ten sets of single, 5-, or 10-cells were isolated using a micromanipulator under direct visualization with an inverted microscope. Additionally, two groups of 10 individual DTC, each isolated from bone marrow of 2 patients with metastatic PCa were obtained. RNA was amplified using the WT-Ovation™ One-Direct Amplification System. The amplified material was hybridized on a 44K Whole Human Gene Expression Microarray. A high stringency threshold, a mean Alexa Fluor® 3 signal intensity above 300, was used for gene detection. Relative expression levels were validated for select genes using real-time PCR (RT-qPCR). Using this approach, 22,410, 20,423, and 17,009 probes were positive on the arrays from 10-cell pools, 5-cell pools, and single-cells, respectively. The sensitivity and specificity of gene detection on the single-cell analyses were 0.739 and 0.972 respectively when compared to 10-cell pools, and 0.814 and 0.979 respectively when compared to 5-cell pools, demonstrating a low false positive rate. Among 10,000 randomly selected pairs of genes, the Pearson correlation coefficient was 0.875 between the single-cell and 5-cell pools and 0.783 between the single-cell and 10-cell pools. As expected, abundant transcripts in the 5- and 10-cell samples were detected by RT-qPCR in the single-cell isolates, while lower abundance messages were not. Using the same stringency, 16,039 probes were positive on the patient single-cell arrays. Cluster analysis showed that all 10 DTC grouped together within each patient. A transcriptomic profile can be reliably obtained from a single cell using commercially available technology. As expected, fewer amplified genes are detected from a single-cell sample than from pooled-cell samples, however this method can be used to reliably obtain a transcriptomic profile from DTC isolated from the bone marrow of patients with PCa.

Genomic, transcriptomic, and proteomic approaches towards understanding the molecular mechanisms of salt tolerance in Frankia strains isolated from Casuarina trees.

PubMed

Oshone, Rediet; Ngom, Mariama; Chu, Feixia; Mansour, Samira; Sy, Mame Ourèye; Champion, Antony; Tisa, Louis S

2017-08-18

Soil salinization is a worldwide problem that is intensifying because of the effects of climate change. An effective method for the reclamation of salt-affected soils involves initiating plant succession using fast growing, nitrogen fixing actinorhizal trees such as the Casuarina. The salt tolerance of Casuarina is enhanced by the nitrogen-fixing symbiosis that they form with the actinobacterium Frankia. Identification and molecular characterization of salt-tolerant Casuarina species and associated Frankia is imperative for the successful utilization of Casuarina trees in saline soil reclamation efforts. In this study, salt-tolerant and salt-sensitive Casuarina associated Frankia strains were identified and comparative genomics, transcriptome profiling, and proteomics were employed to elucidate the molecular mechanisms of salt and osmotic stress tolerance. Salt-tolerant Frankia strains (CcI6 and Allo2) that could withstand up to 1000 mM NaCl and a salt-sensitive Frankia strain (CcI3) which could withstand only up to 475 mM NaCl were identified. The remaining isolates had intermediate levels of salt tolerance with MIC values ranging from 650 mM to 750 mM. Comparative genomic analysis showed that all of the Frankia isolates from Casuarina belonged to the same species (Frankia casuarinae). Pangenome analysis revealed a high abundance of singletons among all Casuarina isolates. The two salt-tolerant strains contained 153 shared single copy genes (most of which code for hypothetical proteins) that were not found in the salt-sensitive(CcI3) and moderately salt-tolerant (CeD) strains. RNA-seq analysis of one of the two salt-tolerant strains (Frankia sp. strain CcI6) revealed hundreds of genes differentially expressed under salt and/or osmotic stress. Among the 153 genes, 7 and 7 were responsive to salt and osmotic stress, respectively. Proteomic profiling confirmed the transcriptome results and identified 19 and 8 salt and/or osmotic stress-responsive proteins in the salt-tolerant (CcI6) and the salt-sensitive (CcI3) strains, respectively. Genetic differences between salt-tolerant and salt-sensitive Frankia strains isolated from Casuarina were identified. Transcriptome and proteome profiling of a salt-tolerant strain was used to determine molecular differences correlated with differential salt-tolerance and several candidate genes were identified. Mechanisms involving transcriptional and translational regulation, cell envelop remodeling, and previously uncharacterized proteins appear to be important for salt tolerance. Physiological and mutational analyses will further shed light on the molecular mechanism of salt tolerance in Casuarina associated Frankia isolates.

Transcript abundance on its own cannot be used to infer fluxes in central metabolism

DOE PAGES

Schwender, Jorg; Konig, Christina; Klapperstuck, Matthias; ...

2014-11-28

An attempt has been made to define the extent to which metabolic flux in central plant metabolism is reflected by changes in the transcriptome and metabolome, based on an analysis of in vitro cultured immature embryos of two oilseed rape (Brassica napus) accessions which contrast for seed lipid accumulation. Metabolic flux analysis (MFA) was used to constrain a flux balance metabolic model which included 671 biochemical and transport reactions within the central metabolism. This highly confident flux information was eventually used for comparative analysis of flux vs. transcript (metabolite). Metabolite profiling succeeded in identifying 79 intermediates within the central metabolism,more » some of which differed quantitatively between the two accessions and displayed a significant shift corresponding to flux. An RNA-Seq based transcriptome analysis revealed a large number of genes which were differentially transcribed in the two accessions, including some enzymes/proteins active in major metabolic pathways. With a few exceptions, differential activity in the major pathways (glycolysis, TCA cycle, amino acid, and fatty acid synthesis) was not reflected in contrasting abundances of the relevant transcripts. The conclusion was that transcript abundance on its own cannot be used to infer metabolic activity/fluxes in central plant metabolism. Lastly, this limitation needs to be borne in mind in evaluating transcriptome data and designing metabolic engineering experiments.« less

Alternative Splicing Profile and Sex-Preferential Gene Expression in the Female and Male Pacific Abalone Haliotis discus hannai.

PubMed

Kim, Mi Ae; Rhee, Jae-Sung; Kim, Tae Ha; Lee, Jung Sick; Choi, Ah-Young; Choi, Beom-Soon; Choi, Ik-Young; Sohn, Young Chang

2017-03-09

In order to characterize the female or male transcriptome of the Pacific abalone and further increase genomic resources, we sequenced the mRNA of full-length complementary DNA (cDNA) libraries derived from pooled tissues of female and male Haliotis discus hannai by employing the Iso-Seq protocol of the PacBio RSII platform. We successfully assembled whole full-length cDNA sequences and constructed a transcriptome database that included isoform information. After clustering, a total of 15,110 and 12,145 genes that coded for proteins were identified in female and male abalones, respectively. A total of 13,057 putative orthologs were retained from each transcriptome in abalones. Overall Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analyzed in each database showed a similar composition between sexes. In addition, a total of 519 and 391 isoforms were genome-widely identified with at least two isoforms from female and male transcriptome databases. We found that the number of isoforms and their alternatively spliced patterns are variable and sex-dependent. This information represents the first significant contribution to sex-preferential genomic resources of the Pacific abalone. The availability of whole female and male transcriptome database and their isoform information will be useful to improve our understanding of molecular responses and also for the analysis of population dynamics in the Pacific abalone.

Alternative Splicing Profile and Sex-Preferential Gene Expression in the Female and Male Pacific Abalone Haliotis discus hannai

PubMed Central

Kim, Mi Ae; Rhee, Jae-Sung; Kim, Tae Ha; Lee, Jung Sick; Choi, Ah-Young; Choi, Beom-Soon; Choi, Ik-Young; Sohn, Young Chang

2017-01-01

In order to characterize the female or male transcriptome of the Pacific abalone and further increase genomic resources, we sequenced the mRNA of full-length complementary DNA (cDNA) libraries derived from pooled tissues of female and male Haliotis discus hannai by employing the Iso-Seq protocol of the PacBio RSII platform. We successfully assembled whole full-length cDNA sequences and constructed a transcriptome database that included isoform information. After clustering, a total of 15,110 and 12,145 genes that coded for proteins were identified in female and male abalones, respectively. A total of 13,057 putative orthologs were retained from each transcriptome in abalones. Overall Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analyzed in each database showed a similar composition between sexes. In addition, a total of 519 and 391 isoforms were genome-widely identified with at least two isoforms from female and male transcriptome databases. We found that the number of isoforms and their alternatively spliced patterns are variable and sex-dependent. This information represents the first significant contribution to sex-preferential genomic resources of the Pacific abalone. The availability of whole female and male transcriptome database and their isoform information will be useful to improve our understanding of molecular responses and also for the analysis of population dynamics in the Pacific abalone. PMID:28282934

Systems perspectives on erythromycin biosynthesis by comparative genomic and transcriptomic analyses of S. erythraea E3 and NRRL23338 strains

PubMed Central

2013-01-01

Background S. erythraea is a Gram-positive filamentous bacterium used for the industrial-scale production of erythromycin A which is of high clinical importance. In this work, we sequenced the whole genome of a high-producing strain (E3) obtained by random mutagenesis and screening from the wild-type strain NRRL23338, and examined time-series expression profiles of both E3 and NRRL23338. Based on the genomic data and transcriptpmic data of these two strains, we carried out comparative analysis of high-producing strain and wild-type strain at both the genomic level and the transcriptomic level. Results We observed a large number of genetic variants including 60 insertions, 46 deletions and 584 single nucleotide variations (SNV) in E3 in comparison with NRRL23338, and the analysis of time series transcriptomic data indicated that the genes involved in erythromycin biosynthesis and feeder pathways were significantly up-regulated during the 60 hours time-course. According to our data, BldD, a previously identified ery cluster regulator, did not show any positive correlations with the expression of ery cluster, suggesting the existence of alternative regulation mechanisms of erythromycin synthesis in S. erythraea. Several potential regulators were then proposed by integration analysis of genomic and transcriptomic data. Conclusion This is a demonstration of the functional comparative genomics between an industrial S. erythraea strain and the wild-type strain. These findings help to understand the global regulation mechanisms of erythromycin biosynthesis in S. erythraea, providing useful clues for genetic and metabolic engineering in the future. PMID:23902230

Transcriptome analysis of sika deer in China.

PubMed

Jia, Bo-Yin; Ba, Heng-Xing; Wang, Gui-Wu; Yang, Ying; Cui, Xue-Zhe; Peng, Ying-Hua; Zheng, Jun-Jun; Xing, Xiu-Mei; Yang, Fu-He

2016-10-01

Sika deer is of great commercial value because their antlers are used in tonics and alternative medicine and their meat is healthy and delicious. The goal of this study was to generate transcript sequences from sika deer for functional genomic analyses and to identify the transcripts that demonstrate tissue-specific, age-dependent differential expression patterns. These sequences could enhance our understanding of the molecular mechanisms underlying sika deer growth and development. In the present study, we performed de novo transcriptome assembly and profiling analysis across ten tissue types and four developmental stages (juvenile, adolescent, adult, and aged) of sika deer, using Illumina paired-end tag (PET) sequencing technology. A total of 1,752,253 contigs with an average length of 799 bp were generated, from which 1,348,618 unigenes with an average length of 590 bp were defined. Approximately 33.2 % of these (447,931 unigenes) were then annotated in public protein databases. Many sika deer tissue-specific, age-dependent unigenes were identified. The testes have the largest number of tissue-enriched unigenes, and some of them were prone to develop new functions for other tissues. Additionally, our transcriptome revealed that the juvenile-adolescent transition was the most complex and important stage of the sika deer life cycle. The present work represents the first multiple tissue transcriptome analysis of sika deer across four developmental stages. The generated data not only provide a functional genomics resource for future biological research on sika deer but also guide the selection and manipulation of genes controlling growth and development.

Optimization Of A High-Throughput Transcriptomic (HTTr) Bioactivity Screen In MCF7 Cells Using Targeted RNA-Seq (SOT)

EPA Science Inventory

Recent advances in targeted RNA-Seq technology allow researchers to efficiently and cost-effectively obtain whole transcriptome profiles using picograms of mRNA from human cell lysates. Low mRNA input requirements and sample multiplexing capabilities has made time- and concentrat...

Obesity modulates inflammation and lipid metabolism oocyte gene expression: A single cell transcriptome perspective

USDA-ARS?s Scientific Manuscript database

This study aimed to compare oocyte gene expression profiles and follicular fluid (FF) content from overweight/obese (OW) women and normal weight (NW) women who were undergoing fertility treatments. Using single cell transcriptomic analyses, we investigated oocyte gene expression using RNA-seq. Serum...

The enhanced value of combining conventional and 'omics' analyses in early assessment of drug-induced hepatobiliary injury

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ellinger-Ziegelbauer, Heidrun, E-mail: heidrun.ellinger-ziegelbauer@bayerhealthcare.com; Adler, Melanie; Amberg, Alexander

2011-04-15

The InnoMed PredTox consortium was formed to evaluate whether conventional preclinical safety assessment can be significantly enhanced by incorporation of molecular profiling ('omics') technologies. In short-term toxicological studies in rats, transcriptomics, proteomics and metabolomics data were collected and analyzed in relation to routine clinical chemistry and histopathology. Four of the sixteen hepato- and/or nephrotoxicants given to rats for 1, 3, or 14 days at two dose levels induced similar histopathological effects. These were characterized by bile duct necrosis and hyperplasia and/or increased bilirubin and cholestasis, in addition to hepatocyte necrosis and regeneration, hepatocyte hypertrophy, and hepatic inflammation. Combined analysis ofmore » liver transcriptomics data from these studies revealed common gene expression changes which allowed the development of a potential sequence of events on a mechanistic level in accordance with classical endpoint observations. This included genes implicated in early stress responses, regenerative processes, inflammation with inflammatory cell immigration, fibrotic processes, and cholestasis encompassing deregulation of certain membrane transporters. Furthermore, a preliminary classification analysis using transcriptomics data suggested that prediction of cholestasis may be possible based on gene expression changes seen at earlier time-points. Targeted bile acid analysis, based on LC-MS metabonomics data demonstrating increased levels of conjugated or unconjugated bile acids in response to individual compounds, did not provide earlier detection of toxicity as compared to conventional parameters, but may allow distinction of different types of hepatobiliary toxicity. Overall, liver transcriptomics data delivered mechanistic and molecular details in addition to the classical endpoint observations which were further enhanced by targeted bile acid analysis using LC/MS metabonomics.« less

Transcriptomics-based analysis using RNA-Seq of the coconut (Cocos nucifera) leaf in response to yellow decline phytoplasma infection.

PubMed

Nejat, Naghmeh; Cahill, David M; Vadamalai, Ganesan; Ziemann, Mark; Rookes, James; Naderali, Neda

2015-10-01

Invasive phytoplasmas wreak havoc on coconut palms worldwide, leading to high loss of income, food insecurity and extreme poverty of farmers in producing countries. Phytoplasmas as strictly biotrophic insect-transmitted bacterial pathogens instigate distinct changes in developmental processes and defence responses of the infected plants and manipulate plants to their own advantage; however, little is known about the cellular and molecular mechanisms underlying host-phytoplasma interactions. Further, phytoplasma-mediated transcriptional alterations in coconut palm genes have not yet been identified. This study evaluated the whole transcriptome profiles of naturally infected leaves of Cocos nucifera ecotype Malayan Red Dwarf in response to yellow decline phytoplasma from group 16SrXIV, using RNA-Seq technique. Transcriptomics-based analysis reported here identified genes involved in coconut innate immunity. The number of down-regulated genes in response to phytoplasma infection exceeded the number of genes up-regulated. Of the 39,873 differentially expressed unigenes, 21,860 unigenes were suppressed and 18,013 were induced following infection. Comparative analysis revealed that genes associated with defence signalling against biotic stimuli were significantly overexpressed in phytoplasma-infected leaves versus healthy coconut leaves. Genes involving cell rescue and defence, cellular transport, oxidative stress, hormone stimulus and metabolism, photosynthesis reduction, transcription and biosynthesis of secondary metabolites were differentially represented. Our transcriptome analysis unveiled a core set of genes associated with defence of coconut in response to phytoplasma attack, although several novel defence response candidate genes with unknown function have also been identified. This study constitutes valuable sequence resource for uncovering the resistance genes and/or susceptibility genes which can be used as genetic tools in disease resistance breeding.

RNA-seq analysis of the gonadal transcriptome during Alligator mississippiensis temperature-dependent sex determination and differentiation.

PubMed

Yatsu, Ryohei; Miyagawa, Shinichi; Kohno, Satomi; Parrott, Benjamin B; Yamaguchi, Katsushi; Ogino, Yukiko; Miyakawa, Hitoshi; Lowers, Russell H; Shigenobu, Shuji; Guillette, Louis J; Iguchi, Taisen

2016-01-25

The American alligator (Alligator mississippiensis) displays temperature-dependent sex determination (TSD), in which incubation temperature during embryonic development determines the sexual fate of the individual. However, the molecular mechanisms governing this process remain a mystery, including the influence of initial environmental temperature on the comprehensive gonadal gene expression patterns occurring during TSD. Our characterization of transcriptomes during alligator TSD allowed us to identify novel candidate genes involved in TSD initiation. High-throughput RNA sequencing (RNA-seq) was performed on gonads collected from A. mississippiensis embryos incubated at both a male and a female producing temperature (33.5 °C and 30 °C, respectively) in a time series during sexual development. RNA-seq yielded 375.2 million paired-end reads, which were mapped and assembled, and used to characterize differential gene expression. Changes in the transcriptome occurring as a function of both development and sexual differentiation were extensively profiled. Forty-one differentially expressed genes were detected in response to incubation at male producing temperature, and included genes such as Wnt signaling factor WNT11, histone demethylase KDM6B, and transcription factor C/EBPA. Furthermore, comparative analysis of development- and sex-dependent differential gene expression revealed 230 candidate genes involved in alligator sex determination and differentiation, and early details of the suspected male-fate commitment were profiled. We also discovered sexually dimorphic expression of uncharacterized ncRNAs and other novel elements, such as unique expression patterns of HEMGN and ARX. Twenty-five of the differentially expressed genes identified in our analysis were putative transcriptional regulators, among which were MYBL2, MYCL, and HOXC10, in addition to conventional sex differentiation genes such as SOX9, and FOXL2. Inferred gene regulatory network was constructed, and the gene-gene and temperature-gene interactions were predicted. Gonadal global gene expression kinetics during sex determination has been extensively profiled for the first time in a TSD species. These findings provide insights into the genetic framework underlying TSD, and expand our current understanding of the developmental fate pathways during vertebrate sex determination.

Identifying potential RNAi targets in grain aphid (Sitobion avenae F.) based on transcriptome profiling of its alimentary canal after feeding on wheat plants.

PubMed

Zhang, Min; Zhou, Yuwen; Wang, Hui; Jones, Huw; Gao, Qiang; Wang, Dahai; Ma, Youzhi; Xia, Lanqin

2013-08-16

The grain aphid (Sitobion avenae F.) is a major agricultural pest which causes significant yield losses of wheat in China, Europe and North America annually. Transcriptome profiling of the grain aphid alimentary canal after feeding on wheat plants could provide comprehensive gene expression information involved in feeding, ingestion and digestion. Furthermore, selection of aphid-specific RNAi target genes would be essential for utilizing a plant-mediated RNAi strategy to control aphids via a non-toxic mode of action. However, due to the tiny size of the alimentary canal and lack of genomic information on grain aphid as a whole, selection of the RNAi targets is a challenging task that as far as we are aware, has never been documented previously. In this study, we performed de novo transcriptome assembly and gene expression analyses of the alimentary canals of grain aphids before and after feeding on wheat plants using Illumina RNA sequencing. The transcriptome profiling generated 30,427 unigenes with an average length of 664 bp. Furthermore, comparison of the transcriptomes of alimentary canals of pre- and post feeding grain aphids indicated that 5490 unigenes were differentially expressed, among which, diverse genes and/or pathways were identified and annotated. Based on the RPKM values of these unigenes, 16 of them that were significantly up or down-regulated upon feeding were selected for dsRNA artificial feeding assay. Of these, 5 unigenes led to higher mortality and developmental stunting in an artificial feeding assay due to the down-regulation of the target gene expression. Finally, by adding fluorescently labelled dsRNA into the artificial diet, the spread of fluorescence signal in the whole body tissues of grain aphid was observed. Comparison of the transcriptome profiles of the alimentary canals of pre- and post-feeding grain aphids on wheat plants provided comprehensive gene expression information that could facilitate our understanding of the molecular mechanisms underlying feeding, ingestion and digestion. Furthermore, five novel and effective potential RNAi target genes were identified in grain aphid for the first time. This finding would provide a fundamental basis for aphid control in wheat through plant mediated RNAi strategy.

Hepatic transcriptome analysis and identification of differentially expressed genes response to dietary oxidized fish oil in loach Misgurnus anguillicaudatus.

PubMed

Zhang, Yin; Li, Yang; Liang, Xiao; Cao, Xiaojuan; Huang, Longfei; Yan, Jie; Wei, Yanxing; Gao, Jian

2017-01-01

RNA sequencing and short-read assembly were utilized to produce a transcriptome of livers from loaches (Misgurnus anguillicaudatus) fed with three different diets respectively containing fresh fish oil (FO group), medium oxidized fish oil (MO group) and high oxidized fish oil (HO group). A total of 60,663 unigenes were obtained in this study, with mean length 848.74 bp. 50,814, 49,584 and 49,814 unigenes were respectively obtained from FO, MO and HO groups. There were 2,343 differentially expressed genes between FO and MO, with 855 down- and 1,488 up-regulated genes in the MO group. 2,813 genes were differentially expressed between FO and HO, including 1,256 down- and 1,552 up-regulated genes in the HO group. 2,075 differentially expressed genes were found in the comparison of MO and HO, including 1,074 up- and 1,001 down-regulated genes in the MO group. Some differentially expressed genes, such as fatty acid transport protein (fatp), fatty acid binding protein (fabp), apolipoprotein (apo), peroxisome proliferator activated receptor-gamma (ppar-γ), acetyl-CoA synthetase (acs) and arachidonate 5-lipoxygenase (alox5), were involved in lipid metabolism, suggesting these genes in the loach were responsive to dietary oxidized fish oil. Results of transcriptome profilings here were validated using quantitative real time PCR in fourteen randomly selected unigenes. The present study provides insights into hepatic transcriptome profile of the loach, which is a valuable resource for studies of loach genomics. More importantly, this study identifies some important genes responsible for dietary oxidized fish oil, which will benefit researches of lipid metabolism in fish.

Transcriptome Profiles of the Protoscoleces of Echinococcus granulosus Reveal that Excretory-Secretory Products Are Essential to Metabolic Adaptation

PubMed Central

Pan, Wei; Shen, Yujuan; Han, Xiuming; Wang, Ying; Liu, Hua; Jiang, Yanyan; Zhang, Yumei; Wang, Yanjuan; Xu, Yuxin; Cao, Jianping

2014-01-01

Background Cystic hydatid disease (CHD) is caused by the larval stages of the cestode and affects humans and domestic animals worldwide. Protoscoleces (PSCs) are one component of the larval stages that can interact with both definitive and intermediate hosts. Previous genomic and transcriptomic data have provided an overall snapshot of the genomics of the growth and development of this parasite. However, our understanding of how PSCs subvert the immune response of hosts and maintains metabolic adaptation remains unclear. In this study, we used Roche 454 sequencing technology and in silico secretome analysis to explore the transcriptome profiles of the PSCs from E. granulosus and elucidate the potential functions of the excretory-secretory proteins (ESPs) released by the parasite. Methodology/Principal Findings A large number of nonredundant sequences as unigenes were generated (26,514), of which 22,910 (86.4%) were mapped to the newly published E. granulosus genome and 17,705 (66.8%) were distributed within the coding sequence (CDS) regions. Of the 2,280 ESPs predicted from the transcriptome, 138 ESPs were inferred to be involved in the metabolism of carbohydrates, while 124 ESPs were inferred to be involved in the metabolism of protein. Eleven ESPs were identified as intracellular enzymes that regulate glycolysis/gluconeogenesis (GL/GN) pathways, while a further 44 antigenic proteins, 25 molecular chaperones and four proteases were highly represented. Many proteins were also found to be significantly enriched in development-related signaling pathways, such as the TGF-β receptor pathways and insulin pathways. Conclusions/Significance This study provides valuable information on the metabolic adaptation of parasites to their hosts that can be used to aid the development of novel intervention targets for hydatid treatment and control. PMID:25500817

Transcriptomic Analysis of Persistent Infection with Foot-and-Mouth Disease Virus in Cattle Suggests Impairment of Apoptosis and Cell-Mediated Immunity in the Nasopharynx

DOE PAGES

Eschbaumer, Michael; Stenfeldt, Carolina; Smoliga, George R.; ...

2016-09-19

In order to investigate the mechanisms of persistent foot-and-mouth disease virus (FMDV) infection in cattle, transcriptome alterations associated with the FMDV carrier state were characterized using a bovine whole-transcriptome microarray. Eighteen cattle (8 vaccinated with a recombinant FMDV A vaccine, 10 non-vaccinated) were challenged with FMDV A 24 Cruzeiro, and the gene expression profiles of nasopharyngeal tissues collected between 21 and 35 days after challenge were compared between 11 persistently infected carriers and 7 non-carriers. Carriers and non-carrierswere further compared to 2 naive animals that had been neither vaccinated nor challenged. At a controlled false-discovery rate of 10% and amore » minimum difference in expression of 50%, 648 genes were differentially expressed between FMDV carriers and non-carriers, and most (467) had higher expression in carriers.Among these, genes associated with cellular proliferation and the immune response–such as chemokines, cytokines and genes regulating T and B cells–were significantly over represented. Differential gene expression was significantly correlated between non-vaccinated and vaccinated animals (biological correlation +0.97), indicating a similar transcriptome profile across these groups. Genes related to prostaglandin E 2 production and the induction of regulatoryT cells were over expressed in carriers. In contrast, tissues from non-carrier animals expressed higher levels of complement regulators and pro-apoptotic genes that could promote virus clearance. Furthermore, based on these findings, we propose a working hypothesis for FMDV persistence in nasopharyngeal tissues of cattle, in which the virus may be maintained by an impairment of apoptosis and the local suppression of cell-mediated antiviral immunity by inducible regulatoryT cells.« less

Transcriptomic Analysis of Persistent Infection with Foot-and-Mouth Disease Virus in Cattle Suggests Impairment of Apoptosis and Cell-Mediated Immunity in the Nasopharynx

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eschbaumer, Michael; Stenfeldt, Carolina; Smoliga, George R.

In order to investigate the mechanisms of persistent foot-and-mouth disease virus (FMDV) infection in cattle, transcriptome alterations associated with the FMDV carrier state were characterized using a bovine whole-transcriptome microarray. Eighteen cattle (8 vaccinated with a recombinant FMDV A vaccine, 10 non-vaccinated) were challenged with FMDV A 24 Cruzeiro, and the gene expression profiles of nasopharyngeal tissues collected between 21 and 35 days after challenge were compared between 11 persistently infected carriers and 7 non-carriers. Carriers and non-carrierswere further compared to 2 naive animals that had been neither vaccinated nor challenged. At a controlled false-discovery rate of 10% and amore » minimum difference in expression of 50%, 648 genes were differentially expressed between FMDV carriers and non-carriers, and most (467) had higher expression in carriers.Among these, genes associated with cellular proliferation and the immune response–such as chemokines, cytokines and genes regulating T and B cells–were significantly over represented. Differential gene expression was significantly correlated between non-vaccinated and vaccinated animals (biological correlation +0.97), indicating a similar transcriptome profile across these groups. Genes related to prostaglandin E 2 production and the induction of regulatoryT cells were over expressed in carriers. In contrast, tissues from non-carrier animals expressed higher levels of complement regulators and pro-apoptotic genes that could promote virus clearance. Furthermore, based on these findings, we propose a working hypothesis for FMDV persistence in nasopharyngeal tissues of cattle, in which the virus may be maintained by an impairment of apoptosis and the local suppression of cell-mediated antiviral immunity by inducible regulatoryT cells.« less

Integration of Transcriptome, Proteome and Metabolism Data Reveals the Alkaloids Biosynthesis in Macleaya cordata and Macleaya microcarpa

PubMed Central

Liu, Fuqing; Huang, Peng; Zhu, Pengcheng; Chen, Jinjun; Shi, Mingming; Guo, Fang; Cheng, Pi; Zeng, Jing; Liao, Yifang; Gong, Jing; Zhang, Hong-Mei; Wang, Depeng; Guo, An-Yuan; Xiong, Xingyao

2013-01-01

Background The Macleaya spp., including Macleaya cordata and Macleaya microcarpa, are traditional anti-virus, inflammation eliminating, and insecticide herb medicines for their isoquinoline alkaloids. They are also known as the basis of the popular natural animal food addictive in Europe. However, few studies especially at genomics level were conducted on them. Hence, we performed the Macleaya spp. transcriptome and integrated it with iTRAQ proteome analysis in order to identify potential genes involved in alkaloids biosynthesis. Methodology and Principal Findings We elaborately designed the transcriptome, proteome and metabolism profiling for 10 samples of both species to explore their alkaloids biosynthesis. From the transcriptome data, we obtained 69367 and 78255 unigenes for M. cordata and M. microcarpa, in which about two thirds of them were similar to sequences in public databases. By metabolism profiling, reverse patterns for alkaloids sanguinarine, chelerythrine, protopine, and allocryptopine were observed in different organs of two species. We characterized the expressions of enzymes in alkaloid biosynthesis pathways. We also identified more than 1000 proteins from iTRAQ proteome data. Our results strongly suggest that the root maybe the organ for major alkaloids biosynthesis of Macleaya spp. Except for biosynthesis, the alkaloids storage and transport were also important for their accumulation. The ultrastructure of laticifers by SEM helps us to prove the alkaloids maybe accumulated in the mature roots. Conclusions/Significance To our knowledge this is the first study to elucidate the genetic makeup of Macleaya spp. This work provides clues to the identification of the potential modulate genes involved in alkaloids biosynthesis in Macleaya spp., and sheds light on researches for non-model medicinal plants by integrating different high-throughput technologies. PMID:23326424

High-throughput full-length single-cell mRNA-seq of rare cells.

PubMed

Ooi, Chin Chun; Mantalas, Gary L; Koh, Winston; Neff, Norma F; Fuchigami, Teruaki; Wong, Dawson J; Wilson, Robert J; Park, Seung-Min; Gambhir, Sanjiv S; Quake, Stephen R; Wang, Shan X

2017-01-01

Single-cell characterization techniques, such as mRNA-seq, have been applied to a diverse range of applications in cancer biology, yielding great insight into mechanisms leading to therapy resistance and tumor clonality. While single-cell techniques can yield a wealth of information, a common bottleneck is the lack of throughput, with many current processing methods being limited to the analysis of small volumes of single cell suspensions with cell densities on the order of 107 per mL. In this work, we present a high-throughput full-length mRNA-seq protocol incorporating a magnetic sifter and magnetic nanoparticle-antibody conjugates for rare cell enrichment, and Smart-seq2 chemistry for sequencing. We evaluate the efficiency and quality of this protocol with a simulated circulating tumor cell system, whereby non-small-cell lung cancer cell lines (NCI-H1650 and NCI-H1975) are spiked into whole blood, before being enriched for single-cell mRNA-seq by EpCAM-functionalized magnetic nanoparticles and the magnetic sifter. We obtain high efficiency (> 90%) capture and release of these simulated rare cells via the magnetic sifter, with reproducible transcriptome data. In addition, while mRNA-seq data is typically only used for gene expression analysis of transcriptomic data, we demonstrate the use of full-length mRNA-seq chemistries like Smart-seq2 to facilitate variant analysis of expressed genes. This enables the use of mRNA-seq data for differentiating cells in a heterogeneous population by both their phenotypic and variant profile. In a simulated heterogeneous mixture of circulating tumor cells in whole blood, we utilize this high-throughput protocol to differentiate these heterogeneous cells by both their phenotype (lung cancer versus white blood cells), and mutational profile (H1650 versus H1975 cells), in a single sequencing run. This high-throughput method can help facilitate single-cell analysis of rare cell populations, such as circulating tumor or endothelial cells, with demonstrably high-quality transcriptomic data.

Combined venomics, antivenomics and venom gland transcriptome analysis of the monocoled cobra (Naja kaouthia) from China.

PubMed

Xu, Ning; Zhao, Hong-Yan; Yin, Yin; Shen, Shan-Shan; Shan, Lin-Lin; Chen, Chuan-Xi; Zhang, Yan-Xia; Gao, Jian-Fang; Ji, Xiang

2017-04-21

We conducted an omics-analysis of the venom of Naja kaouthia from China. Proteomics analysis revealed six protein families [three-finger toxins (3-FTx), phospholipase A 2 (PLA 2 ), nerve growth factor, snake venom metalloproteinase (SVMP), cysteine-rich secretory protein and ohanin], and venom-gland transcriptomics analysis revealed 28 protein families from 79 unigenes. 3-FTx (56.5% in proteome/82.0% in transcriptome) and PLA 2 (26.9%/13.6%) were identified as the most abundant families in venom proteome and venom-gland transcriptome. Furthermore, N. kaouthia venom expressed strong lethality (i.p. LD 50 : 0.79μg/g) and myotoxicity (CK: 5939U/l) in mice, and showed notable activity in PLA 2 but weak activity in SVMP, l-amino acid oxidase or 5' nucleotidase. Antivenomic assessment revealed that several venom components (nearly 17.5% of total venom) from N. kaouthia could not be thoroughly immunocaptured by commercial Naja atra antivenom. ELISA analysis revealed that there was no difference in the cross-reaction between N. kaouthia and N. atra venoms against the N. atra antivenom. The use of commercial N. atra antivenom in treatment of snakebites caused by N. kaouthia is reasonable, but design of novel antivenom with the attention on enhancing the immune response of non-immunocaptured components should be encouraged. The venomics, antivenomics and venom-gland transcriptome of the monocoled cobra (Naja kaouthia) from China have been elucidated. Quantitative and qualitative differences are evident when venom proteomic and venom-gland transcriptomic profiles are compared. Two protein families (3-FTx and PLA 2 ) are found to be the predominated components in N. kaouthia venom, and considered as the major players in functional role of venom. Other protein families with relatively low abundance appear to be minor in the functional significance. Antivenomics and ELISA evaluation reveal that the N. kaouthia venom can be effectively immunorecognized by commercial N. atra antivenom, but still a small number of venom components could not be thoroughly immunocaptured. The findings indicate that exploring the precise composition of snake venom should be executed by an integrated omics-approach, and elucidating the venom composition is helpful in understanding composition-function relationships and will facilitate the clinical application of antivenoms. Copyright © 2017 Elsevier B.V. All rights reserved.

Quantitative high-throughput profiling of snake venom gland transcriptomes and proteomes (Ovophis okinavensis and Protobothrops flavoviridis)

PubMed Central

2013-01-01

Background Advances in DNA sequencing and proteomics have facilitated quantitative comparisons of snake venom composition. Most studies have employed one approach or the other. Here, both Illumina cDNA sequencing and LC/MS were used to compare the transcriptomes and proteomes of two pit vipers, Protobothrops flavoviridis and Ovophis okinavensis, which differ greatly in their biology. Results Sequencing of venom gland cDNA produced 104,830 transcripts. The Protobothrops transcriptome contained transcripts for 103 venom-related proteins, while the Ovophis transcriptome contained 95. In both, transcript abundances spanned six orders of magnitude. Mass spectrometry identified peptides from 100% of transcripts that occurred at higher than contaminant (e.g. human keratin) levels, including a number of proteins never before sequenced from snakes. These transcriptomes reveal fundamentally different envenomation strategies. Adult Protobothrops venom promotes hemorrhage, hypotension, incoagulable blood, and prey digestion, consistent with mammalian predation. Ovophis venom composition is less readily interpreted, owing to insufficient pharmacological data for venom serine and metalloproteases, which comprise more than 97.3% of Ovophis transcripts, but only 38.0% of Protobothrops transcripts. Ovophis venom apparently represents a hybrid strategy optimized for frogs and small mammals. Conclusions This study illustrates the power of cDNA sequencing combined with MS profiling. The former quantifies transcript composition, allowing detection of novel proteins, but cannot indicate which proteins are actually secreted, as does MS. We show, for the first time, that transcript and peptide abundances are correlated. This means that MS can be used for quantitative, non-invasive venom profiling, which will be beneficial for studies of endangered species. PMID:24224955

Combined Analysis of the Fruit Metabolome and Transcriptome Reveals Candidate Genes Involved in Flavonoid Biosynthesis in Actinidia arguta.

PubMed

Li, Yukuo; Fang, Jinbao; Qi, Xiujuan; Lin, Miaomiao; Zhong, Yunpeng; Sun, Leiming; Cui, Wen

2018-05-15

To assess the interrelation between the change of metabolites and the change of fruit color, we performed a combined metabolome and transcriptome analysis of the flesh in two different Actinidia arguta cultivars: "HB" ("Hongbaoshixing") and "YF" ("Yongfengyihao") at two different fruit developmental stages: 70d (days after full bloom) and 100d (days after full bloom). Metabolite and transcript profiling was obtained by ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometer and high-throughput RNA sequencing, respectively. The identification and quantification results of metabolites showed that a total of 28,837 metabolites had been obtained, of which 13,715 were annotated. In comparison of HB100 vs. HB70, 41 metabolites were identified as being flavonoids, 7 of which, with significant difference, were identified as bracteatin, luteolin, dihydromyricetin, cyanidin, pelargonidin, delphinidin and (-)-epigallocatechin. Association analysis between metabolome and transcriptome revealed that there were two metabolic pathways presenting significant differences during fruit development, one of which was flavonoid biosynthesis, in which 14 structural genes were selected to conduct expression analysis, as well as 5 transcription factor genes obtained by transcriptome analysis. RT-qPCR results and cluster analysis revealed that AaF3H , AaLDOX , AaUFGT , AaMYB , AabHLH , and AaHB2 showed the best possibility of being candidate genes. A regulatory network of flavonoid biosynthesis was established to illustrate differentially expressed candidate genes involved in accumulation of metabolites with significant differences, inducing red coloring during fruit development. Such a regulatory network linking genes and flavonoids revealed a system involved in the pigmentation of all-red-fleshed and all-green-fleshed A. arguta , suggesting this conjunct analysis approach is not only useful in understanding the relationship between genotype and phenotype, but is also a powerful tool for providing more valuable information for breeding.

The Role of Vitamin D in the Transcriptional Program of Human Pregnancy

PubMed Central

Al-Garawi, Amal; Carey, Vincent J.; Chhabra, Divya; Morrow, Jarrett; Lasky-Su, Jessica; Qiu, Weiliang; Laranjo, Nancy; Litonjua, Augusto A.; Weiss, Scott T.

2016-01-01

Background Patterns of gene expression of human pregnancy are poorly understood. In a trial of vitamin D supplementation in pregnant women, peripheral blood transcriptomes were measured longitudinally on 30 women and used to characterize gene co-expression networks. Objective Studies suggest that increased maternal Vitamin D levels may reduce the risk of asthma in early life, yet the underlying mechanisms have not been examined. In this study, we used a network-based approach to examine changes in gene expression profiles during the course of normal pregnancy and evaluated their association with maternal Vitamin D levels. Design The VDAART study is a randomized clinical trial of vitamin D supplementation in pregnancy for reduction of pediatric asthma risk. The trial enrolled 881 women at 10–18 weeks of gestation. Longitudinal gene expression measures were obtained on thirty pregnant women, using RNA isolated from peripheral blood samples obtained in the first and third trimesters. Differentially expressed genes were identified using significance of analysis of microarrays (SAM), and clustered using a weighted gene co-expression network analysis (WGCNA). Gene-set enrichment was performed to identify major biological pathways. Results Comparison of transcriptional profiles between first and third trimesters of pregnancy identified 5839 significantly differentially expressed genes (FDR<0.05). Weighted gene co-expression network analysis clustered these transcripts into 14 co-expression modules of which two showed significant correlation with maternal vitamin D levels. Pathway analysis of these two modules revealed genes enriched in immune defense pathways and extracellular matrix reorganization as well as genes enriched in notch signaling and transcription factor networks. Conclusion Our data show that gene expression profiles of healthy pregnant women change during the course of pregnancy and suggest that maternal Vitamin D levels influence transcriptional profiles. These alterations of the maternal transcriptome may contribute to fetal immune imprinting and reduce allergic sensitization in early life. Trial Registration clinicaltrials.gov NCT00920621 PMID:27711190

Genomic and transcriptomic predictors of triglyceride response to regular exercise

PubMed Central

Sarzynski, Mark A; Davidsen, Peter K; Sung, Yun Ju; Hesselink, Matthijs K C; Schrauwen, Patrick; Rice, Treva K; Rao, D C; Falciani, Francesco; Bouchard, Claude

2015-01-01

Aim We performed genome-wide and transcriptome-wide profiling to identify genes and single nucleotide polymorphisms (SNPs) associated with the response of triglycerides (TG) to exercise training. Methods Plasma TG levels were measured before and after a 20-week endurance training programme in 478 white participants from the HERITAGE Family Study. Illumina HumanCNV370-Quad v3.0 BeadChips were genotyped using the Illumina BeadStation 500GX platform. Affymetrix HG-U133+2 arrays were used to quantitate gene expression levels from baseline muscle biopsies of a subset of participants (N=52). Genome-wide association study (GWAS) analysis was performed using MERLIN, while transcriptomic predictor models were developed using the R-package GALGO. Results The GWAS results showed that eight SNPs were associated with TG training-response (ΔTG) at p<9.9×10−6, while another 31 SNPs showed p values <1×10−4. In multivariate regression models, the top 10 SNPs explained 32.0% of the variance in ΔTG, while conditional heritability analysis showed that four SNPs statistically accounted for all of the heritability of ΔTG. A molecular signature based on the baseline expression of 11 genes predicted 27% of ΔTG in HERITAGE, which was validated in an independent study. A composite SNP score based on the top four SNPs, each from the genomic and transcriptomic analyses, was the strongest predictor of ΔTG (R2=0.14, p=3.0×10−68). Conclusions Our results indicate that skeletal muscle transcript abundance at 11 genes and SNPs at a number of loci contribute to TG response to exercise training. Combining data from genomics and transcriptomics analyses identified a SNP-based gene signature that should be further tested in independent samples. PMID:26491034

Transcriptome Profiling of Wheat Inflorescence Development from Spikelet Initiation to Floral Patterning Identified Stage-Specific Regulatory Genes1[OPEN

PubMed Central

Feng, Nan; Song, Gaoyuan; Guan, Jiantao; Chen, Kai; Jia, Meiling; Huang, Dehua; Wu, Jiajie; Zhang, Lichao; Kong, Xiuying; Geng, Shuaifeng

2017-01-01

Early reproductive development in cereals is crucial for final grain number per spike and hence the yield potential of the crop. To date, however, no systematic analyses of gene expression profiles during this important process have been conducted for common wheat (Triticum aestivum). Here, we studied the transcriptome profiles at four stages of early wheat reproductive development, from spikelet initiation to floral organ differentiation. K-means clustering and stage-specific transcript identification detected dynamically expressed homeologs of important transcription regulators in spikelet and floral meristems that may be involved in spikelet initiation, floret meristem specification, and floral organ patterning, as inferred from their homologs in model plants. Small RNA transcriptome sequencing discovered key microRNAs that were differentially expressed during wheat inflorescence development alongside their target genes, suggesting that miRNA-mediated regulatory mechanisms for floral development may be conserved in cereals and Arabidopsis. Our analysis was further substantiated by the functional characterization of the ARGONAUTE1d (AGO1d) gene, which was initially expressed in stamen primordia and later in the tapetum during anther maturation. In agreement with its stage-specific expression pattern, the loss of function of the predominantly expressed B homeolog of AGO1d in a tetraploid durum wheat mutant resulted in smaller anthers with more infertile pollens than the wild type and a reduced grain number per spike. Together, our work provides a first glimpse of the gene regulatory networks in wheat inflorescence development that may be pivotal for floral and grain development, highlighting potential targets for genetic manipulation to improve future wheat yields. PMID:28515146

Assessing the accuracy of blood RNA profiles to identify patients with post-concussion syndrome: A pilot study in a military patient population.

PubMed

Hardy, Jimmaline J; Mooney, Scott R; Pearson, Andrea N; McGuire, Dawn; Correa, Daniel J; Simon, Roger P; Meller, Robert

2017-01-01

Mild traumatic brain injury (mTBI) is a complex, neurophysiological condition that can have detrimental outcomes. Yet, to date, no objective method of diagnosis exists. Physical damage to the blood-brain-barrier and normal waste clearance via the lymphatic system may enable the detection of biomarkers of mTBI in peripheral circulation. Here we evaluate the accuracy of whole transcriptome analysis of blood to predict the clinical diagnosis of post-concussion syndrome (PCS) in a military cohort. Sixty patients with clinically diagnosed chronic concussion and controls (no history of concussion) were recruited (retrospective study design). Male patients (46) were split into a training set comprised of 20 long-term concussed (> 6 months and symptomatic) and 12 controls (no documented history of concussion). Models were validated in a testing set (control = 9, concussed = 5). RNA_Seq libraries were prepared from whole blood samples for sequencing using a SOLiD5500XL sequencer and aligned to hg19 reference genome. Patterns of differential exon expression were used for diagnostic modeling using support vector machine classification, and then validated in a second patient cohort. The accuracy of RNA profiles to predict the clinical diagnosis of post-concussion syndrome patients from controls was 86% (sensitivity 80%; specificity 89%). In addition, RNA profiles reveal duration of concussion. This pilot study shows the potential utility of whole transcriptome analysis to establish the clinical diagnosis of chronic concussion syndrome.

Transcriptomic Response of Porcine PBMCs to Vaccination with Tetanus Toxoid as a Model Antigen

PubMed Central

Adler, Marcel; Murani, Eduard; Brunner, Ronald; Ponsuksili, Siriluck; Wimmers, Klaus

2013-01-01

The aim of the present study was to characterize in vivo genome-wide transcriptional responses to immune stimulation in order to get insight into the resulting changes of allocation of resources. Vaccination with tetanus toxoid was used as a model for a mixed Th1 and Th2 immune response in pig. Expression profiles of PBMCs (peripheral blood mononuclear cells) before and at 12 time points over a period of four weeks after initial and booster vaccination at day 14 were studied by use of Affymetrix GeneChip microarrays and Ingenuity Pathway Analysis (IPA). The transcriptome data in total comprised more than 5000 genes with different transcript abundances (DE-genes). Within the single time stages the numbers of DE-genes were between several hundred and more than 1000. Ingenuity Pathway Analysis mainly revealed canonical pathways of cellular immune response and cytokine signaling as well as a broad range of processes in cellular and organismal growth, proliferation and development, cell signaling, biosynthesis and metabolism. Significant changes in the expression profiles of PBMCs already occurred very early after immune stimulation. At two hours after the first vaccination 679 DE-genes corresponding to 110 canonical pathways of cytokine signaling, cellular immune response and other multiple cellular functions were found. Immune competence and global disease resistance are heritable but difficult to measure and to address by breeding. Besides QTL mapping of immune traits gene expression profiling facilitates the detection of functional gene networks and thus functional candidate genes. PMID:23536793

Transcriptomic response of porcine PBMCs to vaccination with tetanus toxoid as a model antigen.

PubMed

Adler, Marcel; Murani, Eduard; Brunner, Ronald; Ponsuksili, Siriluck; Wimmers, Klaus

2013-01-01

The aim of the present study was to characterize in vivo genome-wide transcriptional responses to immune stimulation in order to get insight into the resulting changes of allocation of resources. Vaccination with tetanus toxoid was used as a model for a mixed Th1 and Th2 immune response in pig. Expression profiles of PBMCs (peripheral blood mononuclear cells) before and at 12 time points over a period of four weeks after initial and booster vaccination at day 14 were studied by use of Affymetrix GeneChip microarrays and Ingenuity Pathway Analysis (IPA). The transcriptome data in total comprised more than 5000 genes with different transcript abundances (DE-genes). Within the single time stages the numbers of DE-genes were between several hundred and more than 1000. Ingenuity Pathway Analysis mainly revealed canonical pathways of cellular immune response and cytokine signaling as well as a broad range of processes in cellular and organismal growth, proliferation and development, cell signaling, biosynthesis and metabolism. Significant changes in the expression profiles of PBMCs already occurred very early after immune stimulation. At two hours after the first vaccination 679 DE-genes corresponding to 110 canonical pathways of cytokine signaling, cellular immune response and other multiple cellular functions were found. Immune competence and global disease resistance are heritable but difficult to measure and to address by breeding. Besides QTL mapping of immune traits gene expression profiling facilitates the detection of functional gene networks and thus functional candidate genes.

Transcriptome sequencing and microarray development for the woodrat (Neotoma spp.): custom genetic tools for exploring herbivore ecology.

PubMed

Malenke, J R; Milash, B; Miller, A W; Dearing, M D

2013-07-01

Massively parallel sequencing has enabled the creation of novel, in-depth genetic tools for nonmodel, ecologically important organisms. We present the de novo transcriptome sequencing, analysis and microarray development for a vertebrate herbivore, the woodrat (Neotoma spp.). This genus is of ecological and evolutionary interest, especially with respect to ingestion and hepatic metabolism of potentially toxic plant secondary compounds. We generated a liver transcriptome of the desert woodrat (Neotoma lepida) using the Roche 454 platform. The assembled contigs were well annotated using rodent references (99.7% annotation), and biotransformation function was reflected in the gene ontology. The transcriptome was used to develop a custom microarray (eArray, Agilent). We tested the microarray with three experiments: one across species with similar habitat (thus, dietary) niches, one across species with different habitat niches and one across populations within a species. The resulting one-colour arrays had high technical and biological quality. Probes designed from the woodrat transcriptome performed significantly better than functionally similar probes from the Norway rat (Rattus norvegicus). There were a multitude of expression differences across the woodrat treatments, many of which related to biotransformation processes and activities. The pattern and function of the differences indicate shared ecological pressures, and not merely phylogenetic distance, play an important role in shaping gene expression profiles of woodrat species and populations. The quality and functionality of the woodrat transcriptome and custom microarray suggest these tools will be valuable for expanding the scope of herbivore biology, as well as the exploration of conceptual topics in ecology. © 2013 John Wiley & Sons Ltd.

Comparative Transcriptome Analysis Reveals Critical Function of Sucrose Metabolism Related-Enzymes in Starch Accumulation in the Storage Root of Sweet Potato

PubMed Central

Zhang, Kai; Wu, Zhengdan; Tang, Daobin; Luo, Kai; Lu, Huixiang; Liu, Yingying; Dong, Jie; Wang, Xin; Lv, Changwen; Wang, Jichun; Lu, Kun

2017-01-01

The starch properties of the storage root (SR) affect the quality of sweet potato (Ipomoea batatas (L.) Lam.). Although numerous studies have analyzed the accumulation and properties of starch in sweet potato SRs, the transcriptomic variation associated with starch properties in SR has not been quantified. In this study, we measured the starch and sugar contents and analyzed the transcriptome profiles of SRs harvested from sweet potatoes with high, medium, and extremely low starch contents, at five developmental stages [65, 80, 95, 110, and 125 days after transplanting (DAP)]. We found that differences in both water content and starch accumulation in the dry matter affect the starch content of SRs in different sweet potato genotypes. Based on transcriptome sequencing data, we assembled 112336 unigenes, and identified several differentially expressed genes (DEGs) involved in starch and sucrose metabolism, and revealed the transcriptional regulatory network controlling starch and sucrose metabolism in sweet potato SRs. Correlation analysis between expression patterns and starch and sugar contents suggested that the sugar–starch conversion steps catalyzed by sucrose synthase (SuSy) and UDP-glucose pyrophosphorylase (UGPase) may be essential for starch accumulation in the dry matter of SRs, and IbβFRUCT2, a vacuolar acid invertase, might also be a key regulator of starch content in the SRs. Our results provide valuable resources for future investigations aimed at deciphering the molecular mechanisms determining the starch properties of sweet potato SRs. PMID:28690616

Molecular profiles of Quadriceps muscle in myostatin-null mice reveal PI3K and apoptotic pathways as myostatin targets

PubMed Central

Chelh, Ilham; Meunier, Bruno; Picard, Brigitte; Reecy, Mark James; Chevalier, Catherine; Hocquette, Jean-François; Cassar-Malek, Isabelle

2009-01-01

Background Myostatin (MSTN), a member of the TGF-β superfamily, has been identified as a negative regulator of skeletal muscle mass. Inactivating mutations in the MSTN gene are responsible for the development of a hypermuscular phenotype. In this study, we performed transcriptomic and proteomic analyses to detect altered expression/abundance of genes and proteins. These differentially expressed genes and proteins may represent new molecular targets of MSTN and could be involved in the regulation of skeletal muscle mass. Results Transcriptomic analysis of the Quadriceps muscles of 5-week-old MSTN-null mice (n = 4) and their controls (n = 4) was carried out using microarray (human and murine oligonucleotide sequences) of 6,473 genes expressed in muscle. Proteomic profiles were analysed using two-dimensional gel electrophoresis coupled with mass spectrometry. Comparison of the transcriptomic profiles revealed 192 up- and 245 down- regulated genes. Genes involved in the PI3K pathway, insulin/IGF pathway, carbohydrate metabolism and apoptosis regulation were up-regulated. Genes belonging to canonical Wnt, calcium signalling pathways and cytokine-receptor cytokine interaction were down-regulated. Comparison of the protein profiles revealed 20 up- and 18 down-regulated proteins spots. Knockout of the MSTN gene was associated with up-regulation of proteins involved in glycolytic shift of the muscles and down-regulation of proteins involved in oxidative energy metabolism. In addition, an increased abundance of survival/anti-apoptotic factors were observed. Conclusion All together, these results showed a differential expression of genes and proteins related to the muscle energy metabolism and cell survival/anti-apoptotic pathway (e.g. DJ-1, PINK1, 14-3-3ε protein, TCTP/GSK-3β). They revealed the PI3K and apoptotic pathways as MSTN targets and are in favour of a role of MSTN as a modulator of cell survival in vivo. PMID:19397818

Conjunctival transcriptome profiling of Solomon Islanders with active trachoma in the absence of Chlamydia trachomatis infection.

PubMed

Vasileva, Hristina; Butcher, Robert; Pickering, Harry; Sokana, Oliver; Jack, Kelvin; Solomon, Anthony W; Holland, Martin J; Roberts, Chrissy H

2018-02-21

Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Islands, but the majority of these individuals have no serological evidence of previous infection with Chlamydia trachomatis. In Temotu and Rennell and Bellona provinces, ocular infections with C. trachomatis were seldom detected among children with active trachoma; a similar lack of association was seen between active trachoma and other common bacterial and viral causes of follicular conjunctivitis. Here, we set out to characterise patterns of gene expression at the conjunctivae of children in these provinces with and without clinical signs of trachomatous inflammation-follicular (TF) and C. trachomatis infection. Purified RNA from children with and without active trachoma was run on Affymetrix GeneChip Human Transcriptome Array 2.0 microarrays. Profiles were compared between individuals with ocular C. trachomatis infection and TF (group DI; n = 6), individuals with TF but no C. trachomatis infection (group D; n = 7), and individuals without TF or C. trachomatis infection (group N; n = 7). Differential gene expression and gene set enrichment for pathway membership were assessed. Conjunctival gene expression profiles were more similar within-group than between-group. Principal components analysis indicated that the first and second principal components combined explained almost 50% of the variance in the dataset. When comparing the DI group to the N group, genes involved in T-cell proliferation, B-cell signalling and CD8+ T cell signalling pathways were differentially regulated. When comparing the DI group to the D group, CD8+ T-cell regulation, interferon-gamma and IL17 production pathways were enriched. Genes involved in RNA transcription and translation pathways were upregulated when comparing the D group to the N group. Gene expression profiles in children in the Solomon Islands indicate immune responses consistent with bacterial infection when TF and C. trachomatis infection are concurrent. The transcriptomes of children with TF but without identified infection were not consistent with allergic or viral conjunctivitis.

ARG1 Functions in the Physiological Adaptation of Undifferentiated Plant Cells to Spaceflight.

PubMed

Zupanska, Agata K; Schultz, Eric R; Yao, JiQiang; Sng, Natasha J; Zhou, Mingqi; Callaham, Jordan B; Ferl, Robert J; Paul, Anna-Lisa

2017-11-01

Scientific access to spaceflight and especially the International Space Station has revealed that physiological adaptation to spaceflight is accompanied or enabled by changes in gene expression that significantly alter the transcriptome of cells in spaceflight. A wide range of experiments have shown that plant physiological adaptation to spaceflight involves gene expression changes that alter cell wall and other metabolisms. However, while transcriptome profiling aptly illuminates changes in gene expression that accompany spaceflight adaptation, mutation analysis is required to illuminate key elements required for that adaptation. Here we report how transcriptome profiling was used to gain insight into the spaceflight adaptation role of Altered response to gravity 1 (Arg1), a gene known to affect gravity responses in plants on Earth. The study compared expression profiles of cultured lines of Arabidopsis thaliana derived from wild-type (WT) cultivar Col-0 to profiles from a knock-out line deficient in the gene encoding ARG1 (ARG1 KO), both on the ground and in space. The cell lines were launched on SpaceX CRS-2 as part of the Cellular Expression Logic (CEL) experiment of the BRIC-17 spaceflight mission. The cultured cell lines were grown within 60 mm Petri plates in Petri Dish Fixation Units (PDFUs) that were housed within the Biological Research In Canisters (BRIC) hardware. Spaceflight samples were fixed on orbit. Differentially expressed genes were identified between the two environments (spaceflight and comparable ground controls) and the two genotypes (WT and ARG1 KO). Each genotype engaged unique genes during physiological adaptation to the spaceflight environment, with little overlap. Most of the genes altered in expression in spaceflight in WT cells were found to be Arg1-dependent, suggesting a major role for that gene in the physiological adaptation of undifferentiated cells to spaceflight. Key Words: ARG1-Spaceflight-Gene expression-Physiological adaptation-BRIC. Astrobiology 17, 1077-1111.

De novo sequencing and comparative transcriptome analysis of the male and hermaphroditic flowers provide insights into the regulation of flower formation in andromonoecious taihangia rupestris.

PubMed

Li, Weiguo; Zhang, Lihui; Ding, Zhan; Wang, Guodong; Zhang, Yandi; Gong, Hongmei; Chang, Tianjun; Zhang, Yanwen

2017-02-28

Taihangia rupestris, an andromonoecious plant species, bears both male and hermaphroditic flowers within the same individual. However, the establishment and development of male and hermaphroditic flowers in andromonoecious Taihangia remain poorly understood, due to the limited genetic and sequence information. To investigate the potential molecular mechanism in the regulation of Taihangia flower formation, we used de novo RNA sequencing to compare the transcriptome profiles of male and hermaphroditic flowers at early and late developmental stages. Four cDNA libraries, including male floral bud, hermaphroditic floral bud, male flower, and hermaphroditic flower, were constructed and sequenced by using the Illumina RNA-Seq method. Totally, 84,596,426 qualified Illumina reads were obtained and then assembled into 59,064 unigenes, of which 24,753 unigenes were annotated in the NCBI non-redundant protein database. In addition, 12,214, 7,153, and 8,115 unigenes were assigned into 53 Gene Ontology (GO) functional groups, 25 Clusters of Orthologous Group (COG) categories, and 126 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, respectively. By pairwise comparison of unigene abundance between the samples, we identified 1,668 differential expressed genes (DEGs), including 176 transcription factors (TFs) between the male and hermaphroditic flowers. At the early developmental stage, we found 263 up-regulated genes and 436 down-regulated genes expressed in hermaphroditic floral buds, while 844 up-regulated genes and 314 down-regulated genes were detected in hermaphroditic flowers at the late developmental stage. GO and KEGG enrichment analyses showed that a large number of DEGs were associated with a wide range of functions, including cell cycle, epigenetic processes, flower development, and biosynthesis of unsaturated fatty acid pathway. Finally, real-time quantitative PCR was conducted to validate the DEGs identified in the present study. In this study, transcriptome data of this rare andromonoecious Taihangia were reported for the first time. Comparative transcriptome analysis revealed the significant differences in gene expression profiles between male and hermaphroditic flowers at early and late developmental stages. The transcriptome data of Taihangia would be helpful to improve the understanding of the underlying molecular mechanisms in regulation of flower formation and unisexual flower establishment in andromonoecious plants.

De novo transcriptome profiling of cold-stressed siliques during pod filling stages in Indian mustard (Brassica juncea L.)

PubMed Central

Sinha, Somya; Raxwal, Vivek K.; Joshi, Bharat; Jagannath, Arun; Katiyar-Agarwal, Surekha; Goel, Shailendra; Kumar, Amar; Agarwal, Manu

2015-01-01

Low temperature is a major abiotic stress that impedes plant growth and development. Brassica juncea is an economically important oil seed crop and is sensitive to freezing stress during pod filling subsequently leading to abortion of seeds. To understand the cold stress mediated global perturbations in gene expression, whole transcriptome of B. juncea siliques that were exposed to sub-optimal temperature was sequenced. Manually self-pollinated siliques at different stages of development were subjected to either short (6 h) or long (12 h) durations of chilling stress followed by construction of RNA-seq libraries and deep sequencing using Illumina's NGS platform. De-novo assembly of B. juncea transcriptome resulted in 133,641 transcripts, whose combined length was 117 Mb and N50 value was 1428 bp. We identified 13,342 differentially regulated transcripts by pair-wise comparison of 18 transcriptome libraries. Hierarchical clustering along with Spearman correlation analysis identified that the differentially expressed genes segregated in two major clusters representing early (5–15 DAP) and late stages (20–30 DAP) of silique development. Further analysis led to the discovery of sub-clusters having similar patterns of gene expression. Two of the sub-clusters (one each from the early and late stages) comprised of genes that were inducible by both the durations of cold stress. Comparison of transcripts from these clusters led to identification of 283 transcripts that were commonly induced by cold stress, and were referred to as “core cold-inducible” transcripts. Additionally, we found that 689 and 100 transcripts were specifically up-regulated by cold stress in early and late stages, respectively. We further explored the expression patterns of gene families encoding for transcription factors (TFs), transcription regulators (TRs) and kinases, and found that cold stress induced protein kinases only during early silique development. We validated the digital gene expression profiles of selected transcripts by qPCR and found a high degree of concordance between the two analyses. To our knowledge this is the first report of transcriptome sequencing of cold-stressed B. juncea siliques. The data generated in this study would be a valuable resource for not only understanding the cold stress signaling pathway but also for introducing cold hardiness in B. juncea. PMID:26579175

De novo assembly and characterization of fruit transcriptome in Litchi chinensis Sonn and analysis of differentially regulated genes in fruit in response to shading

PubMed Central

2013-01-01

Background Litchi (Litchi chinensis Sonn.) is one of the most important fruit trees cultivated in tropical and subtropical areas. However, a lack of transcriptomic and genomic information hinders our understanding of the molecular mechanisms underlying fruit set and fruit development in litchi. Shading during early fruit development decreases fruit growth and induces fruit abscission. Here, high-throughput RNA sequencing (RNA-Seq) was employed for the de novo assembly and characterization of the fruit transcriptome in litchi, and differentially regulated genes, which are responsive to shading, were also investigated using digital transcript abundance(DTA)profiling. Results More than 53 million paired-end reads were generated and assembled into 57,050 unigenes with an average length of 601 bp. These unigenes were annotated by querying against various public databases, with 34,029 unigenes found to be homologous to genes in the NCBI GenBank database and 22,945 unigenes annotated based on known proteins in the Swiss-Prot database. In further orthologous analyses, 5,885 unigenes were assigned with one or more Gene Ontology terms, 10,234 hits were aligned to the 24 Clusters of Orthologous Groups classifications and 15,330 unigenes were classified into 266 Kyoto Encyclopedia of Genes and Genomes pathways. Based on the newly assembled transcriptome, the DTA profiling approach was applied to investigate the differentially expressed genes related to shading stress. A total of 3.6 million and 3.5 million high-quality tags were generated from shaded and non-shaded libraries, respectively. As many as 1,039 unigenes were shown to be significantly differentially regulated. Eleven of the 14 differentially regulated unigenes, which were randomly selected for more detailed expression comparison during the course of shading treatment, were identified as being likely to be involved in the process of fruitlet abscission in litchi. Conclusions The assembled transcriptome of litchi fruit provides a global description of expressed genes in litchi fruit development, and could serve as an ideal repository for future functional characterization of specific genes. The DTA analysis revealed that more than 1000 differentially regulated unigenes respond to the shading signal, some of which might be involved in the fruitlet abscission process in litchi, shedding new light on the molecular mechanisms underlying organ abscission. PMID:23941440

Transcriptional signature associated with early rheumatoid arthritis and healthy individuals at high risk to develop the disease

PubMed Central

Macías-Segura, N.; Bastian, Y.; Santiago-Algarra, D.; Castillo-Ortiz, J. D.; Alemán-Navarro, A. L.; Jaime-Sánchez, E.; Gomez-Moreno, M.; Saucedo-Toral, C. A.; Lara-Ramírez, Edgar E.; Zapata-Zuñiga, M.; Enciso-Moreno, L.; González-Amaro, R.; Ramos-Remus, C.; Enciso-Moreno, J. A.

2018-01-01

Background Little is known regarding the mechanisms underlying the loss of tolerance in the early and preclinical stages of autoimmune diseases. The aim of this work was to identify the transcriptional profile and signaling pathways associated to non-treated early rheumatoid arthritis (RA) and subjects at high risk. Several biomarker candidates for early RA are proposed. Methods Whole blood total RNA was obtained from non-treated early RA patients with <1 year of evolution as well as from healthy first-degree relatives of patients with RA (FDR) classified as ACCP+ and ACCP- according to their antibodies serum levels against cyclic citrullinated peptides. Complementary RNA (cRNA) was synthetized and hybridized to high-density microarrays. Data was analyzed in Genespring Software and functional categories were assigned to a specific transcriptome identified in subjects with RA and FDR ACCP positive. Specific signaling pathways for genes associated to RA were identified. Gene expression was evaluated by qPCR. Receiver operating characteristic (ROC) analysis was used to evaluate these genes as biomarkers. Results A characteristic transcriptome of 551 induced genes and 4,402 repressed genes were identified in early RA patients. Bioinformatics analysis of the data identified a specific transcriptome in RA patients. Moreover, some overlapped transcriptional profiles between patients with RA and ACCP+ were identified, suggesting an up-regulated distinctive transcriptome from the preclinical stages up to progression to an early RA state. A total of 203 pathways have up-regulated genes that are shared between RA and ACCP+. Some of these genes show potential to be used as progression biomarkers for early RA with area under the curve of ROC > 0.92. These genes come from several functional categories associated to inflammation, Wnt signaling and type I interferon pathways. Conclusion The presence of a specific transcriptome in whole blood of RA patients suggests the activation of a specific inflammatory transcriptional signature in early RA development. The set of overexpressed genes in early RA patients that are shared with ACCP+ subjects but not with ACCP- subjects, can represent a transcriptional signature involved with the transition of a preclinical to a clinical RA stage. Some of these particular up-regulated and down-regulated genes are related to inflammatory processes and could be considered as biomarker candidates for disease progression in subjects at risk to develop RA. PMID:29584756

Gene Expression Profiles of Chlamydophila pneumoniae during the Developmental Cycle and Iron Depletion–Mediated Persistence

PubMed Central

Mäurer, André P; Mehlitz, Adrian; Mollenkopf, Hans J; Meyer, Thomas F

2007-01-01

The obligate intracellular, gram-negative bacterium Chlamydophila pneumoniae (Cpn) has impact as a human pathogen. Little is known about changes in the Cpn transcriptome during its biphasic developmental cycle (the acute infection) and persistence. The latter stage has been linked to chronic diseases. To analyze Cpn CWL029 gene expression, we designed a pathogen-specific oligo microarray and optimized the extraction method for pathogen RNA. Throughout the acute infection, ratio expression profiles for each gene were generated using 48 h post infection as a reference. Based on these profiles, significantly expressed genes were separated into 12 expression clusters using self-organizing map clustering and manual sorting into the “early”, “mid”, “late”, and “tardy” cluster classes. The latter two were differentiated because the “tardy” class showed steadily increasing expression at the end of the cycle. The transcriptome of the Cpn elementary body (EB) and published EB proteomics data were compared to the cluster profile of the acute infection. We found an intriguing association between “late” genes and genes coding for EB proteins, whereas “tardy” genes were mainly associated with genes coding for EB mRNA. It has been published that iron depletion leads to Cpn persistence. We compared the gene expression profiles during iron depletion–mediated persistence with the expression clusters of the acute infection. This led to the finding that establishment of iron depletion–mediated persistence is more likely a mid-cycle arrest in development rather than a completely distinct gene expression pattern. Here, we describe the Cpn transcriptome during the acute infection, differentiating “late” genes, which correlate to EB proteins, and “tardy” genes, which lead to EB mRNA. Expression profiles during iron mediated–persistence led us to propose the hypothesis that the transcriptomic “clock” is arrested during acute mid-cycle. PMID:17590080

RNASeq-based genome annotation and identification of long-noncoding RNAs in the grapevine cultivar 'Riesling'

USDA-ARS?s Scientific Manuscript database

The technological advances of RNA-seq and de novo transcriptome assembly have enabled genome annotation and transcriptome profiling in heterozygous species. This is a promising approach to improving the annotation of the reference genome sequence of grapevine (Vitis vinifera L.), a species of high-l...

Comprehensive transcriptome profiling reveals long noncoding RNA expression and alternative splicing regulation during fruit development and ripening in kiwifruit (Actinidia chinensis)

USDA-ARS?s Scientific Manuscript database

Genomic and transcriptomic data on kiwifruit (Actinidia chinensis) in public databases are very limited despite its nutritional and economic value. Previously, we have constructed and sequenced nine fruit RNA-Seq libraries of A. chinensis cv. 'Hongyang' at immature, mature, and postharvest ripening...

Comparative Transcriptome Profiling of Rice Near-Isogenic Line Carrying Xa23 under Infection of Xanthomonas oryzae pv. oryzae.

PubMed

Tariq, Rezwan; Wang, Chunlian; Qin, Tengfei; Xu, Feifei; Tang, Yongchao; Gao, Ying; Ji, Zhiyuan; Zhao, Kaijun

2018-03-02

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae ( Xoo ), is an overwhelming disease in rice-growing regions worldwide. Our previous studies revealed that the executor R gene Xa23 confers broad-spectrum disease resistance to all naturally occurring biotypes of Xoo . In this study, comparative transcriptomic profiling of two near-isogenic lines (NILs), CBB23 (harboring Xa23 ) and JG30 (without Xa23 ), before and after infection of the Xoo strain, PXO99 A , was done by RNA sequencing, to identify genes associated with the resistance. After high throughput sequencing, 1645 differentially expressed genes (DEGs) were identified between CBB23 and JG30 at different time points. Gene Ontlogy (GO) analysis categorized the DEGs into biological process, molecular function, and cellular component. KEGG analysis categorized the DEGs into different pathways, and phenylpropanoid biosynthesis was the most prominent pathway, followed by biosynthesis of plant hormones, flavonoid biosynthesis, and glycolysis/gluconeogenesis. Further analysis led to the identification of differentially expressed transcription factors (TFs) and different kinase responsive genes in CBB23, than that in JG30. Besides TFs and kinase responsive genes, DEGs related to ethylene, jasmonic acid, and secondary metabolites were also identified in both genotypes after PXO99 A infection. The data of DEGs are a precious resource for further clarifying the network of Xa23 -mediated resistance.

Comparative Transcriptome Profiling of Rice Near-Isogenic Line Carrying Xa23 under Infection of Xanthomonas oryzae pv. oryzae

PubMed Central

Tariq, Rezwan; Wang, Chunlian; Qin, Tengfei; Xu, Feifei; Tang, Yongchao; Gao, Ying; Ji, Zhiyuan; Zhao, Kaijun

2018-01-01

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo), is an overwhelming disease in rice-growing regions worldwide. Our previous studies revealed that the executor R gene Xa23 confers broad-spectrum disease resistance to all naturally occurring biotypes of Xoo. In this study, comparative transcriptomic profiling of two near-isogenic lines (NILs), CBB23 (harboring Xa23) and JG30 (without Xa23), before and after infection of the Xoo strain, PXO99A, was done by RNA sequencing, to identify genes associated with the resistance. After high throughput sequencing, 1645 differentially expressed genes (DEGs) were identified between CBB23 and JG30 at different time points. Gene Ontlogy (GO) analysis categorized the DEGs into biological process, molecular function, and cellular component. KEGG analysis categorized the DEGs into different pathways, and phenylpropanoid biosynthesis was the most prominent pathway, followed by biosynthesis of plant hormones, flavonoid biosynthesis, and glycolysis/gluconeogenesis. Further analysis led to the identification of differentially expressed transcription factors (TFs) and different kinase responsive genes in CBB23, than that in JG30. Besides TFs and kinase responsive genes, DEGs related to ethylene, jasmonic acid, and secondary metabolites were also identified in both genotypes after PXO99A infection. The data of DEGs are a precious resource for further clarifying the network of Xa23-mediated resistance. PMID:29498672

Identification of the Neuromuscular Junction Transcriptome of Extraocular Muscle by Laser Capture Microdissection

PubMed Central

Ketterer, Caroline; Zeiger, Ulrike; Budak, Murat T.; Rubinstein, Neal A.; Khurana, Tejvir S.

2010-01-01

Purpose. To examine and characterize the profile of genes expressed at the synapses or neuromuscular junctions (NMJs) of extraocular muscles (EOMs) compared with those expressed at the tibialis anterior (TA). Methods. Adult rat eyeballs with rectus EOMs attached and TAs were dissected, snap frozen, serially sectioned, and stained for acetylcholinesterase (AChE) to identify the NMJs. Approximately 6000 NMJs for rectus EOM (EOMsyn), 6000 NMJs for TA (TAsyn), equal amounts of NMJ-free fiber regions (EOMfib, TAfib), and underlying myonuclei and RNAs were captured by laser capture microdissection (LCM). RNA was processed for microarray-based expression profiling. Expression profiles and interaction lists were generated for genes differentially expressed at synaptic and nonsynaptic regions of EOM (EOMsyn versus EOMfib) and TA (TAsyn versus TAfib). Profiles were validated by using real-time quantitative polymerase chain reaction (qPCR). Results. The regional transcriptomes associated with NMJs of EOMs and TAs were identified. Two hundred seventy-five genes were preferentially expressed in EOMsyn (compared with EOMfib), 230 in TAsyn (compared with TAfib), and 288 additional transcripts expressed in both synapses. Identified genes included novel genes as well as well-known, evolutionarily conserved synaptic markers (e.g., nicotinic acetylcholine receptor (AChR) alpha (Chrna) and epsilon (Chrne) subunits and nestin (Nes). Conclusions. Transcriptome level differences exist between EOM synaptic regions and TA synaptic regions. The definition of the synaptic transcriptome provides insight into the mechanism of formation and functioning of the unique synapses of EOM and their differential involvement in diseases noted in the EOM allotype. PMID:20393109

Nasopharyngeal Microbiota, Host Transcriptome, and Disease Severity in Children with Respiratory Syncytial Virus Infection.

PubMed

de Steenhuijsen Piters, Wouter A A; Heinonen, Santtu; Hasrat, Raiza; Bunsow, Eleonora; Smith, Bennett; Suarez-Arrabal, Maria-Carmen; Chaussabel, Damien; Cohen, Daniel M; Sanders, Elisabeth A M; Ramilo, Octavio; Bogaert, Debby; Mejias, Asuncion

2016-11-01

Respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory tract infections and hospitalizations in infants worldwide. Known risk factors, however, incompletely explain the variability of RSV disease severity, especially among healthy children. We postulate that the severity of RSV infection is influenced by modulation of the host immune response by the local bacterial ecosystem. To assess whether specific nasopharyngeal microbiota (clusters) are associated with distinct host transcriptome profiles and disease severity in children less than 2 years of age with RSV infection. We characterized the nasopharyngeal microbiota profiles of young children with mild and severe RSV disease and healthy children by 16S-rRNA sequencing. In parallel, using multivariable models, we analyzed whole-blood transcriptome profiles to study the relationship between microbial community composition, the RSV-induced host transcriptional response, and clinical disease severity. We identified five nasopharyngeal microbiota clusters characterized by enrichment of either Haemophilus influenzae, Streptococcus, Corynebacterium, Moraxella, or Staphylococcus aureus. RSV infection and RSV hospitalization were positively associated with H. influenzae and Streptococcus and negatively associated with S. aureus abundance, independent of age. Children with RSV showed overexpression of IFN-related genes, independent of the microbiota cluster. In addition, transcriptome profiles of children with RSV infection and H. influenzae- and Streptococcus-dominated microbiota were characterized by greater overexpression of genes linked to Toll-like receptor and by neutrophil and macrophage activation and signaling. Our data suggest that interactions between RSV and nasopharyngeal microbiota might modulate the host immune response, potentially affecting clinical disease severity.

Genome-scale analysis and comparison of gene expression profiles in developing and germinated pollen in Oryza sativa

PubMed Central

2010-01-01

Background Pollen development from the microspore involves a series of coordinated cellular events, and the resulting mature pollen has a specialized function to quickly germinate, produce a polar-growth pollen tube derived from the vegetative cell, and deliver two sperm cells into the embryo sac for double fertilization. The gene expression profiles of developing and germinated pollen have been characterised by use of the eudicot model plant Arabidopsis. Rice, one of the most important cereal crops, has been used as an excellent monocot model. A comprehensive analysis of transcriptome profiles of developing and germinated pollen in rice is important to understand the conserved and diverse mechanism underlying pollen development and germination in eudicots and monocots. Results We used Affymetrix GeneChip® Rice Genome Array to comprehensively analyzed the dynamic changes in the transcriptomes of rice pollen at five sequential developmental stages from microspores to germinated pollen. Among the 51,279 transcripts on the array, we found 25,062 pollen-preferential transcripts, among which 2,203 were development stage-enriched. The diversity of transcripts decreased greatly from microspores to mature and germinated pollen, whereas the number of stage-enriched transcripts displayed a "U-type" change, with the lowest at the bicellular pollen stage; and a transition of overrepresented stage-enriched transcript groups associated with different functional categories, which indicates a shift in gene expression program at the bicellular pollen stage. About 54% of the now-annotated rice F-box protein genes were expressed preferentially in pollen. The transcriptome profile of germinated pollen was significantly and positively correlated with that of mature pollen. Analysis of expression profiles and coexpressed features of the pollen-preferential transcripts related to cell cycle, transcription, the ubiquitin/26S proteasome system, phytohormone signalling, the kinase system and defense/stress response revealed five expression patterns, which are compatible with changes in major cellular events during pollen development and germination. A comparison of pollen transcriptomes between rice and Arabidopsis revealed that 56.6% of the rice pollen preferential genes had homologs in Arabidopsis genome, but 63.4% of these homologs were expressed, with a small proportion being expressed preferentially, in Arabidopsis pollen. Rice and Arabidopsis pollen had non-conservative transcription factors each. Conclusions Our results demonstrated that rice pollen expressed a set of reduced but specific transcripts in comparison with vegetative tissues, and the number of stage-enriched transcripts displayed a "U-type" change during pollen development, with the lowest at the bicellular pollen stage. These features are conserved in rice and Arabidopsis. The shift in gene expression program at the bicellular pollen stage may be important to the transition from earlier cell division to later pollen maturity. Pollen at maturity pre-synthesized transcripts needed for germination and early pollen tube growth. The transcription regulation associated with pollen development would have divergence between the two species. Our results also provide novel insights into the molecular program and key components of the regulatory network regulating pollen development and germination. PMID:20507633

Single-lineage transcriptome analysis reveals key regulatory pathways in primitive erythroid progenitors in the mouse embryo

PubMed Central

Isern, Joan; He, Zhiyong; Fraser, Stuart T.; Nowotschin, Sonja; Ferrer-Vaquer, Anna; Moore, Rebecca; Hadjantonakis, Anna-Katerina; Schulz, Vincent; Tuck, David; Gallagher, Patrick G.

2011-01-01

Primitive erythroid (EryP) progenitors are the first cell type specified from the mesoderm late in gastrulation. We used a transgenic reporter to image and purify the earliest blood progenitors and their descendants from developing mouse embryos. EryP progenitors exhibited remarkable proliferative capacity in the yolk sac immediately before the onset of circulation, when these cells comprise nearly half of all cells of the embryo. Global expression profiles generated at 24-hour intervals from embryonic day 7.5 through 2.5 revealed 2 abrupt changes in transcript diversity that coincided with the entry of EryPs into the circulation and with their late maturation and enucleation, respectively. These changes were paralleled by the expression of critical regulatory factors. Experiments designed to test predictions from these data demonstrated that the Wnt-signaling pathway is active in EryP progenitors, which display an aerobic glycolytic profile and the numbers of which are regulated by transforming growth factor-β1 and hypoxia. This is the first transcriptome assembled for a single hematopoietic lineage of the embryo over the course of its differentiation. PMID:21263157

Global transcriptomic profiling of aspen trees under elevated [CO2] to identify potential molecular mechanisms responsible for enhanced radial growth.

PubMed

Wei, Hairong; Gou, Jiqing; Yordanov, Yordan; Zhang, Huaxin; Thakur, Ramesh; Jones, Wendy; Burton, Andrew

2013-03-01

Aspen (Populus tremuloides) trees growing under elevated [CO(2)] at a free-air CO(2) enrichment (FACE) site produced significantly more biomass than control trees. We investigated the molecular mechanisms underlying the observed increase in biomass by producing transcriptomic profiles of the vascular cambium zone (VCZ) and leaves, and then performed a comparative study to identify significantly changed genes and pathways after 12 years exposure to elevated [CO(2)]. In leaves, elevated [CO(2)] enhanced expression of genes related to Calvin cycle activity and linked pathways. In the VCZ, the pathways involved in cell growth, cell division, hormone metabolism, and secondary cell wall formation were altered while auxin conjugation, ABA synthesis, and cytokinin glucosylation and degradation were inhibited. Similarly, the genes involved in hemicellulose and pectin biosynthesis were enhanced, but some genes that catalyze important steps in lignin biosynthesis pathway were inhibited. Evidence from systemic analysis supported the functioning of multiple molecular mechanisms that underpin the enhanced radial growth in response to elevated [CO(2)].

Transcriptome profiling reveals mosaic genomic origins of modern cultivated barley.

PubMed

Dai, Fei; Chen, Zhong-Hua; Wang, Xiaolei; Li, Zefeng; Jin, Gulei; Wu, Dezhi; Cai, Shengguan; Wang, Ning; Wu, Feibo; Nevo, Eviatar; Zhang, Guoping

2014-09-16

The domestication of cultivated barley has been used as a model system for studying the origins and early spread of agrarian culture. Our previous results indicated that the Tibetan Plateau and its vicinity is one of the centers of domestication of cultivated barley. Here we reveal multiple origins of domesticated barley using transcriptome profiling of cultivated and wild-barley genotypes. Approximately 48-Gb of clean transcript sequences in 12 Hordeum spontaneum and 9 Hordeum vulgare accessions were generated. We reported 12,530 de novo assembled transcripts in all of the 21 samples. Population structure analysis showed that Tibetan hulless barley (qingke) might have existed in the early stage of domestication. Based on the large number of unique genomic regions showing the similarity between cultivated and wild-barley groups, we propose that the genomic origin of modern cultivated barley is derived from wild-barley genotypes in the Fertile Crescent (mainly in chromosomes 1H, 2H, and 3H) and Tibet (mainly in chromosomes 4H, 5H, 6H, and 7H). This study indicates that the domestication of barley may have occurred over time in geographically distinct regions.

Profiling the transcriptome of Gracilaria changii (Rhodophyta) in response to light deprivation.

PubMed

Ho, Chai-Ling; Teoh, Seddon; Teo, Swee-Sen; Rahim, Raha Abdul; Phang, Siew-Moi

2009-01-01

Light regulates photosynthesis, growth and reproduction, yield and properties of phycocolloids, and starch contents in seaweeds. Despite its importance as an environmental cue that regulates many developmental, physiological, and biochemical processes, the network of genes involved during light deprivation are obscure. In this study, we profiled the transcriptome of Gracilaria changii at two different irradiance levels using a cDNA microarray containing more than 3,000 cDNA probes. Microarray analysis revealed that 93 and 105 genes were up- and down-regulated more than 3-fold under light deprivation, respectively. However, only 50% of the transcripts have significant matches to the nonredundant peptide sequences in the database. The transcripts that accumulated under light deprivation include vanadium chloroperoxidase, thioredoxin, ferredoxin component, and reduced nicotinamide adenine dinucleotide dehydrogenase. Among the genes that were down-regulated under light deprivation were genes encoding light harvesting protein, light harvesting complex I, phycobilisome 7.8 kDa linker polypeptide, low molecular weight early light-inducible protein, and vanadium bromoperoxidase. Our findings also provided important clues to the functions of many unknown sequences that could not be annotated using sequence comparison.

A cell-based systems biology assessment of human blood to monitor immune responses after influenza vaccination.

PubMed

Hoek, Kristen L; Samir, Parimal; Howard, Leigh M; Niu, Xinnan; Prasad, Nripesh; Galassie, Allison; Liu, Qi; Allos, Tara M; Floyd, Kyle A; Guo, Yan; Shyr, Yu; Levy, Shawn E; Joyce, Sebastian; Edwards, Kathryn M; Link, Andrew J

2015-01-01

Systems biology is an approach to comprehensively study complex interactions within a biological system. Most published systems vaccinology studies have utilized whole blood or peripheral blood mononuclear cells (PBMC) to monitor the immune response after vaccination. Because human blood is comprised of multiple hematopoietic cell types, the potential for masking responses of under-represented cell populations is increased when analyzing whole blood or PBMC. To investigate the contribution of individual cell types to the immune response after vaccination, we established a rapid and efficient method to purify human T and B cells, natural killer (NK) cells, myeloid dendritic cells (mDC), monocytes, and neutrophils from fresh venous blood. Purified cells were fractionated and processed in a single day. RNA-Seq and quantitative shotgun proteomics were performed to determine expression profiles for each cell type prior to and after inactivated seasonal influenza vaccination. Our results show that transcriptomic and proteomic profiles generated from purified immune cells differ significantly from PBMC. Differential expression analysis for each immune cell type also shows unique transcriptomic and proteomic expression profiles as well as changing biological networks at early time points after vaccination. This cell type-specific information provides a more comprehensive approach to monitor vaccine responses.

A Comparative Analysis of Industrial Escherichia coli K–12 and B Strains in High-Glucose Batch Cultivations on Process-, Transcriptome- and Proteome Level

PubMed Central

Marisch, Karoline; Bayer, Karl; Scharl, Theresa; Mairhofer, Juergen; Krempl, Peter M.; Hummel, Karin; Razzazi-Fazeli, Ebrahim; Striedner, Gerald

2013-01-01

Escherichia coli K–12 and B strains are among the most frequently used bacterial hosts for production of recombinant proteins on an industrial scale. To improve existing processes and to accelerate bioprocess development, we performed a detailed host analysis. We investigated the different behaviors of the E. coli production strains BL21, RV308, and HMS174 in response to high-glucose concentrations. Tightly controlled cultivations were conducted under defined environmental conditions for the in-depth analysis of physiological behavior. In addition to acquisition of standard process parameters, we also used DNA microarray analysis and differential gel electrophoresis (EttanTM DIGE). Batch cultivations showed different yields of the distinct strains for cell dry mass and growth rate, which were highest for BL21. In addition, production of acetate, triggered by excess glucose supply, was much higher for the K–12 strains compared to the B strain. Analysis of transcriptome data showed significant alteration in 347 of 3882 genes common among all three hosts. These differentially expressed genes included, for example, those involved in transport, iron acquisition, and motility. The investigation of proteome patterns additionally revealed a high number of differentially expressed proteins among the investigated hosts. The subsequently selected 38 spots included proteins involved in transport and motility. The results of this comprehensive analysis delivered a full genomic picture of the three investigated strains. Differentially expressed groups for targeted host modification were identified like glucose transport or iron acquisition, enabling potential optimization of strains to improve yield and process quality. Dissimilar growth profiles of the strains confirm different genotypes. Furthermore, distinct transcriptome patterns support differential regulation at the genome level. The identified proteins showed high agreement with the transcriptome data and suggest similar regulation within a host at both levels for the identified groups. Such host attributes need to be considered in future process design and operation. PMID:23950949

A comparative analysis of industrial Escherichia coli K-12 and B strains in high-glucose batch cultivations on process-, transcriptome- and proteome level.

PubMed

Marisch, Karoline; Bayer, Karl; Scharl, Theresa; Mairhofer, Juergen; Krempl, Peter M; Hummel, Karin; Razzazi-Fazeli, Ebrahim; Striedner, Gerald

2013-01-01

Escherichia coli K-12 and B strains are among the most frequently used bacterial hosts for production of recombinant proteins on an industrial scale. To improve existing processes and to accelerate bioprocess development, we performed a detailed host analysis. We investigated the different behaviors of the E. coli production strains BL21, RV308, and HMS174 in response to high-glucose concentrations. Tightly controlled cultivations were conducted under defined environmental conditions for the in-depth analysis of physiological behavior. In addition to acquisition of standard process parameters, we also used DNA microarray analysis and differential gel electrophoresis (Ettan(TM) DIGE). Batch cultivations showed different yields of the distinct strains for cell dry mass and growth rate, which were highest for BL21. In addition, production of acetate, triggered by excess glucose supply, was much higher for the K-12 strains compared to the B strain. Analysis of transcriptome data showed significant alteration in 347 of 3882 genes common among all three hosts. These differentially expressed genes included, for example, those involved in transport, iron acquisition, and motility. The investigation of proteome patterns additionally revealed a high number of differentially expressed proteins among the investigated hosts. The subsequently selected 38 spots included proteins involved in transport and motility. The results of this comprehensive analysis delivered a full genomic picture of the three investigated strains. Differentially expressed groups for targeted host modification were identified like glucose transport or iron acquisition, enabling potential optimization of strains to improve yield and process quality. Dissimilar growth profiles of the strains confirm different genotypes. Furthermore, distinct transcriptome patterns support differential regulation at the genome level. The identified proteins showed high agreement with the transcriptome data and suggest similar regulation within a host at both levels for the identified groups. Such host attributes need to be considered in future process design and operation.

Mitochondria, oligodendrocytes and inflammation in bipolar disorder: evidence from transcriptome studies points to intriguing parallels with multiple sclerosis

PubMed Central

Konradi, Christine; Sillivan, Stephanie E.; Clay, Hayley B.

2011-01-01

Gene expression studies of bipolar disorder (BPD) have shown changes in transcriptome profiles in multiple brain regions. Here we summarize the most consistent findings in the scientific literature, and compare them to data from schizophrenia (SZ) and major depressive disorder (MDD). The transcriptome profiles of all three disorders overlap, making the existence of a BPD-specific profile unlikely. Three groups of functionally related genes are consistently expressed at altered levels in BPD, SZ and MDD. Genes involved in energy metabolism and mitochondrial function are downregulated, genes involved in immune response and inflammation are upregulated, and genes expressed in oligodendrocytes are downregulated. Experimental paradigms for multiple sclerosis demonstrate a tight link between energy metabolism, inflammation and demyelination. These studies also show variabilities in the extent of oligodendrocyte stress, which can vary from a downregulation of oligodendrocyte genes, such as observed in psychiatric disorders, to cell death and brain lesions seen in multiple sclerosis. We conclude that experimental models of multiple sclerosis could be of interest for the research of BPD, SZ and MDD. PMID:21310238

Single cell transcriptomics of neighboring hyphae of Aspergillus niger

PubMed Central

2011-01-01

Single cell profiling was performed to assess differences in RNA accumulation in neighboring hyphae of the fungus Aspergillus niger. A protocol was developed to isolate and amplify RNA from single hyphae or parts thereof. Microarray analysis resulted in a present call for 4 to 7% of the A. niger genes, of which 12% showed heterogeneous RNA levels. These genes belonged to a wide range of gene categories. PMID:21816052

Changes in the transcriptomic profiles of maize roots in response to iron-deficiency stress.

PubMed

Li, Yan; Wang, Nian; Zhao, Fengtao; Song, Xuejiao; Yin, Zhaohua; Huang, Rong; Zhang, Chunqing

2014-07-01

Plants are often subjected to iron (Fe)-deficiency stress because of its low solubility. Plants have evolved two distinct strategies to solubilize and transport Fe to acclimate to this abiotic stress condition. Transcriptomic profiling analysis was performed using Illumina digital gene expression to understand the mechanism underlying resistance responses of roots to Fe starvation in maize, an important Strategy II plant. A total of 3,427, 4,069, 4,881, and 2,610 genes had significantly changed expression levels after Fe-deficiency treatments of 1, 2, 4 or 7 days, respectively. Genes involved in 2'-deoxymugineic acid (DMA) synthesis, secretion, and Fe(III)-DMA uptake were significantly induced. Many genes related to plant hormones, protein kinases, and protein phosphatases responded to Fe-deficiency stress, suggesting their regulatory roles in response to the Fe-deficiency stress. Functional annotation clustering analysis, using the Database for Annotation, Visualization and Integrated Discovery, revealed maize root responses to Fe starvation. This resulted in 38 functional annotation clusters: 25 for up-regulated genes, and 13 for down-regulated ones. These included genes encoding enzymes involved in the metabolism of carboxylic acids, isoprenoids and aromatic compounds, transporters, and stress response proteins. Our work provides integrated information for understanding maize response to Fe-deficiency stress.

Gene Expression Profiles in Rice Developing Ovules Provided Evidence for the Role of Sporophytic Tissue in Female Gametophyte Development.

PubMed

Wu, Ya; Yang, Liyu; Cao, Aqin; Wang, Jianbo

2015-01-01

The development of ovule in rice (Oryza sativa) is vital during its life cycle. To gain more understanding of the molecular events associated with the ovule development, we used RNA sequencing approach to perform transcriptome-profiling analysis of the leaf and ovules at four developmental stages. In total, 25,401, 23,343, 23,647 and 23,806 genes were identified from the four developmental stages of the ovule, respectively. We identified a number of differently expressed genes (DEGs) from three adjacent stage comparisons, which may play crucial roles in ovule development. The DEGs were then conducted functional annotations and Kyoto encyclopedia of genes and genomes (KEGG) pathway analyses. Genes related to cellular component biogenesis, membrane-bounded organelles and reproductive regulation were identified to be highly expressed during the ovule development. Different expression levels of auxin-related and cytokinin-related genes were also identified at various stages, providing evidence for the role of sporophytic ovule tissue in female gametophyte development from the aspect of gene expression. Generally, an overall transcriptome analysis for rice ovule development has been conducted. These results increased our knowledge of the complex molecular and cellular events that occur during the development of rice ovule and provided foundation for further studies on rice ovule development.

Transcriptome profiling and pathway analysis of hepatotoxicity induced by tris (2-ethylhexyl) trimellitate (TOTM) in mice.

PubMed

Chen, Xian-Hua; Ma, Li; Hu, Yi-Xiang; Wang, Dan-Xian; Fang, Li; Li, Xue-Lai; Zhao, Jin-Chuan; Yu, Hai-Rong; Ying, Hua-Zhong; Yu, Chen-Huan

2016-01-01

Tris (2-ethylhexyl) trimellitate (TOTM) is commonly used as an alternative plasticizer for medical devices. But very little information was available on its biological effects. In this study, we investigated toxicity effects of TOTM on hepatic differential gene expression analyzed by using high-throughput sequencing analysis for over-represented functions and phenotypically anchored to complementary histopathologic, and biochemical data in the liver of mice. Among 1668 candidate genes, 694 genes were up-regulated and 974 genes were down-regulated after TOTM exposure. Using Gene Ontology analysis, TOTM affected three processes: the cell cycle, metabolic process and oxidative activity. Furthermore, 11 key genes involved in the above processes were validated by real time PCR. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that these genes were involved in the cell cycle pathway, lipid metabolism and oxidative process. It revealed the transcriptome gene expression response to TOTM exposure in mouse, and these data could contribute to provide a clearer understanding of the molecular mechanisms of TOTM-induced hepatotoxicity in human. Copyright © 2015 Elsevier B.V. All rights reserved.

Genome-wide retinal transcriptome analysis of endotoxin-induced uveitis in mice with next-generation sequencing

PubMed Central

Qiu, Yiguo; Yu, Peng; Lin, Ru; Fu, Xinyu; Hao, Bingtao

2017-01-01

Purpose Endotoxin-induced uveitis (EIU) is a well-established mouse model for studying human acute inflammatory uveitis. The purpose of this study is to investigate the genome-wide retinal transcriptome profile of EIU. Methods The anterior segment of the mice was examined with a slit-lamp, and clinical scores were evaluated simultaneously. The histological changes in the posterior segment of the eyes were evaluated with hematoxylin and eosin (H&E) staining. A high throughput RNA sequencing (RNA-seq) strategy using the Illumina Hiseq 2500 platform was applied to characterize the retinal transcriptome profile from lipopolysaccharide (LPS)-treated and untreated mice. The validation of the differentially expressed genes (DEGs) was analyzed with real-time PCR. Results At the 24th hour after challenge, the clinical score of the LPS group was significantly higher (3.83±0.75, mean ± standard deviation [SD]) than that of the control group (0.08±0.20, mean ± SD; p<0.001). The histological evaluation showed a large number of inflammatory cells infiltrated into the vitreous cavity in the LPS group compared with the control group. A total of 478 DEGs were identified with RNA-seq. Among these genes, 406 were upregulated and 72 were downregulated in the LPS group. Gene Ontology (GO) enrichment showed three significantly enriched upregulated terms. Twenty-one upregulated and seven downregulated pathways were remarkably enriched by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. Eleven inflammatory response–, complement system–, fibrinolytic system–, and cell stress–related genes were validated to show similar results as the RNA-seq. Conclusions We first reported the retinal transcriptome profile of the EIU mouse with RNA-seq. The results indicate that the abnormal changes in the inflammatory response–, complement system–, fibrinolytic system–, and cell stress–related genes occurred concurrently in EIU. These genes may play an important role in the pathogenesis of EIU. This study will lead to a better understanding of the underlying mechanisms and shed light on discovering novel therapeutic targets for ocular inflammation. PMID:28706439

Genome-wide retinal transcriptome analysis of endotoxin-induced uveitis in mice with next-generation sequencing.

PubMed

Qiu, Yiguo; Yu, Peng; Lin, Ru; Fu, Xinyu; Hao, Bingtao; Lei, Bo

2017-01-01

Endotoxin-induced uveitis (EIU) is a well-established mouse model for studying human acute inflammatory uveitis. The purpose of this study is to investigate the genome-wide retinal transcriptome profile of EIU. The anterior segment of the mice was examined with a slit-lamp, and clinical scores were evaluated simultaneously. The histological changes in the posterior segment of the eyes were evaluated with hematoxylin and eosin (H&E) staining. A high throughput RNA sequencing (RNA-seq) strategy using the Illumina Hiseq 2500 platform was applied to characterize the retinal transcriptome profile from lipopolysaccharide (LPS)-treated and untreated mice. The validation of the differentially expressed genes (DEGs) was analyzed with real-time PCR. At the 24th hour after challenge, the clinical score of the LPS group was significantly higher (3.83±0.75, mean ± standard deviation [SD]) than that of the control group (0.08±0.20, mean ± SD; p<0.001). The histological evaluation showed a large number of inflammatory cells infiltrated into the vitreous cavity in the LPS group compared with the control group. A total of 478 DEGs were identified with RNA-seq. Among these genes, 406 were upregulated and 72 were downregulated in the LPS group. Gene Ontology (GO) enrichment showed three significantly enriched upregulated terms. Twenty-one upregulated and seven downregulated pathways were remarkably enriched by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. Eleven inflammatory response-, complement system-, fibrinolytic system-, and cell stress-related genes were validated to show similar results as the RNA-seq. We first reported the retinal transcriptome profile of the EIU mouse with RNA-seq. The results indicate that the abnormal changes in the inflammatory response-, complement system-, fibrinolytic system-, and cell stress-related genes occurred concurrently in EIU. These genes may play an important role in the pathogenesis of EIU. This study will lead to a better understanding of the underlying mechanisms and shed light on discovering novel therapeutic targets for ocular inflammation.

Salt-Responsive Transcriptome Profiling of Suaeda glauca via RNA Sequencing

PubMed Central

Jin, Hangxia; Dong, Dekun; Yang, Qinghua; Zhu, Danhua

2016-01-01

Background Suaeda glauca, a succulent halophyte of the Chenopodiaceae family, is widely distributed in coastal areas of China. Suaeda glauca is highly resistant to salt and alkali stresses. In the present study, the salt-responsive transcriptome of Suaeda glauca was analyzed to identify genes involved in salt tolerance and study halophilic mechanisms in this halophyte. Results Illumina HiSeq 2500 was used to sequence cDNA libraries from salt-treated and control samples with three replicates each treatment. De novo assembly of the six transcriptomes identified 75,445 unigenes. A total of 23,901 (31.68%) unigenes were annotated. Compared with transcriptomes from the three salt-treated and three salt-free samples, 231 differentially expressed genes (DEGs) were detected (including 130 up-regulated genes and 101 down-regulated genes), and 195 unigenes were functionally annotated. Based on the Gene Ontology (GO), Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) classifications of the DEGs, more attention should be paid to transcripts associated with signal transduction, transporters, the cell wall and growth, defense metabolism and transcription factors involved in salt tolerance. Conclusions This report provides a genome-wide transcriptional analysis of a halophyte, Suaeda glauca, under salt stress. Further studies of the genetic basis of salt tolerance in halophytes are warranted. PMID:26930632

EchinoDB, an application for comparative transcriptomics of deeply-sampled clades of echinoderms.

PubMed

Janies, Daniel A; Witter, Zach; Linchangco, Gregorio V; Foltz, David W; Miller, Allison K; Kerr, Alexander M; Jay, Jeremy; Reid, Robert W; Wray, Gregory A

2016-01-22

One of our goals for the echinoderm tree of life project (http://echinotol.org) is to identify orthologs suitable for phylogenetic analysis from next-generation transcriptome data. The current dataset is the largest assembled for echinoderm phylogeny and transcriptomics. We used RNA-Seq to profile adult tissues from 42 echinoderm specimens from 24 orders and 37 families. In order to achieve sampling members of clades that span key evolutionary divergence, many of our exemplars were collected from deep and polar seas. A small fraction of the transcriptome data we produced is being used for phylogenetic reconstruction. Thus to make a larger dataset available to researchers with a wide variety of interests, we made a web-based application, EchinoDB (http://echinodb.uncc.edu). EchinoDB is a repository of orthologous transcripts from echinoderms that is searchable via keywords and sequence similarity. From transcripts we identified 749,397 clusters of orthologous loci. We have developed the information technology to manage and search the loci their annotations with respect to the Sea Urchin (Strongylocentrotus purpuratus) genome. Several users have already taken advantage of these data for spin-off projects in developmental biology, gene family studies, and neuroscience. We hope others will search EchinoDB to discover datasets relevant to a variety of additional questions in comparative biology.

Maternal Pre-Pregnancy Obesity Is Associated with Altered Placental Transcriptome.

PubMed

Altmäe, Signe; Segura, Maria Teresa; Esteban, Francisco J; Bartel, Sabine; Brandi, Pilar; Irmler, Martin; Beckers, Johannes; Demmelmair, Hans; López-Sabater, Carmen; Koletzko, Berthold; Krauss-Etschmann, Susanne; Campoy, Cristina

2017-01-01

Maternal obesity has a major impact on pregnancy outcomes. There is growing evidence that maternal obesity has a negative influence on placental development and function, thereby adversely influencing offspring programming and health outcomes. However, the molecular mechanisms underlying these processes are poorly understood. We analysed ten term placenta's whole transcriptomes in obese (n = 5) and normal weight women (n = 5), using the Affymetrix microarray platform. Analyses of expression data were carried out using non-parametric methods. Hierarchical clustering and principal component analysis showed a clear distinction in placental transcriptome between obese and normal weight women. We identified 72 differentially regulated genes, with most being down-regulated in obesity (n = 61). Functional analyses of the targets using DAVID and IPA confirm the dysregulation of previously identified processes and pathways in the placenta from obese women, including inflammation and immune responses, lipid metabolism, cancer pathways, and angiogenesis. In addition, we detected new molecular aspects of obesity-derived effects on the placenta, involving the glucocorticoid receptor signalling pathway and dysregulation of several genes including CCL2, FSTL3, IGFBP1, MMP12, PRG2, PRL, QSOX1, SERPINE2 and TAC3. Our global gene expression profiling approach demonstrates that maternal obesity creates a unique in utero environment that impairs the placental transcriptome.

Transcriptome analysis of Aedes aegypti in response to mono-infections and co-infections of dengue virus-2 and chikungunya virus.

PubMed

Shrinet, Jatin; Srivastava, Pratibha; Sunil, Sujatha

2017-10-28

Chikungunya virus (CHIKV) and Dengue virus (DENV) spread via the bite of infected Aedes mosquitoes. Both these viruses exist as co-infections in the host as well as the vector and are known to exploit their cellular machinery for their replication. While there are studies reporting the changes in Aedes transcriptome when infected with DENV and CHIKV individually, the effect both these viruses have on the mosquitoes when present as co-infections is not clearly understood. In the present study, we infected Aedes aegypti mosquitoes with DENV and CHIKV individually and as co-infection through nanoinjections. We performed high throughput RNA sequencing of the infected Aedes aegypti to understand the changes in the Aedes transcriptome during the early stages of infection, i.e., 24 h post infection and compared the transcriptome profiles during DENV and CHIKV mono-infections with that of co-infections. We identified 190 significantly regulated genes identified in CHIKV infected library, 37 genes from DENV library and 100 genes from co-infected library and they were classified into different pathways. Our study reveal that distinct pathways and transcripts are being regulated during the three types of infection states in Aedes aegypti mosquitoes. Copyright © 2017 Elsevier Inc. All rights reserved.

Comprehensive Transcriptome Profiling and Functional Analysis of the Frog (Bombina maxima) Immune System

PubMed Central

Zhao, Feng; Yan, Chao; Wang, Xuan; Yang, Yang; Wang, Guangyin; Lee, Wenhui; Xiang, Yang; Zhang, Yun

2014-01-01

Amphibians occupy a key phylogenetic position in vertebrates and evolution of the immune system. But, the resources of its transcriptome or genome are still little now. Bombina maxima possess strong ability to survival in very harsh environment with a more mature immune system. We obtained a comprehensive transcriptome by RNA-sequencing technology. 14.3% of transcripts were identified to be skin-specific genes, most of which were not isolated from skin secretion in previous works or novel non-coding RNAs. 27.9% of transcripts were mapped into 242 predicted KEGG pathways and 6.16% of transcripts related to human disease and cancer. Of 39 448 transcripts with the coding sequence, at least 1501 transcripts (570 genes) related to the immune system process. The molecules of immune signalling pathway were almost presented, several transcripts with high expression in skin and stomach. Experiments showed that lipopolysaccharide or bacteria challenge stimulated pro-inflammatory cytokine production and activation of pro-inflammatory caspase-1. These frog's data can remarkably expand the existing genome or transcriptome resources of amphibians, especially immunity data. The entity of the data provides a valuable platform for further investigation on more detailed immune response in B. maxima and a comparative study with other amphibians. PMID:23942912

Beyond quantification: in situ analysis of transcriptome and pre-mRNA alternative splicing at the nanoscale.

PubMed

Cui, Yi; Liu, Jing; Irudayaraj, Joseph

2017-07-01

In situ analysis offers a venue for dissecting the complex transcriptome in its natural context to tap into cellular processes that could explain the phenotypic physiology and pathology yet to be understood. Over the past decades, enormous progress has been made to improve the resolution, sensitivity, and specificity of single-cell technologies. The continued efforts in RNA research not only facilitates mechanistic studies of molecular biology but also provides state-of-the-art strategies for diagnostic purposes. The implementation of novel bio-imaging platforms has yielded valuable information for inspecting gene expression, mapping regulatory networks, and classifying cell types. In this article, we discuss the merits and technical challenges in single-molecule in situ RNA profiling. Advanced in situ hybridization methodologies developed for a variety of detection modalities are reviewed. Considering the fact that in mammalian cells the number of protein products immensely exceeds that of the actual coding genes due to pre-mRNA alternative splicing, tools capable of elucidating this process in intact cells are highlighted. To conclude, we point out future directions for in situ transcriptome analysis and expect a plethora of opportunities and discoveries in this field. WIREs Nanomed Nanobiotechnol 2017, 9:e1443. doi: 10.1002/wnan.1443 For further resources related to this article, please visit the WIREs website. © 2016 Wiley Periodicals, Inc.

Liver Transcriptome Analysis of the Large Yellow Croaker (Larimichthys crocea) during Fasting by Using RNA-Seq

PubMed Central

Qian, Baoying; Xue, Liangyi; Huang, Hongli

2016-01-01

The large yellow croaker (Larimichthys crocea) is an economically important fish species in Chinese mariculture industry. To understand the molecular basis underlying the response to fasting, Illumina HiSeqTM 2000 was used to analyze the liver transcriptome of fasting large yellow croakers. A total of 54,933,550 clean reads were obtained and assembled into 110,364 contigs. Annotation to the NCBI database identified a total of 38,728 unigenes, of which 19,654 were classified into Gene Ontology and 22,683 were found in Kyoto Encyclopedia of Genes and Genomes (KEGG). Comparative analysis of the expression profiles between fasting fish and normal-feeding fish identified a total of 7,623 differentially expressed genes (P < 0.05), including 2,500 upregulated genes and 5,123 downregulated genes. Dramatic differences were observed in the genes involved in metabolic pathways such as fat digestion and absorption, citrate cycle, and glycolysis/gluconeogenesis, and the similar results were also found in the transcriptome of skeletal muscle. Further qPCR analysis confirmed that the genes encoding the factors involved in those pathways significantly changed in terms of expression levels. The results of the present study provide insights into the molecular mechanisms underlying the metabolic response of the large yellow croaker to fasting as well as identified areas that require further investigation. PMID:26967898

Existence of Inverted Profile in Chemically Responsive Molecular Pathways in the Zebrafish Liver

PubMed Central

Zhang, Xun; Li, Hu; Ma, Jing; Zhang, Louxin; Li, Baowen; Gong, Zhiyuan

2011-01-01

How a living organism maintains its healthy equilibrium in response to endless exposure of potentially harmful chemicals is an important question in current biology. By transcriptomic analysis of zebrafish livers treated by various chemicals, we defined hubs as molecular pathways that are frequently perturbed by chemicals and have high degree of functional connectivity to other pathways. Our network analysis revealed that these hubs were organized into two groups showing inverted functionality with each other. Intriguingly, the inverted activity profiles in these two groups of hubs were observed to associate only with toxicopathological states but not with physiological changes. Furthermore, these inverted profiles were also present in rat, mouse, and human under certain toxicopathological conditions. Thus, toxicopathological-associated anti-correlated profiles in hubs not only indicate their potential use in diagnosis but also development of systems-based therapeutics to modulate gene expression by chemical approach in order to rewire the deregulated activities of hubs back to normal physiology. PMID:22140468

Massively parallel digital transcriptional profiling of single cells

PubMed Central

Zheng, Grace X. Y.; Terry, Jessica M.; Belgrader, Phillip; Ryvkin, Paul; Bent, Zachary W.; Wilson, Ryan; Ziraldo, Solongo B.; Wheeler, Tobias D.; McDermott, Geoff P.; Zhu, Junjie; Gregory, Mark T.; Shuga, Joe; Montesclaros, Luz; Underwood, Jason G.; Masquelier, Donald A.; Nishimura, Stefanie Y.; Schnall-Levin, Michael; Wyatt, Paul W.; Hindson, Christopher M.; Bharadwaj, Rajiv; Wong, Alexander; Ness, Kevin D.; Beppu, Lan W.; Deeg, H. Joachim; McFarland, Christopher; Loeb, Keith R.; Valente, William J.; Ericson, Nolan G.; Stevens, Emily A.; Radich, Jerald P.; Mikkelsen, Tarjei S.; Hindson, Benjamin J.; Bielas, Jason H.

2017-01-01

Characterizing the transcriptome of individual cells is fundamental to understanding complex biological systems. We describe a droplet-based system that enables 3′ mRNA counting of tens of thousands of single cells per sample. Cell encapsulation, of up to 8 samples at a time, takes place in ∼6 min, with ∼50% cell capture efficiency. To demonstrate the system's technical performance, we collected transcriptome data from ∼250k single cells across 29 samples. We validated the sensitivity of the system and its ability to detect rare populations using cell lines and synthetic RNAs. We profiled 68k peripheral blood mononuclear cells to demonstrate the system's ability to characterize large immune populations. Finally, we used sequence variation in the transcriptome data to determine host and donor chimerism at single-cell resolution from bone marrow mononuclear cells isolated from transplant patients. PMID:28091601

A transcriptome-based assessment of the astrocytic dystrophin-associated complex in the developing human brain.

PubMed

Simon, Matthew J; Murchison, Charles; Iliff, Jeffrey J

2018-02-01

Astrocytes play a critical role in regulating the interface between the cerebral vasculature and the central nervous system. Contributing to this is the astrocytic endfoot domain, a specialized structure that ensheathes the entirety of the vasculature and mediates signaling between endothelial cells, pericytes, and neurons. The astrocytic endfoot has been implicated as a critical element of the glymphatic pathway, and changes in protein expression profiles in this cellular domain are linked to Alzheimer's disease pathology. Despite this, basic physiological properties of this structure remain poorly understood including the developmental timing of its formation, and the protein components that localize there to mediate its functions. Here we use human transcriptome data from male and female subjects across several developmental stages and brain regions to characterize the gene expression profile of the dystrophin-associated complex (DAC), a known structural component of the astrocytic endfoot that supports perivascular localization of the astroglial water channel aquaporin-4. Transcriptomic profiling is also used to define genes exhibiting parallel expression profiles to DAC elements, generating a pool of candidate genes that encode gene products that may contribute to the physiological function of the perivascular astrocytic endfoot domain. We found that several genes encoding transporter proteins are transcriptionally associated with DAC genes. © 2017 Wiley Periodicals, Inc.

Transcriptome profiling to discover putative genes associated with paraquat resistance in goosegrass (Eleusine indica L.).

PubMed

An, Jing; Shen, Xuefeng; Ma, Qibin; Yang, Cunyi; Liu, Simin; Chen, Yong

2014-01-01

Goosegrass (Eleusine indica L.), a serious annual weed in the world, has evolved resistance to several herbicides including paraquat, a non-selective herbicide. The mechanism of paraquat resistance in weeds is only partially understood. To further study the molecular mechanism underlying paraquat resistance in goosegrass, we performed transcriptome analysis of susceptible and resistant biotypes of goosegrass with or without paraquat treatment. The RNA-seq libraries generated 194,716,560 valid reads with an average length of 91.29 bp. De novo assembly analysis produced 158,461 transcripts with an average length of 1153.74 bp and 100,742 unigenes with an average length of 712.79 bp. Among these, 25,926 unigenes were assigned to 65 GO terms that contained three main categories. A total of 13,809 unigenes with 1,208 enzyme commission numbers were assigned to 314 predicted KEGG metabolic pathways, and 12,719 unigenes were categorized into 25 KOG classifications. Furthermore, our results revealed that 53 genes related to reactive oxygen species scavenging, 10 genes related to polyamines and 18 genes related to transport were differentially expressed in paraquat treatment experiments. The genes related to polyamines and transport are likely potential candidate genes that could be further investigated to confirm their roles in paraquat resistance of goosegrass. This is the first large-scale transcriptome sequencing of E. indica using the Illumina platform. Potential genes involved in paraquat resistance were identified from the assembled sequences. The transcriptome data may serve as a reference for further analysis of gene expression and functional genomics studies, and will facilitate the study of paraquat resistance at the molecular level in goosegrass.

Transcriptome Profiling to Discover Putative Genes Associated with Paraquat Resistance in Goosegrass (Eleusine indica L.)

PubMed Central

An, Jing; Shen, Xuefeng; Ma, Qibin; Yang, Cunyi; Liu, Simin; Chen, Yong

2014-01-01

Background Goosegrass (Eleusine indica L.), a serious annual weed in the world, has evolved resistance to several herbicides including paraquat, a non-selective herbicide. The mechanism of paraquat resistance in weeds is only partially understood. To further study the molecular mechanism underlying paraquat resistance in goosegrass, we performed transcriptome analysis of susceptible and resistant biotypes of goosegrass with or without paraquat treatment. Results The RNA-seq libraries generated 194,716,560 valid reads with an average length of 91.29 bp. De novo assembly analysis produced 158,461 transcripts with an average length of 1153.74 bp and 100,742 unigenes with an average length of 712.79 bp. Among these, 25,926 unigenes were assigned to 65 GO terms that contained three main categories. A total of 13,809 unigenes with 1,208 enzyme commission numbers were assigned to 314 predicted KEGG metabolic pathways, and 12,719 unigenes were categorized into 25 KOG classifications. Furthermore, our results revealed that 53 genes related to reactive oxygen species scavenging, 10 genes related to polyamines and 18 genes related to transport were differentially expressed in paraquat treatment experiments. The genes related to polyamines and transport are likely potential candidate genes that could be further investigated to confirm their roles in paraquat resistance of goosegrass. Conclusion This is the first large-scale transcriptome sequencing of E. indica using the Illumina platform. Potential genes involved in paraquat resistance were identified from the assembled sequences. The transcriptome data may serve as a reference for further analysis of gene expression and functional genomics studies, and will facilitate the study of paraquat resistance at the molecular level in goosegrass. PMID:24927422

Coupling Deep Transcriptome Analysis with Untargeted Metabolic Profiling in Ophiorrhiza pumila to Further the Understanding of the Biosynthesis of the Anti-Cancer Alkaloid Camptothecin and Anthraquinones

PubMed Central

Yamazaki, Mami; Mochida, Keiichi; Asano, Takashi; Nakabayashi, Ryo; Chiba, Motoaki; Udomson, Nirin; Yamazaki, Yasuyo; Goodenowe, Dayan B.; Sankawa, Ushio; Yoshida, Takuhiro; Toyoda, Atsushi; Totoki, Yasushi; Sakaki, Yoshiyuki; Góngora-Castillo, Elsa; Buell, C. Robin; Sakurai, Tetsuya; Saito, Kazuki

2013-01-01

The Rubiaceae species, Ophiorrhiza pumila, accumulates camptothecin, an anti-cancer alkaloid with a potent DNA topoisomerase I inhibitory activity, as well as anthraquinones that are derived from the combination of the isochorismate and hemiterpenoid pathways. The biosynthesis of these secondary products is active in O. pumila hairy roots yet very low in cell suspension culture. Deep transcriptome analysis was conducted in O. pumila hairy roots and cell suspension cultures using the Illumina platform, yielding a total of 2 Gb of sequence for each sample. We generated a hybrid transcriptome assembly of O. pumila using the Illumina-derived short read sequences and conventional Sanger-derived expressed sequence tag clones derived from a full-length cDNA library constructed using RNA from hairy roots. Among 35,608 non-redundant unigenes, 3,649 were preferentially expressed in hairy roots compared with cell suspension culture. Candidate genes involved in the biosynthetic pathway for the monoterpenoid indole alkaloid camptothecin were identified; specifically, genes involved in post-strictosamide biosynthetic events and genes involved in the biosynthesis of anthraquinones and chlorogenic acid. Untargeted metabolomic analysis by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) indicated that most of the proposed intermediates in the camptothecin biosynthetic pathway accumulated in hairy roots in a preferential manner compared with cell suspension culture. In addition, a number of anthraquinones and chlorogenic acid preferentially accumulated in hairy roots compared with cell suspension culture. These results suggest that deep transcriptome and metabolome data sets can facilitate the identification of genes and intermediates involved in the biosynthesis of secondary products including camptothecin in O. pumila. PMID:23503598

Transcriptome analysis of starch and sucrose metabolism across bulb development in Sagittaria sagittifolia.

PubMed

Gao, Meiping; Zhang, Shangwen; Luo, Cong; He, Xinhua; Wei, Shaolong; Jiang, Wen; He, Fanglian; Lin, Zhicheng; Yan, Meixin; Dong, Weiqong

2018-04-05

Sagittaria sagittifolia L is an important bulb vegetable that has high nutritional and medical value. Bulb formation and development are crucial to Sagittaria sagittifolia; however, its sucrose metabolism is poorly understood and there are a lack of sufficient transcriptomic and genomic data available to fully understand the molecular mechanisms underlying bulb formation and development as well as the bulb transcriptome. Five cDNA libraries were constructed at different developmental stages and sequenced using high-throughput Illumina RNA sequencing. From approximately 63.53 Gb clean reads, a total of 60,884 unigenes, with an average length of 897.34 bp and N50 of 1.368 kb, were obtained. A total of 36,590 unigenes were successfully annotated using five public databases. Across different developmental stages, 4195, 827, 832, 851, and 1494 were differentially expressed in T02, T03, T04, T05, and T06 libraries, respectively. Gene ontology (GO) analysis revealed several differentially-expressed genes (DEGs) associated with catalytic activity, binding, and transporter activity. The Kyoto encyclopedia of genes and genomes (KEGG) revealed that these DEGs are involved in physiological and biochemical processes. RT-qPCR was used to profile the expression of these unigenes and revealed that the expression patterns of the DEGs were consistent with the transcriptome data. In this study, we conducted a comparative gene expression analysis at the transcriptional level using RNA-seq across the different developmental stages of Sagittaria sagittifolia. We identified a set of genes that might contribute to starch and sucrose metabolism, and the genetic mechanisms related to bulblet development were also explored. This study provides important data for future studies of the genetic and molecular mechanisms underlying bulb formation and development in Sagittaria sagittifolia. Copyright © 2018. Published by Elsevier B.V.

Transcriptional profiles of Arabidopsis stomataless mutants reveal developmental and physiological features of life in the absence of stomata

PubMed Central

de Marcos, Alberto; Triviño, Magdalena; Pérez-Bueno, María Luisa; Ballesteros, Isabel; Barón, Matilde; Mena, Montaña; Fenoll, Carmen

2015-01-01

Loss of function of the positive stomata development regulators SPCH or MUTE in Arabidopsis thaliana renders stomataless plants; spch-3 and mute-3 mutants are extreme dwarfs, but produce cotyledons and tiny leaves, providing a system to interrogate plant life in the absence of stomata. To this end, we compared their cotyledon transcriptomes with that of wild-type plants. K-means clustering of differentially expressed genes generated four clusters: clusters 1 and 2 grouped genes commonly regulated in the mutants, while clusters 3 and 4 contained genes distinctively regulated in mute-3. Classification in functional categories and metabolic pathways of genes in clusters 1 and 2 suggested that both mutants had depressed secondary, nitrogen and sulfur metabolisms, while only a few photosynthesis-related genes were down-regulated. In situ quenching analysis of chlorophyll fluorescence revealed limited inhibition of photosynthesis. This and other fluorescence measurements matched the mutant transcriptomic features. Differential transcriptomes of both mutants were enriched in growth-related genes, including known stomata development regulators, which paralleled their epidermal phenotypes. Analysis of cluster 3 was not informative for developmental aspects of mute-3. Cluster 4 comprised genes differentially up−regulated in mute−3, 35% of which were direct targets for SPCH and may relate to the unique cell types of mute−3. A screen of T-DNA insertion lines in genes differentially expressed in the mutants identified a gene putatively involved in stomata development. A collection of lines for conditional overexpression of transcription factors differentially expressed in the mutants rendered distinct epidermal phenotypes, suggesting that these proteins may be novel stomatal development regulators. Thus, our transcriptome analysis represents a useful source of new genes for the study of stomata development and for characterizing physiology and growth in the absence of stomata. PMID:26157447

20180312 - Application of a Multiplexed High Content Imaging (HCI) Based Cell Viability and Apoptosis Chemical Screening Assay with Results in MCF-7 Cells (SOT)

EPA Science Inventory

The NCCT high throughput transcriptomics (HTTr) screening program uses whole transcriptome profiling assay in human-derived cells to collect concentration-response data for large numbers (100s-1000s) of environmental chemicals. To contextualize HTTr data, chemical effects on cell...

Analysis of hepatic transcript profile and plasma lipid profile in early lactating dairy cows fed grape seed and grape marc meal extract.

PubMed

Gessner, Denise K; Winkler, Anne; Koch, Christian; Dusel, Georg; Liebisch, Gerhard; Ringseis, Robert; Eder, Klaus

2017-03-23

It was recently reported that dairy cows fed a polyphenol-rich grape seed and grape marc meal extract (GSGME) during the transition period had an increased milk yield, but the underlying reasons remained unclear. As polyphenols exert a broad spectrum of metabolic effects, we hypothesized that feeding of GSGME influences metabolic pathways in the liver which could account for the positive effects of GSGME in dairy cows. In order to identify these pathways, we performed genome-wide transcript profiling in the liver and lipid profiling in plasma of dairy cows fed GSGME during the transition period at 1 week postpartum. Transcriptomic analysis of the liver revealed 207 differentially expressed transcripts, from which 156 were up- and 51 were down-regulated, between cows fed GSGME and control cows. Gene set enrichment analysis of the 155 up-regulated mRNAs showed that the most enriched gene ontology (GO) biological process terms were dealing with cell cycle regulation and the most enriched Kyoto Encyclopedia of Genes and Genomes pathways were p53 signaling and cell cycle. Functional analysis of the 43 down-regulated mRNAs revealed that a great part of these genes are involved in endoplasmic reticulum (ER) stress-induced unfolded protein response (UPR) and inflammatory processes. Accordingly, protein folding, response to unfolded protein, unfolded protein binding, chemokine activity and heat shock protein binding were identified as one of the most enriched GO biological process and molecular function terms assigned to the down-regulated genes. In line with the transcriptomics data the plasma concentrations of the acute phase proteins serum amyloid A (SAA) and haptoglobin were reduced in cows fed GSGME compared to control cows. Lipidomic analysis of plasma revealed no differences in the concentrations of individual species of major and minor lipid classes between cows fed GSGME and control cows. Analysis of hepatic transcript profile in cows fed GSGME during the transition period at 1 week postpartum indicates that polyphenol-rich feed components are able to inhibit ER stress-induced UPR and inflammatory processes, both of which are considered to contribute to liver-associated diseases and to impair milk performance in dairy cows, in the liver of dairy cows during early lactation.

Transcriptome profiling of Diachasmimorpha longicaudata towards useful molecular tools for population management.

PubMed

Mannino, M Constanza; Rivarola, Máximo; Scannapieco, Alejandra C; González, Sergio; Farber, Marisa; Cladera, Jorge L; Lanzavecchia, Silvia B

2016-10-12

Diachasmimorpha longicaudata (Hymenoptera: Braconidae) is a solitary parasitoid of Tephritidae (Diptera) fruit flies of economic importance currently being mass-reared in bio-factories and successfully used worldwide. A peculiar biological aspect of Hymenoptera is its haplo-diploid life cycle, where females (diploid) develop from fertilized eggs and males (haploid) from unfertilized eggs. Diploid males were described in many species and recently evidenced in D. longicaudata by mean of inbreeding studies. Sex determination in this parasitoid is based on the Complementary Sex Determination (CSD) system, with alleles from at least one locus involved in early steps of this pathway. Since limited information is available about genetics of this parasitoid species, a deeper analysis on D. longicaudata's genomics is required to provide molecular tools for achieving a more cost effective production under artificial rearing conditions. We report here the first transcriptome analysis of male-larvae, adult females and adult males of D. longicaudata using 454-pyrosequencing. A total of 469766 reads were analyzed and 8483 high-quality isotigs were assembled. After functional annotation, a total of 51686 unigenes were produced, from which, 7021 isotigs and 20227 singletons had at least one BLAST hit against the NCBI non-redundant protein database. A preliminary comparison of adult female and male evidenced that 98 transcripts showed differential expression profiles, with at least a 10-fold difference. Among the functionally annotated transcripts we detected four sequences potentially involved in sex determination and three homologues to two known genes involved in the sex determination cascade. Finally, a total of 4674SimpleSequence Repeats (SSRs) were in silico identified and characterized. The information obtained here will significantly contribute to the development of D. longicaudata functional genomics, genetics and population-based genome studies. Thousands of new microsatellite markers were identified as toolkits for population genetics analysis. The transcriptome characterized here is the starting point to elucidate the molecular bases of the sex determination mechanism in this species.

Extensive Transcriptomic and Genomic Analysis Provides New Insights about Luminal Breast Cancers

PubMed Central

Tishchenko, Inna; Milioli, Heloisa Helena; Riveros, Carlos; Moscato, Pablo

2016-01-01

Despite constituting approximately two thirds of all breast cancers, the luminal A and B tumours are poorly classified at both clinical and molecular levels. There are contradictory reports on the nature of these subtypes: some define them as intrinsic entities, others as a continuum. With the aim of addressing these uncertainties and identifying molecular signatures of patients at risk, we conducted a comprehensive transcriptomic and genomic analysis of 2,425 luminal breast cancer samples. Our results indicate that the separation between the molecular luminal A and B subtypes—per definition—is not associated with intrinsic characteristics evident in the differentiation between other subtypes. Moreover, t-SNE and MST-kNN clustering approaches based on 10,000 probes, associated with luminal tumour initiation and/or development, revealed the close connections between luminal A and B tumours, with no evidence of a clear boundary between them. Thus, we considered all luminal tumours as a single heterogeneous group for analysis purposes. We first stratified luminal tumours into two distinct groups by their HER2 gene cluster co-expression: HER2-amplified luminal and ordinary-luminal. The former group is associated with distinct transcriptomic and genomic profiles, and poor prognosis; it comprises approximately 8% of all luminal cases. For the remaining ordinary-luminal tumours we further identified the molecular signature correlated with disease outcomes, exhibiting an approximately continuous gene expression range from low to high risk. Thus, we employed four virtual quantiles to segregate the groups of patients. The clinico-pathological characteristics and ratios of genomic aberrations are concordant with the variations in gene expression profiles, hinting at a progressive staging. The comparison with the current separation into luminal A and B subtypes revealed a substantially improved survival stratification. Concluding, we suggest a review of the definition of luminal A and B subtypes. A proposition for a revisited delineation is provided in this study. PMID:27341628

Genome-Wide Identification of Differentially Expressed Genes Associated with the High Yielding of Oleoresin in Secondary Xylem of Masson Pine (Pinus massoniana Lamb) by Transcriptomic Analysis

PubMed Central

Liu, Qinghua; Zhou, Zhichun; Wei, Yongcheng; Shen, Danyu; Feng, Zhongping; Hong, Shanping

2015-01-01

Masson pine is an important timber and resource for oleoresin in South China. Increasing yield of oleoresin in stems can raise economic benefits and enhance the resistance to bark beetles. However, the genetic mechanisms for regulating the yield of oleoresin were still unknown. Here, high-throughput sequencing technology was used to investigate the transcriptome and compare the gene expression profiles of high and low oleoresin-yielding genotypes. A total of 40,690,540 reads were obtained and assembled into 137,499 transcripts from the secondary xylem tissues. We identified 84,842 candidate unigenes based on sequence annotation using various databases and 96 unigenes were candidates for terpenoid backbone biosynthesis in pine. By comparing the expression profiles of high and low oleoresin-yielding genotypes, 649 differentially expressed genes (DEGs) were identified. GO enrichment analysis of DEGs revealed that multiple pathways were related to high yield of oleoresin. Nine candidate genes were validated by QPCR analysis. Among them, the candidate genes encoding geranylgeranyl diphosphate synthase (GGPS) and (-)-alpha/beta-pinene synthase were up-regulated in the high oleoresin-yielding genotype, while tricyclene synthase revealed lower expression level, which was in good agreement with the GC/MS result. In addition, DEG encoding ABC transporters, pathogenesis-related proteins (PR5 and PR9), phosphomethylpyrimidine synthase, non-specific lipid-transfer protein-like protein and ethylene responsive transcription factors (ERFs) were also confirmed to be critical for the biosynthesis of oleoresin. The next-generation sequencing strategy used in this study has proven to be a powerful means for analyzing transcriptome variation related to the yield of oleoresin in masson pine. The candidate genes encoding GGPS, (-)-alpha/beta-pinene, tricyclene synthase, ABC transporters, non-specific lipid-transfer protein-like protein, phosphomethylpyrimidine synthase, ERFs and pathogen responses may play important roles in regulating the yield of oleoresin. These DEGs are worthy of special attention in future studies. PMID:26167875

Transcriptome-based investigation of cirrus development and identifying microsatellite markers in rattan (Daemonorops jenkinsiana)

PubMed Central

Zhao, Hansheng; Sun, Huayu; Li, Lichao; Lou, Yongfeng; Li, Rongsheng; Qi, Lianghua; Gao, Zhimin

2017-01-01

Rattan is an important group of regenerating non-wood climbing palm in tropical forests. The cirrus is an essential climbing organ and provides morphological evidence for evolutionary and taxonomic studies. However, limited data are available on the molecular mechanisms underlying the development of the cirrus. Thus, we performed in-depth transcriptomic sequencing analyses to characterize the cirrus development at different developmental stages of Daemonorops jenkinsiana. The result showed 404,875 transcripts were assembled, including 61,569 high-quality unigenes were identified, of which approximately 76.16% were annotated and classified by seven authorized databases. Moreover, a comprehensive analysis of the gene expression profiles identified differentially expressed genes (DEGs) concentrated in developmental pathways, cell wall metabolism, and hook formation between the different stages of the cirri. Among them, 37 DEGs were validated by qRT-PCR. Furthermore, 14,693 transcriptome-based microsatellites were identified. Of the 168 designed SSR primer pairs, 153 were validated and 16 pairs were utilized for the polymorphic analysis of 25 rattan accessions. These findings can be used to interpret the molecular mechanisms of cirrus development, and the developed microsatellites markers provide valuable data for assisting rattan taxonomy and expanding the understanding of genomic study in rattan. PMID:28383053

Comparative transcriptome assembly and genome-guided profiling for Brettanomyces bruxellensis LAMAP2480 during p-coumaric acid stress.

PubMed

Godoy, Liliana; Vera-Wolf, Patricia; Martinez, Claudio; Ugalde, Juan A; Ganga, María Angélica

2016-09-28

Brettanomyces bruxellensis has been described as the main contaminant yeast in wine production, due to its ability to convert the hydroxycinnamic acids naturally present in the grape phenolic derivatives, into volatile phenols. Currently, there are no studies in B. bruxellensis which explains the resistance mechanisms to hydroxycinnamic acids, and in particular to p-coumaric acid which is directly involved in alterations to wine. In this work, we performed a transcriptome analysis of B. bruxellensis LAMAP248rown in the presence and absence of p-coumaric acid during lag phase. Because of reported genetic variability among B. bruxellensis strains, to complement de novo assembly of the transcripts, we used the high-quality genome of B. bruxellensis AWRI1499, as well as the draft genomes of strains CBS2499 and0 g LAMAP2480. The results from the transcriptome analysis allowed us to propose a model in which the entrance of p-coumaric acid to the cell generates a generalized stress condition, in which the expression of proton pump and efflux of toxic compounds are induced. In addition, these mechanisms could be involved in the outflux of nitrogen compounds, such as amino acids, decreasing the overall concentration and triggering the expression of nitrogen metabolism genes.

Comparative transcriptome assembly and genome-guided profiling for Brettanomyces bruxellensis LAMAP2480 during p-coumaric acid stress

PubMed Central

Godoy, Liliana; Vera-Wolf, Patricia; Martinez, Claudio; Ugalde, Juan A.; Ganga, María Angélica

2016-01-01

Brettanomyces bruxellensis has been described as the main contaminant yeast in wine production, due to its ability to convert the hydroxycinnamic acids naturally present in the grape phenolic derivatives, into volatile phenols. Currently, there are no studies in B. bruxellensis which explains the resistance mechanisms to hydroxycinnamic acids, and in particular to p-coumaric acid which is directly involved in alterations to wine. In this work, we performed a transcriptome analysis of B. bruxellensis LAMAP248rown in the presence and absence of p-coumaric acid during lag phase. Because of reported genetic variability among B. bruxellensis strains, to complement de novo assembly of the transcripts, we used the high-quality genome of B. bruxellensis AWRI1499, as well as the draft genomes of strains CBS2499 and0 g LAMAP2480. The results from the transcriptome analysis allowed us to propose a model in which the entrance of p-coumaric acid to the cell generates a generalized stress condition, in which the expression of proton pump and efflux of toxic compounds are induced. In addition, these mechanisms could be involved in the outflux of nitrogen compounds, such as amino acids, decreasing the overall concentration and triggering the expression of nitrogen metabolism genes. PMID:27678167

Garlic (Allium sativum L.) fertility: transcriptome and proteome analyses provide insight into flower and pollen development

PubMed Central

Shemesh-Mayer, Einat; Ben-Michael, Tomer; Rotem, Neta; Rabinowitch, Haim D.; Doron-Faigenboim, Adi; Kosmala, Arkadiusz; Perlikowski, Dawid; Sherman, Amir; Kamenetsky, Rina

2015-01-01

Commercial cultivars of garlic, a popular condiment, are sterile, making genetic studies and breeding of this plant challenging. However, recent fertility restoration has enabled advanced physiological and genetic research and hybridization in this important crop. Morphophysiological studies, combined with transcriptome and proteome analyses and quantitative PCR validation, enabled the identification of genes and specific processes involved in gametogenesis in fertile and male-sterile garlic genotypes. Both genotypes exhibit normal meiosis at early stages of anther development, but in the male-sterile plants, tapetal hypertrophy after microspore release leads to pollen degeneration. Transcriptome analysis and global gene-expression profiling showed that >16,000 genes are differentially expressed in the fertile vs. male-sterile developing flowers. Proteome analysis and quantitative comparison of 2D-gel protein maps revealed 36 significantly different protein spots, 9 of which were present only in the male-sterile genotype. Bioinformatic and quantitative PCR validation of 10 candidate genes exhibited significant expression differences between male-sterile and fertile flowers. A comparison of morphophysiological and molecular traits of fertile and male-sterile garlic flowers suggests that respiratory restrictions and/or non-regulated programmed cell death of the tapetum can lead to energy deficiency and consequent pollen abortion. Potential molecular markers for male fertility and sterility in garlic are proposed. PMID:25972879

De novo analysis of the Nilaparvata lugens (Stål) antenna transcriptome and expression patterns of olfactory genes.

PubMed

Zhou, Shuang-Shuang; Sun, Ze; Ma, Weihua; Chen, Wei; Wang, Man-Qun

2014-03-01

We sequenced the antenna transcriptome of the brown planthopper (BPH), Nilaparvata lugens (Stål), a global rice pest, and performed transcriptome analysis on BPH antenna. We obtained about 40million 90bp reads that were assembled into 75,874 unigenes with a mean size of 456bp. Among the antenna transcripts, 32,856 (43%) showed significant similarity (E-value <1e(-5)) to known proteins in the NCBI database. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were used to classify functions of BPH antenna genes. We identified 10 odorant-binding proteins (OBPs), including 7 previously unidentified, and 11 chemosensory proteins (CSPs), including two new members. The expression profiles of 4 OBPs and 2 CSPs were determined by q-PCR for antenna, abdomen, leg and wing of insects of different age, gender, and mating status including two BPH adult wing-morphology types. NlugCSP10 and 4 OBPs appeared to be antenna-specific because they were highly and differentially expressed in male and female antennae. NlugCSP11 was expressed ubiquitously, with particularly high expression in wings. The transcript levels of several olfactory genes depended on adult wing form, age, gender, and mating status, although no clear expression patterns were determined. Copyright © 2013 Elsevier Inc. All rights reserved.

Characterization of the global transcriptome for cotton (Gossypium hirsutum L.) anther and development of SSR marker.

PubMed

Zhang, Xianwen; Ye, Zhenwei; Wang, Tiankang; Xiong, Hairong; Yuan, Xiaoling; Zhang, Zhigang; Yuan, Youlu; Liu, Zhi

2014-11-10

Cotton is an important fiber plant, and it's attractive to elucidate the molecular mechanism of anther development due to the close relationship between the anther fertility and boll-setting, and also fiber yield. In the present paper, 47.2 million paired-end reads with average length of 82.87 bp from the anthers of TM-1 (Gossypium hirsutum L.), a genetic standard line, were generated through transcriptome sequencing, and 210,965 unigenes of more than 100 bp were obtained. BLAST, KEGG, COG, and GO analyses showed that the genes were enriched in the processes of transcription, translation, and post-translation as well as hormone signal transduction, the transcription factor families, and cell wall-related genes mainly participating in cell expansion and carbohydrate metabolism. Further analysis identified 11,153 potential SSRs. A suit of 5122 primer pair sequences were designed, and 82 of 300 randomly selected primer pairs produced reproducible amplicons that were polymorphic among 22 cotton accessions from G. hirsutum, Gossypium barbadense and Gossypium arboreum. The UPGMA clustering analysis further confirmed high quality and effectiveness of these novel SSR markers. The present study provided insights into the transcriptome profile of the cotton and established a public information platform for functional genomics and molecular breeding. Copyright © 2014 Elsevier B.V. All rights reserved.

RNA-Seq analysis of global transcriptomic changes suggests a roles for the MAPK pathway and carbon metabolism in cell wall maintenance in a Saccharomyces cerevisiae FKS1 mutant.

PubMed

Huang, Cong; Zhao, Fengguang; Lin, Ying; Zheng, Suiping; Liang, Shuli; Han, Shuangyan

2018-06-07

FKS1 encodes a β-1,3-glucan synthase, which is a key player in cell wall assembly in Saccharomyces cerevisiae. Here we analyzed the global transcriptomic changes in the FKS1 mutant to establish a correlation between the changes in the cell wall of the FKS1 mutant and the molecular mechanism of cell wall maintenance. These transcriptomic profiles showed that there are 1151 differentially expressed genes (DEGs) in the FKS1 mutant. Through KEGG pathway analysis of the DEGs, the MAPK pathway and seven pathways involved in carbon metabolism were significantly enriched. We found that the MAPK pathway is activated for FKS1 mutant survival and the synthesis of cell wall components are reinforced in the FKS1 mutant. Our results confirm that the FKS1 mutant has a β-1,3-glucan defect that affects the cell wall and partly elucidate the molecular mechanism responsible for cell wall synthesis. Our greater understanding of these mechanisms helps to explain how the FKS1 mutant survives, has useful implications for the study of similar pathways in other fungi, and increases the theoretical foundation for the regulation of the cell wall in S. cerevisiae. Copyright © 2018 Elsevier Inc. All rights reserved.

Transcriptome analysis on the exoskeleton formation in early developmetal stages and reconstruction scenario in growth-moulting in Litopenaeus vannamei.

PubMed

Gao, Yi; Wei, Jiankai; Yuan, Jianbo; Zhang, Xiaojun; Li, Fuhua; Xiang, Jianhai

2017-04-24

Exoskeleton construction is an important issue in shrimp. To better understand the molecular mechanism of exoskeleton formation, development and reconstruction, the transcriptome of the entire developmental process in Litopenaeus vannamei, including nine early developmental stages and eight adult-moulting stages, was sequenced and analysed using Illumina RNA-seq technology. A total of 117,539 unigenes were obtained, with 41.2% unigenes predicting the full-length coding sequence. Gene Ontology, Clusters of Orthologous Group (COG), the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and functional annotation of all unigenes gave a better understanding of the exoskeleton developmental process in L. vannamei. As a result, more than six hundred unigenes related to exoskeleton development were identified both in the early developmental stages and adult-moulting. A cascade of sequential expression events of exoskeleton-related genes were summarized, including exoskeleton formation, regulation, synthesis, degradation, mineral absorption/reabsorption, calcification and hardening. This new insight on major transcriptional events provide a deep understanding for exoskeleton formation and reconstruction in L. vannamei. In conclusion, this is the first study that characterized the integrated transcriptomic profiles cover the entire exoskeleton development from zygote to adult-moulting in a crustacean, and these findings will serve as significant references for exoskeleton developmental biology and aquaculture research.

Transcriptomic and proteomic responses of Serratia marcescens to spaceflight conditions involve large-scale changes in metabolic pathways

NASA Astrophysics Data System (ADS)

Wang, Yajuan; Yuan, Yanting; Liu, Jinwen; Su, Longxiang; Chang, De; Guo, Yinghua; Chen, Zhenhong; Fang, Xiangqun; Wang, Junfeng; Li, Tianzhi; Zhou, Lisha; Fang, Chengxiang; Yang, Ruifu; Liu, Changting

2014-04-01

The microgravity environment of spaceflight expeditions has been associated with altered microbial responses. This study explores the characterization of Serratia marcescensis grown in a spaceflight environment at the phenotypic, transcriptomic and proteomic levels. From November 1, 2011 to November 17, 2011, a strain of S. marcescensis was sent into space for 398 h on the Shenzhou VIII spacecraft, and ground simulation was performed as a control (LCT-SM213). After the flight, two mutant strains (LCT-SM166 and LCT-SM262) were selected for further analysis. Although no changes in the morphology, post-culture growth kinetics, hemolysis or antibiotic sensitivity were observed, the two mutant strains exhibited significant changes in their metabolic profiles after exposure to spaceflight. Enrichment analysis of the transcriptome showed that the differentially expressed genes of the two spaceflight strains and the ground control strain mainly included those involved in metabolism and degradation. The proteome revealed that changes at the protein level were also associated with metabolic functions, such as glycolysis/gluconeogenesis, pyruvate metabolism, arginine and proline metabolism and the degradation of valine, leucine and isoleucine. In summary S. marcescens showed alterations primarily in genes and proteins that were associated with metabolism under spaceflight conditions, which gave us valuable clues for future research.

The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups.

PubMed

Curtis, Christina; Shah, Sohrab P; Chin, Suet-Feung; Turashvili, Gulisa; Rueda, Oscar M; Dunning, Mark J; Speed, Doug; Lynch, Andy G; Samarajiwa, Shamith; Yuan, Yinyin; Gräf, Stefan; Ha, Gavin; Haffari, Gholamreza; Bashashati, Ali; Russell, Roslin; McKinney, Steven; Langerød, Anita; Green, Andrew; Provenzano, Elena; Wishart, Gordon; Pinder, Sarah; Watson, Peter; Markowetz, Florian; Murphy, Leigh; Ellis, Ian; Purushotham, Arnie; Børresen-Dale, Anne-Lise; Brenton, James D; Tavaré, Simon; Caldas, Carlos; Aparicio, Samuel

2012-04-18

The elucidation of breast cancer subgroups and their molecular drivers requires integrated views of the genome and transcriptome from representative numbers of patients. We present an integrated analysis of copy number and gene expression in a discovery and validation set of 997 and 995 primary breast tumours, respectively, with long-term clinical follow-up. Inherited variants (copy number variants and single nucleotide polymorphisms) and acquired somatic copy number aberrations (CNAs) were associated with expression in ~40% of genes, with the landscape dominated by cis- and trans-acting CNAs. By delineating expression outlier genes driven in cis by CNAs, we identified putative cancer genes, including deletions in PPP2R2A, MTAP and MAP2K4. Unsupervised analysis of paired DNA–RNA profiles revealed novel subgroups with distinct clinical outcomes, which reproduced in the validation cohort. These include a high-risk, oestrogen-receptor-positive 11q13/14 cis-acting subgroup and a favourable prognosis subgroup devoid of CNAs. Trans-acting aberration hotspots were found to modulate subgroup-specific gene networks, including a TCR deletion-mediated adaptive immune response in the ‘CNA-devoid’ subgroup and a basal-specific chromosome 5 deletion-associated mitotic network. Our results provide a novel molecular stratification of the breast cancer population, derived from the impact of somatic CNAs on the transcriptome.

Profiles of metabolites and gene expression in rats with chemically induced hepatic necrosis.

PubMed

Heijne, Wilbert H M; Lamers, Robert-Jan A N; van Bladeren, Peter J; Groten, John P; van Nesselrooij, Joop H J; van Ommen, Ben

2005-01-01

This study investigated whether integrated analysis of transcriptomics and metabolomics data increased the sensitivity of detection and provided new insight in the mechanisms of hepatotoxicity. Metabolite levels in plasma or urine were analyzed in relation to changes in hepatic gene expression in rats that received bromobenzene to induce acute hepatic centrilobular necrosis. Bromobenzene-induced lesions were only observed after treatment with the highest of 3 dose levels. Multivariate statistical analysis showed that metabolite profiles of blood plasma were largely different from controls when the rats were treated with bromobenzene, also at doses that did not elicit histopathological changes. Changes in levels of genes and metabolites were related to the degree of necrosis, providing putative novel markers of hepatotoxicity. Levels of endogenous metabolites like alanine, lactate, tyrosine and dimethylglycine differed in plasma from treated and control rats. The metabolite profiles of urine were found to be reflective of the exposure levels. This integrated analysis of hepatic transcriptomics and plasma metabolomics was able to more sensitively detect changes related to hepatotoxicity and discover novel markers. The relation between gene expression and metabolite levels was explored and additional insight in the role of various biological pathways in bromobenzene-induced hepatic necrosis was obtained, including the involvement of apoptosis and changes in glycolysis and amino acid metabolism. The complete Table 2 is available as a supplemental file online at http://taylorandfrancis.metapress.com/openurlasp?genre=journal&issn=0192-6233. To access the file, click on the issue link for 33(4), then select this article. A download option appears at the bottom of this abstract. In order to access the full article online, you must either have an individual subscription or a member subscription accessed through www.toxpath.org.

Transcriptomics analysis of lungs and peripheral blood of crystalline silica-exposed rats

PubMed Central

Sellamuthu, Rajendran; Umbright, Christina; Roberts, Jenny R.; Chapman, Rebecca; Young, Shih-Houng; Richardson, Diana; Cumpston, Jared; McKinney, Walter; Chen, Bean T.; Frazer, David; Li, Shengqiao; Kashon, Michael; Joseph, Pius

2015-01-01

Minimally invasive approaches to detect/predict target organ toxicity have significant practical applications in occupational toxicology. The potential application of peripheral blood transcriptomics as a practical approach to study the mechanisms of silica-induced pulmonary toxicity was investigated. Rats were exposed by inhalation to crystalline silica (15 mg/m3, 6 h/day, 5 days) and pulmonary toxicity and global gene expression profiles of lungs and peripheral blood were determined at 32 weeks following termination of exposure. A significant elevation in bronchoalveolar lavage fluid lactate dehydrogenase activity and moderate histological changes in the lungs, including type II pneumocyte hyperplasia and fibrosis, indicated pulmonary toxicity in the rats. Similarly, significant infiltration of neutrophils and elevated monocyte chemotactic protein-1 levels in the lungs showed pulmonary inflammation in the rats. Microarray analysis of global gene expression profiles identified significant differential expression [>1.5-fold change and false discovery rate (FDR) p < 0.01] of 520 and 537 genes, respectively, in the lungs and blood of the exposed rats. Bioinformatics analysis of the differentially expressed genes demonstrated significant similarity in the biological processes, molecular networks, and canonical pathways enriched by silica exposure in the lungs and blood of the rats. Several genes involved in functions relevant to silica-induced pulmonary toxicity such as inflammation, respiratory diseases, cancer, cellular movement, fibrosis, etc, were found significantly differentially expressed in the lungs and blood of the silica-exposed rats. The results of this study suggested the potential application of peripheral blood gene expression profiling as a toxicologically relevant and minimally invasive surrogate approach to study the mechanisms underlying silica-induced pulmonary toxicity. PMID:22861000

Dissection of Symbiosis and Organ Development by Integrated Transcriptome Analysis of Lotus japonicus Mutant and Wild-Type Plants

PubMed Central

Høgslund, Niels; Radutoiu, Simona; Krusell, Lene; Voroshilova, Vera; Hannah, Matthew A.; Goffard, Nicolas; Sanchez, Diego H.; Lippold, Felix; Ott, Thomas; Sato, Shusei; Tabata, Satoshi; Liboriussen, Poul; Lohmann, Gitte V.; Schauser, Leif; Weiller, Georg F.; Udvardi, Michael K.; Stougaard, Jens

2009-01-01

Genetic analyses of plant symbiotic mutants has led to the identification of key genes involved in Rhizobium-legume communication as well as in development and function of nitrogen fixing root nodules. However, the impact of these genes in coordinating the transcriptional programs of nodule development has only been studied in limited and isolated studies. Here, we present an integrated genome-wide analysis of transcriptome landscapes in Lotus japonicus wild-type and symbiotic mutant plants. Encompassing five different organs, five stages of the sequentially developed determinate Lotus root nodules, and eight mutants impaired at different stages of the symbiotic interaction, our data set integrates an unprecedented combination of organ- or tissue-specific profiles with mutant transcript profiles. In total, 38 different conditions sampled under the same well-defined growth regimes were included. This comprehensive analysis unravelled new and unexpected patterns of transcriptional regulation during symbiosis and organ development. Contrary to expectations, none of the previously characterized nodulins were among the 37 genes specifically expressed in nodules. Another surprise was the extensive transcriptional response in whole root compared to the susceptible root zone where the cellular response is most pronounced. A large number of transcripts predicted to encode transcriptional regulators, receptors and proteins involved in signal transduction, as well as many genes with unknown function, were found to be regulated during nodule organogenesis and rhizobial infection. Combining wild type and mutant profiles of these transcripts demonstrates the activation of a complex genetic program that delineates symbiotic nitrogen fixation. The complete data set was organized into an indexed expression directory that is accessible from a resource database, and here we present selected examples of biological questions that can be addressed with this comprehensive and powerful gene expression data set. PMID:19662091

Hepatic Transcriptome Responses in Mice (Mus musculus) Exposed to the Nafion Membrane and Its Combustion Products

PubMed Central

Feng, Mingbao; Qu, Ruijuan; Habteselassie, Mussie; Wu, Jun; Yang, Shaogui; Sun, Ping; Huang, Qingguo; Wang, Zunyao

2015-01-01

Nafion 117 membrane (N117), an important polymer electrolyte membrane (PEM), has been widely used for numerous chemical technologies. Despite its increasing production and use, the toxicity data for N117 and its combustion products remain lacking. Toxicity studies are necessary to avoid problems related to waste disposal in landfills and incineration that may arise. In this study, we investigated the histopathological alterations, oxidative stress biomarker responses, and transcriptome profiles in the liver of male mice exposed to N117 and its combustion products for 24 days. An ion-chromatography system and liquid chromatography system coupled to a hybrid quadrupole time-of-flight mass spectrometry were used to analyze the chemical compositions of these combustion products. The transcriptomics analysis identified several significantly altered molecular pathways, including the metabolism of xenobiotics, carbohydrates and lipids; signal transduction; cellular processes; immune system; and signaling molecules and interaction. These studies provide preliminary data for the potential toxicity of N117 and its combustion products on living organisms and may fill the information gaps in the toxicity databases for the currently used PEMs. PMID:26057616

A Single-Cell Approach to the Elusive Latent Human Cytomegalovirus Transcriptome.

PubMed

Goodrum, Felicia; McWeeney, Shannon

2018-06-12

Herpesvirus latency has been difficult to understand molecularly due to low levels of viral genomes and gene expression. In the case of the betaherpesvirus human cytomegalovirus (HCMV), this is further complicated by the heterogeneity inherent to hematopoietic subpopulations harboring genomes and, as a consequence, the various patterns of infection that simultaneously exist in a host, ranging from latent to lytic. Single-cell RNA sequencing (scRNA-seq) provides tremendous potential in measuring the gene expression profiles of heterogeneous cell populations for a wide range of applications, including in studies of cancer, immunology, and infectious disease. A recent study by Shnayder et al. (mBio 9:e00013-18, 2018, https://doi.org/10.1128/mBio.00013-18) utilized scRNA-seq to define transcriptomal characteristics of HCMV latency. They conclude that latency-associated gene expression is similar to the late lytic viral program but at lower levels of expression. The study highlights the numerous challenges, from the definition of latency to the analysis of scRNA-seq, that exist in defining a latent transcriptome. Copyright © 2018 Goodrum and McWeeney.

Transcriptome analysis of soiny mullet (Liza haematocheila) spleen in response to Streptococcus dysgalactiae.

PubMed

Qi, Zhitao; Wu, Ping; Zhang, Qihuan; Wei, Youchuan; Wang, Zisheng; Qiu, Ming; Shao, Rong; Li, Yao; Gao, Qian

2016-02-01

Soiny mullet (Liza haematocheila) is becoming an economically important aquaculture mugilid species in China and other Asian countries. However, increasing incidences of bacterial pathogenic diseases has greatly hampered the production of the soiny mullet. Deeper understanding of the soiny mullet immune system and its related genes in response to bacterial infections are necessary for disease control in this species. In this study, the transcriptomic profile of spleen from soiny mullet challenged with Streptococcus dysgalactiae was analyzed by Illumina-based paired-end sequencing method. After assembly, 86,884 unique transcript fragments (unigenes) were assembled, with an average length of 991 bp. Approximately 41,795 (48.1%) unigenes were annotated in the nr NCBI database and 57.9% of the unigenes were similar to that of the Nile tilapia. A total of 24,299 unigenes were categorized into three Gene Ontology (GO) categories (molecular function, cellular component and biological process), 13,570 unigenes into 25 functional Clusters of Orthologous Groups of proteins (COG) categories, and 30,547 unigenes were grouped into 258 known pathways in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Following S. dysgalactiae infection, 11,461 differentially expressed unigenes were identified including 4658 up-regulated unigenes and 6803 down-regulated unigenes. Significant enrichment analysis of these differentially expressed unigenes identified major immune related pathways, including the Toll-like receptor, complement and coagulation cascades, T cell receptor signaling pathway and B cell receptor signaling pathway. In addition, 24,813 simple sequence repeats (SSRs) and 127,503 candidate single nucleotide polymorphisms (SNPs) were identified from the mullet spleen transcriptome. To this date, this study has globally analyzed the transcriptome profile from the spleen of L. haematocheila after S. dysgalactiae infection. Therefore, the results of our study contributes to better on the immune system and defense mechanisms of soiny mullet in response to bacterial infection, and provides valuable references for related studies in mugilidae species which currently lack genomic reference. Copyright © 2015 Elsevier Ltd. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Jing; Ma, Zihao; Carr, Steven A.

Coexpression of mRNAs under multiple conditions is commonly used to infer cofunctionality of their gene products despite well-known limitations of this “guilt-by-association” (GBA) approach. Recent advancements in mass spectrometry-based proteomic technologies have enabled global expression profiling at the protein level; however, whether proteome profiling data can outperform transcriptome profiling data for coexpression based gene function prediction has not been systematically investigated. Here, we address this question by constructing and analyzing mRNA and protein coexpression networks for three cancer types with matched mRNA and protein profiling data from The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC).more » Our analyses revealed a marked difference in wiring between the mRNA and protein coexpression networks. Whereas protein coexpression was driven primarily by functional similarity between coexpressed genes, mRNA coexpression was driven by both cofunction and chromosomal colocalization of the genes. Functionally coherent mRNA modules were more likely to have their edges preserved in corresponding protein networks than functionally incoherent mRNA modules. Proteomic data strengthened the link between gene expression and function for at least 75% of Gene Ontology (GO) biological processes and 90% of KEGG pathways. A web application Gene2Net (http://cptac.gene2net.org) developed based on the three protein coexpression networks revealed novel gene-function relationships, such as linking ERBB2 (HER2) to lipid biosynthetic process in breast cancer, identifying PLG as a new gene involved in complement activation, and identifying AEBP1 as a new epithelial-mesenchymal transition (EMT) marker. Our results demonstrate that proteome profiling outperforms transcriptome profiling for coexpression based gene function prediction. Proteomics should be integrated if not preferred in gene function and human disease studies. Molecular & Cellular Proteomics 16: 10.1074/mcp.M116.060301, 121–134, 2017.« less

Identifying potential RNAi targets in grain aphid (Sitobion avenae F.) based on transcriptome profiling of its alimentary canal after feeding on wheat plants

PubMed Central

2013-01-01

Background The grain aphid (Sitobion avenae F.) is a major agricultural pest which causes significant yield losses of wheat in China, Europe and North America annually. Transcriptome profiling of the grain aphid alimentary canal after feeding on wheat plants could provide comprehensive gene expression information involved in feeding, ingestion and digestion. Furthermore, selection of aphid-specific RNAi target genes would be essential for utilizing a plant-mediated RNAi strategy to control aphids via a non-toxic mode of action. However, due to the tiny size of the alimentary canal and lack of genomic information on grain aphid as a whole, selection of the RNAi targets is a challenging task that as far as we are aware, has never been documented previously. Results In this study, we performed de novo transcriptome assembly and gene expression analyses of the alimentary canals of grain aphids before and after feeding on wheat plants using Illumina RNA sequencing. The transcriptome profiling generated 30,427 unigenes with an average length of 664 bp. Furthermore, comparison of the transcriptomes of alimentary canals of pre- and post feeding grain aphids indicated that 5490 unigenes were differentially expressed, among which, diverse genes and/or pathways were identified and annotated. Based on the RPKM values of these unigenes, 16 of them that were significantly up or down-regulated upon feeding were selected for dsRNA artificial feeding assay. Of these, 5 unigenes led to higher mortality and developmental stunting in an artificial feeding assay due to the down-regulation of the target gene expression. Finally, by adding fluorescently labelled dsRNA into the artificial diet, the spread of fluorescence signal in the whole body tissues of grain aphid was observed. Conclusions Comparison of the transcriptome profiles of the alimentary canals of pre- and post-feeding grain aphids on wheat plants provided comprehensive gene expression information that could facilitate our understanding of the molecular mechanisms underlying feeding, ingestion and digestion. Furthermore, five novel and effective potential RNAi target genes were identified in grain aphid for the first time. This finding would provide a fundamental basis for aphid control in wheat through plant mediated RNAi strategy. PMID:23957588

Gene Expression Profiling of Acute Lymphoblastic Leukemia in Children with Very Early Relapse.

PubMed

Núñez-Enríquez, Juan Carlos; Bárcenas-López, Diego Alberto; Hidalgo-Miranda, Alfredo; Jiménez-Hernández, Elva; Bekker-Méndez, Vilma Carolina; Flores-Lujano, Janet; Solis-Labastida, Karina Anastacia; Martínez-Morales, Gabriela Bibiana; Sánchez-Muñoz, Fausto; Espinoza-Hernández, Laura Eugenia; Velázquez-Aviña, Martha Margarita; Merino-Pasaye, Laura Elizabeth; García Velázquez, Alejandra Jimena; Pérez-Saldívar, María Luisa; Mojica-Espinoza, Raúl; Ramírez-Bello, Julián; Jiménez-Morales, Silvia; Mejía-Aranguré, Juan Manuel

2016-11-01

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer worldwide. Mexican patients have high mortality rates, low frequency of good prognosis biomarkers (i.e., ETV6-RUNX1) and a high proportion is classified at the time of diagnosis with a high risk to relapse according to clinical features. In addition, very early relapses are more frequently observed than in other populations. The aim of the study was to identify new potential biomarkers associated with very early relapse in Mexican ALL children through transcriptome analysis. Microarray gene expression profiling on bone marrow samples of 54 pediatric ALL patients, collected at time of diagnosis and/or at relapse, was performed. Eleven patients presented relapse within the first 18 months after diagnosis. Affymetrix Human Transcriptome Array 2.0 (HTA 2.0) was used to perform gene expression analysis. Annotation and functional enrichment analyses were carried out using Gene Ontology, KEGG pathway analysis and Ingenuity Pathway Analysis tools. BLVRB, ZCCHC7, PAX5, EBF1, TMOD1 and BLNK were differentially expressed (fold-change >2.0 and p value <0.01) between relapsed and non-relapsed patients. Functional analysis of abnormally expressed genes revealed their important role in cellular processes related to the development of hematological diseases, cancer, cell death and survival and in cell-to-cell signaling interaction. Our data support previous findings showing the relevance of PAX5, EBF1 and ZCCHC7 as potential biomarkers to identify a subgroup of ALL children in high risk to relapse. Copyright © 2016 IMSS. Published by Elsevier Inc. All rights reserved.

Spectroscopic and Statistical Techniques for Information Recovery in Metabonomics and Metabolomics

NASA Astrophysics Data System (ADS)

Lindon, John C.; Nicholson, Jeremy K.

2008-07-01

Methods for generating and interpreting metabolic profiles based on nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (MS), and chemometric analysis methods are summarized and the relative strengths and weaknesses of NMR and chromatography-coupled MS approaches are discussed. Given that all data sets measured to date only probe subsets of complex metabolic profiles, we describe recent developments for enhanced information recovery from the resulting complex data sets, including integration of NMR- and MS-based metabonomic results and combination of metabonomic data with data from proteomics, transcriptomics, and genomics. We summarize the breadth of applications, highlight some current activities, discuss the issues relating to metabonomics, and identify future trends.

Spectroscopic and statistical techniques for information recovery in metabonomics and metabolomics.

PubMed

Lindon, John C; Nicholson, Jeremy K

2008-01-01

Methods for generating and interpreting metabolic profiles based on nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (MS), and chemometric analysis methods are summarized and the relative strengths and weaknesses of NMR and chromatography-coupled MS approaches are discussed. Given that all data sets measured to date only probe subsets of complex metabolic profiles, we describe recent developments for enhanced information recovery from the resulting complex data sets, including integration of NMR- and MS-based metabonomic results and combination of metabonomic data with data from proteomics, transcriptomics, and genomics. We summarize the breadth of applications, highlight some current activities, discuss the issues relating to metabonomics, and identify future trends.

Genome wide transcriptional profiling of Herbaspirillum seropedicae SmR1 grown in the presence of naringenin.

PubMed

Tadra-Sfeir, Michelle Z; Faoro, Helisson; Camilios-Neto, Doumit; Brusamarello-Santos, Liziane; Balsanelli, Eduardo; Weiss, Vinicius; Baura, Valter A; Wassem, Roseli; Cruz, Leonardo M; De Oliveira Pedrosa, Fábio; Souza, Emanuel M; Monteiro, Rose A

2015-01-01

Herbaspirillum seropedicae is a diazotrophic bacterium which associates endophytically with economically important gramineae. Flavonoids such as naringenin have been shown to have an effect on the interaction between H. seropedicae and its host plants. We used a high-throughput sequencing based method (RNA-Seq) to access the influence of naringenin on the whole transcriptome profile of H. seropedicae. Three hundred and four genes were downregulated and seventy seven were upregulated by naringenin. Data analysis revealed that genes related to bacterial flagella biosynthesis, chemotaxis and biosynthesis of peptidoglycan were repressed by naringenin. Moreover, genes involved in aromatic metabolism and multidrug transport efllux were actived.

De novo Transcriptome Assembly of Common Wild Rice (Oryza rufipogon Griff.) and Discovery of Drought-Response Genes in Root Tissue Based on Transcriptomic Data.

PubMed

Tian, Xin-Jie; Long, Yan; Wang, Jiao; Zhang, Jing-Wen; Wang, Yan-Yan; Li, Wei-Min; Peng, Yu-Fa; Yuan, Qian-Hua; Pei, Xin-Wu

2015-01-01

The perennial O. rufipogon (common wild rice), which is considered to be the ancestor of Asian cultivated rice species, contains many useful genetic resources, including drought resistance genes. However, few studies have identified the drought resistance and tissue-specific genes in common wild rice. In this study, transcriptome sequencing libraries were constructed, including drought-treated roots (DR) and control leaves (CL) and roots (CR). Using Illumina sequencing technology, we generated 16.75 million bases of high-quality sequence data for common wild rice and conducted de novo assembly and annotation of genes without prior genome information. These reads were assembled into 119,332 unigenes with an average length of 715 bp. A total of 88,813 distinct sequences (74.42% of unigenes) significantly matched known genes in the NCBI NT database. Differentially expressed gene (DEG) analysis showed that 3617 genes were up-regulated and 4171 genes were down-regulated in the CR library compared with the CL library. Among the DEGs, 535 genes were expressed in roots but not in shoots. A similar comparison between the DR and CR libraries showed that 1393 genes were up-regulated and 315 genes were down-regulated in the DR library compared with the CR library. Finally, 37 genes that were specifically expressed in roots were screened after comparing the DEGs identified in the above-described analyses. This study provides a transcriptome sequence resource for common wild rice plants and establishes a digital gene expression profile of wild rice plants under drought conditions using the assembled transcriptome data as a reference. Several tissue-specific and drought-stress-related candidate genes were identified, representing a fully characterized transcriptome and providing a valuable resource for genetic and genomic studies in plants.

Metformin-Induced Changes of the Coding Transcriptome and Non-Coding RNAs in the Livers of Non-Alcoholic Fatty Liver Disease Mice.

PubMed

Guo, Jun; Zhou, Yuan; Cheng, Yafen; Fang, Weiwei; Hu, Gang; Wei, Jie; Lin, Yajun; Man, Yong; Guo, Lixin; Sun, Mingxiao; Cui, Qinghua; Li, Jian

2018-01-01

Recent studies have suggested that changes in non-coding mRNA play a key role in the progression of non-alcoholic fatty liver disease (NAFLD). Metformin is now recommended and effective for the treatment of NAFLD. We hope the current analyses of the non-coding mRNA transcriptome will provide a better presentation of the potential roles of mRNAs and long non-coding RNAs (lncRNAs) that underlie NAFLD and metformin intervention. The present study mainly analysed changes in the coding transcriptome and non-coding RNAs after the application of a five-week metformin intervention. Liver samples from three groups of mice were harvested for transcriptome profiling, which covered mRNA, lncRNA, microRNA (miRNA) and circular RNA (circRNA), using a microarray technique. A systematic alleviation of high-fat diet (HFD)-induced transcriptome alterations by metformin was observed. The metformin treatment largely reversed the correlations with diabetes-related pathways. Our analysis also suggested interaction networks between differentially expressed lncRNAs and known hepatic disease genes and interactions between circRNA and their disease-related miRNA partners. Eight HFD-responsive lncRNAs and three metformin-responsive lncRNAs were noted due to their widespread associations with disease genes. Moreover, seven miRNAs that interacted with multiple differentially expressed circRNAs were highlighted because they were likely to be associated with metabolic or liver diseases. The present study identified novel changes in the coding transcriptome and non-coding RNAs in the livers of NAFLD mice after metformin treatment that might shed light on the underlying mechanism by which metformin impedes the progression of NAFLD. © 2018 The Author(s). Published by S. Karger AG, Basel.

Prior to extension, Transcriptomes of fibroblast-like Synoviocytes from extended and Polyarticular juvenile idiopathic arthritis are indistinguishable.

PubMed

Brescia, AnneMarie C; Simonds, Megan M; McCahan, Suzanne M; Sullivan, Kathleen E; Rose, Carlos D

2018-01-08

Our intent was to identify differences between the transcriptome of fibroblast-like synoviocytes (FLS) in oligoarticular juvenile idiopathic arthritis (JIA) before extension when compared to persistent subtype of JIA, when the two are clinically indistinguishable. Additionally, we sought to determine if differences between the transcriptomes of FLS from extended-to-be and polyarticular course JIA could be detected. Our hypothesis was that intrinsic differences in the transcriptome of the FLS from extended-to-be JIA would distinguish them from persistent oligoarticular JIA, before the course is clinically apparent. Global gene expression was defined in cultured FLS from 6 controls, 12 JIA with persistent course, 7 JIA prior to extension (extended-to-be), 4 JIA with extended course and 6 polyarticular onset, using Affymetrix Human GeneChips 133plus2.0. Bioconductor Linear Models for Microarray Analysis revealed 22 probesets with differential expression between persistent and extended-to-be FLS at 15% FDR, however only 2 probesets distinguished extended-to-be from extended and none distinguished extended-to-be and polyarticular at 15% FDR. Differences in extended and polyarticular gene expression profiles were not detected. Confirmation of select genes was done on the RNA level by RT-qPCR and on the protein level in synovial fluid by ELISA. The transcriptome of FLS from extended-to-be juvenile idiopathic arthritis is distinct from persistent course before a clinical distinction can be made. Additionally, the transcriptome of extended-to-be and polyarticular course, including those who have already extended, are indistinguishable. These gene expression data suggest that FLS already reflect a polyarticular behavior early in disease course, suggesting that extended-to-be may be "latent polyarticular" at onset. These differences can be used to develop early biomarkers of disease course, allowing for better-informed treatment decisions.

Next-Generation Transcriptome Profiling of the Salmon Louse Caligus rogercresseyi Exposed to Deltamethrin (AlphaMax™): Discovery of Relevant Genes and Sex-Related Differences.

PubMed

Chávez-Mardones, Jacqueline; Gallardo-Escárate, Cristian

2015-12-01

Sea lice are one of the main parasites affecting the salmon aquaculture industry, causing significant economic losses worldwide. Increased resistance to traditional chemical treatments has created the need to find alternative control methods. Therefore, the objective of this study was to identify the transcriptome response of the salmon louse Caligus rogercresseyi to the delousing drug deltamethrin (AlphaMax™). Through bioassays with different concentrations of deltamethrin, adult salmon lice transcriptomes were sequenced from cDNA libraries in the MiSeq Illumina platform. A total of 78 million reads for females and males were assembled in 30,212 and 38,536 contigs, respectively. De novo assembly yielded 86,878 high-quality contigs and, based on published data, it was possible to annotate and identify relevant genes involved in several biological processes. RNA-seq analysis in conjunction with heatmap hierarchical clustering evidenced that pyrethroids modify the ectoparasitic transcriptome in adults, affecting molecular processes associated with the nervous system, cuticle formation, oxidative stress, reproduction, and metabolism, among others. Furthermore, sex-related transcriptome differences were evidenced. Specifically, 534 and 1033 exclusive transcripts were identified for males and females, respectively, and 154 were shared between sexes. For males, estradiol 17-beta-dehydrogenase, sphingolipid delta4-desaturase DES1, ketosamine-3-kinase, and arylsulfatase A, among others, were discovered, while for females, vitellogenin 1, glycoprotein G, transaldolase, and nitric oxide synthase were among those identified. The shared transcripts included annotations for tropomyosin, γ-crystallin A, glutamate receptor-metabotropic, glutathione S-transferase, and carboxipeptidase B. The present study reveals that deltamethrin generates a complex transcriptome response in C. rogercresseyi, thus providing valuable genomic information for developing new delousing drugs.

RNA-seq reveals transcriptome changes in goats following myostatin gene knockout

PubMed Central

Cai, Bei; Zhou, Shiwei; Zhu, Haijing; Qu, Lei; Wang, Xiaolong

2017-01-01

Myostatin (MSTN) is a powerful negative regulator of skeletal muscle mass in mammalian species that is primarily expressed in skeletal muscles, and mutations of its encoding gene can result in the double-muscling trait. In this study, the CRISPR/Cas9 technique was used to edit MSTN in Shaanbei Cashmere goats and generate knockout animals. RNA sequencing was used to determine and compare the transcriptome profiles of the muscles from three wild-type (WT) goats, three fibroblast growth factor 5 (FGF5) knockout goats (FGF5+/- group) and three goats with disrupted expression of both the FGF5 and MSTN genes (FM+/- group). The sequence reads were obtained using the Illumina HiSeq 2000 system and mapped to the Capra hircus reference genome using TopHat (v2.0.9). In total, 68.93, 62.04 and 66.26 million clean sequencing reads were obtained from the WT, FM+/- and FGF5+/- groups, respectively. There were 201 differentially expressed genes (DEGs) between the WT and FGF5+/- groups, with 86 down- and 115 up-regulated genes in the FGF5+/- group. Between the WT and FM+/- groups, 121 DEGs were identified, including 81 down- and 40 up-regulated genes in the FM+/- group. A total of 198 DEGs were detected between the FGF5+/- group and FM+/- group, with 128 down- and 70 up-regulated genes in the FM+/- group. At the transcriptome level, we found substantial changes in genes involved in fatty acid metabolism and the biosynthesis of unsaturated fatty acids, such as stearoyl-CoA dehydrogenase, 3-hydroxyacyl-CoA dehydratase 2, ELOVL fatty acid elongase 6 and fatty acid synthase, suggesting that the expression levels of these genes may be directly regulated by MSTN and that these genes are likely downstream targets of MSTN with potential roles in lipid metabolism in goats. Moreover, five randomly selected DEGs were further validated with qRT-PCR, and the results were consistent with the transcriptome analysis. The present study provides insight into the unique transcriptome profile of the MSTN knockout goat, which is a valuable resource for studying goat genomics. PMID:29228005

A detailed gene expression study of the Miscanthus genus reveals changes in the transcriptome associated with the rejuvenation of spring rhizomes.

PubMed

Barling, Adam; Swaminathan, Kankshita; Mitros, Therese; James, Brandon T; Morris, Juliette; Ngamboma, Ornella; Hall, Megan C; Kirkpatrick, Jessica; Alabady, Magdy; Spence, Ashley K; Hudson, Matthew E; Rokhsar, Daniel S; Moose, Stephen P

2013-12-09

The Miscanthus genus of perennial C4 grasses contains promising biofuel crops for temperate climates. However, few genomic resources exist for Miscanthus, which limits understanding of its interesting biology and future genetic improvement. A comprehensive catalog of expressed sequences were generated from a variety of Miscanthus species and tissue types, with an emphasis on characterizing gene expression changes in spring compared to fall rhizomes. Illumina short read sequencing technology was used to produce transcriptome sequences from different tissues and organs during distinct developmental stages for multiple Miscanthus species, including Miscanthus sinensis, Miscanthus sacchariflorus, and their interspecific hybrid Miscanthus × giganteus. More than fifty billion base-pairs of Miscanthus transcript sequence were produced. Overall, 26,230 Sorghum gene models (i.e., ~ 96% of predicted Sorghum genes) had at least five Miscanthus reads mapped to them, suggesting that a large portion of the Miscanthus transcriptome is represented in this dataset. The Miscanthus × giganteus data was used to identify genes preferentially expressed in a single tissue, such as the spring rhizome, using Sorghum bicolor as a reference. Quantitative real-time PCR was used to verify examples of preferential expression predicted via RNA-Seq. Contiguous consensus transcript sequences were assembled for each species and annotated using InterProScan. Sequences from the assembled transcriptome were used to amplify genomic segments from a doubled haploid Miscanthus sinensis and from Miscanthus × giganteus to further disentangle the allelic and paralogous variations in genes. This large expressed sequence tag collection creates a valuable resource for the study of Miscanthus biology by providing detailed gene sequence information and tissue preferred expression patterns. We have successfully generated a database of transcriptome assemblies and demonstrated its use in the study of genes of interest. Analysis of gene expression profiles revealed biological pathways that exhibit altered regulation in spring compared to fall rhizomes, which are consistent with their different physiological functions. The expression profiles of the subterranean rhizome provides a better understanding of the biological activities of the underground stem structures that are essentials for perenniality and the storage or remobilization of carbon and nutrient resources.

Transcriptional analysis of Myceliophthora thermophila on soluble starch and role of regulator AmyR on polysaccharide degradation.

PubMed

Xu, Guanbao; Li, Jingen; Liu, Qian; Sun, Wenliang; Jiang, Min; Tian, Chaoguang

2018-05-24

Thermophilic fungus Myceliophthora thermophila has great capacity for biomass degradation and is an attractive option for use as cell factory to produce chemicals directly from renewable polysaccharides, such as starch, rather than monomer glucose. To date, there has been no transcriptomic analysis of this thermophilic fungus on starch. This study determined the transcriptomic profile of M. thermophila responding to soluble starch and a 342-gene set was identified as a "starch regulon", including the major amylolytic enzyme (Mycth_72393). Its overexpression led to increased amylase activities on starch by 35%. Furthermore, overexpressing the key amylolytic enzyme regulator AmyR in M. thermophila significantly increased amylase activity by 30%. Deletion of amyR by the CRISPR/Cas9 system led to the relief of carbon catabolite repression and 3-fold increased lignocellulase activities on cellulose. This study will accelerate rational fungal strain engineering for biochemical production from biomass substrates such as raw corn starch and even crop straw. Copyright © 2018 Elsevier Ltd. All rights reserved.

SAGE Analysis of Transcriptome Responses in Arabidopsis Roots Exposed to 2,4,6-Trinitrotoluene1

PubMed Central

Ekman, Drew R.; Lorenz, W. Walter; Przybyla, Alan E.; Wolfe, N. Lee; Dean, Jeffrey F.D.

2003-01-01

Serial analysis of gene expression was used to profile transcript levels in Arabidopsis roots and assess their responses to 2,4,6-trinitrotoluene (TNT) exposure. SAGE libraries representing control and TNT-exposed seedling root transcripts were constructed, and each was sequenced to a depth of roughly 32,000 tags. More than 19,000 unique tags were identified overall. The second most highly induced tag (27-fold increase) represented a glutathione S-transferase. Cytochrome P450 enzymes, as well as an ABC transporter and a probable nitroreductase, were highly induced by TNT exposure. Analyses also revealed an oxidative stress response upon TNT exposure. Although some increases were anticipated in light of current models for xenobiotic metabolism in plants, evidence for unsuspected conjugation pathways was also noted. Identifying transcriptome-level responses to TNT exposure will better define the metabolic pathways plants use to detoxify this xenobiotic compound, which should help improve phytoremediation strategies directed at TNT and other nitroaromatic compounds. PMID:14551330

Transcriptome Analysis of the Role of GlnD/GlnBK in Nitrogen Stress Adaptation by Sinorhizobium meliloti Rm1021

PubMed Central

Yurgel, Svetlana N.; Rice, Jennifer; Kahn, Michael L.

2013-01-01

Transcriptional changes in the nitrogen stress response (NSR) of wild type S. meliloti Rm1021, and isogenic strains missing both PII proteins, GlnB and GlnK, or carrying a ΔglnD-sm2 mutation were analyzed using whole-genome microarrays. This approach allowed us to identify a number of new genes involved in the NSR and showed that the response of these bacteria to nitrogen stress overlaps with other stress responses, including induction of the fixK2 transcriptional activator and genes that are part of the phosphate stress response. Our data also show that GlnD and GlnBK proteins may regulate many genes that are not part of the NSR. Analysis of transcriptome profiles of the Rm1021 ΔglnD-sm2 strain allowed us to identify several genes that appear to be regulated by GlnD without the participation of the PII proteins. PMID:23516427

Transcriptome analysis of PCOS arrested 2-cell embryos.

PubMed

Lu, Cuiling; Chi, Hongbin; Wang, Yapeng; Feng, Xue; Wang, Lina; Huang, Shuo; Yan, Liying; Lin, Shengli; Liu, Ping; Qiao, Jie

2018-06-18

In an attempt to explore the early developmental arrest in embryos from polycystic ovarian syndrome (PCOS) patients, we sequenced the transcriptome profiles of PCOS arrested 2-cell embryos, non-PCOS arrested 2-cell embryos and non-arrested 2-cell embryos using single-cell RNA-Seq technique. Differential expression analysis was performed using the DEGSeq R package. Gene Ontology (GO) enrichment was analyzed using the GOseq R package. Data revealed 62 differentially expressed genes between non-PCOS arrested and PCOS arrested embryos and 2217 differentially expressed genes between PCOS arrested and non-arrested 2-cell embryos. A total of 49 differently expressed genes (DEGs) were annotated with GO terms in the up-regulated genes between PCOS arrested and non-PCOS arrested embryos after GO enrichment. A total of 29 DEGs were annotated with GO terms in the down-regulated genes between PCOS arrested and non-arrested 2-cell embryos after GO enrichment. These data can provide a reference for screening specific genes involved in the arrest of PCOS embryos.

Mediterranean versus Red sea corals facing climate change, a transcriptome analysis

NASA Astrophysics Data System (ADS)

Maor-Landaw, Keren; Waldman Ben-Asher, Hiba; Karako-Lampert, Sarit; Salmon-Divon, Mali; Prada, Fiorella; Caroselli, Erik; Goffredo, Stefano; Falini, Giuseppe; Dubinsky, Zvy; Levy, Oren

2017-02-01

The anthropogenic increase in atmospheric CO2 that drives global warming and ocean acidification raises serious concerns regarding the future of corals, the main carbonate biomineralizers. Here we used transcriptome analysis to study the effect of long-term gradual temperature increase (annual rate), combined with lowered pH values, on a sub-tropical Red Sea coral, Stylophora pistillata, and on a temperate Mediterranean symbiotic coral Balanophyllia europaea. The gene expression profiles revealed a strong effect of both temperature increase and pH decrease implying for synergism response. The temperate coral, exposed to a twice as high range of seasonal temperature fluctuations than the Red Sea species, faced stress more effectively. The compensatory strategy for coping apparently involves deviating cellular resources into a massive up-regulation of genes in general, and specifically of genes involved in the generation of metabolic energy. Our results imply that sub-lethal, prolonged exposure to stress can stimulate evolutionary increase in stress resilience.

Gene expression profiling of human breast tissue samples using SAGE-Seq.

PubMed

Wu, Zhenhua Jeremy; Meyer, Clifford A; Choudhury, Sibgat; Shipitsin, Michail; Maruyama, Reo; Bessarabova, Marina; Nikolskaya, Tatiana; Sukumar, Saraswati; Schwartzman, Armin; Liu, Jun S; Polyak, Kornelia; Liu, X Shirley

2010-12-01

We present a powerful application of ultra high-throughput sequencing, SAGE-Seq, for the accurate quantification of normal and neoplastic mammary epithelial cell transcriptomes. We develop data analysis pipelines that allow the mapping of sense and antisense strands of mitochondrial and RefSeq genes, the normalization between libraries, and the identification of differentially expressed genes. We find that the diversity of cancer transcriptomes is significantly higher than that of normal cells. Our analysis indicates that transcript discovery plateaus at 10 million reads/sample, and suggests a minimum desired sequencing depth around five million reads. Comparison of SAGE-Seq and traditional SAGE on normal and cancerous breast tissues reveals higher sensitivity of SAGE-Seq to detect less-abundant genes, including those encoding for known breast cancer-related transcription factors and G protein-coupled receptors (GPCRs). SAGE-Seq is able to identify genes and pathways abnormally activated in breast cancer that traditional SAGE failed to call. SAGE-Seq is a powerful method for the identification of biomarkers and therapeutic targets in human disease.

Single-cell mass cytometry and transcriptome profiling reveal the impact of graphene on human immune cells.

PubMed

Orecchioni, Marco; Bedognetti, Davide; Newman, Leon; Fuoco, Claudia; Spada, Filomena; Hendrickx, Wouter; Marincola, Francesco M; Sgarrella, Francesco; Rodrigues, Artur Filipe; Ménard-Moyon, Cécilia; Cesareni, Gianni; Kostarelos, Kostas; Bianco, Alberto; Delogu, Lucia G

2017-10-24

Understanding the biomolecular interactions between graphene and human immune cells is a prerequisite for its utilization as a diagnostic or therapeutic tool. To characterize the complex interactions between graphene and immune cells, we propose an integrative analytical pipeline encompassing the evaluation of molecular and cellular parameters. Herein, we use single-cell mass cytometry to dissect the effects of graphene oxide (GO) and GO functionalized with amino groups (GONH 2 ) on 15 immune cell populations, interrogating 30 markers at the single-cell level. Next, the integration of single-cell mass cytometry with genome-wide transcriptome analysis shows that the amine groups reduce the perturbations caused by GO on cell metabolism and increase biocompatibility. Moreover, GONH 2 polarizes T-cell and monocyte activation toward a T helper-1/M1 immune response. This study describes an innovative approach for the analysis of the effects of nanomaterials on distinct immune cells, laying the foundation for the incorporation of single-cell mass cytometry on the experimental pipeline.

Differential Gene Expression between Leaf and Rhizome in Atractylodes lancea: A Comparative Transcriptome Analysis

PubMed Central

Huang, Qianqian; Huang, Xiao; Deng, Juan; Liu, Hegang; Liu, Yanwen; Yu, Kun; Huang, Bisheng

2016-01-01

The rhizome of Atractylodes lancea is extensively used in the practice of Traditional Chinese Medicine because of its broad pharmacological activities. This study was designed to characterize the transcriptome profiling of the rhizome and leaf of Atractylodes lancea in an attempt to uncover the molecular mechanisms regulating rhizome formation and growth. Over 270 million clean reads were assembled into 92,366 unigenes, 58% of which are homologous with sequences in public protein databases (NR, Swiss-Prot, GO, and KEGG). Analysis of expression levels showed that genes involved in photosynthesis, stress response, and translation were the most abundant transcripts in the leaf, while transcripts involved in stress response, transcription regulation, translation, and metabolism were dominant in the rhizome. Tissue-specific gene analysis identified distinct gene families active in the leaf and rhizome. Differential gene expression analysis revealed a clear difference in gene expression pattern, identifying 1518 up-regulated genes and 3464 down-regulated genes in the rhizome compared with the leaf, including a series of genes related to signal transduction, primary and secondary metabolism. Transcription factor (TF) analysis identified 42 TF families, with 67 and 60 TFs up-regulated in the rhizome and leaf, respectively. A total of 104 unigenes were identified as candidates for regulating rhizome formation and development. These data offer an overview of the gene expression pattern of the rhizome and leaf and provide essential information for future studies on the molecular mechanisms of controlling rhizome formation and growth. The extensive transcriptome data generated in this study will be a valuable resource for further functional genomics studies of A. lancea. PMID:27066021

PIVOT: platform for interactive analysis and visualization of transcriptomics data.

PubMed

Zhu, Qin; Fisher, Stephen A; Dueck, Hannah; Middleton, Sarah; Khaladkar, Mugdha; Kim, Junhyong

2018-01-05

Many R packages have been developed for transcriptome analysis but their use often requires familiarity with R and integrating results of different packages requires scripts to wrangle the datatypes. Furthermore, exploratory data analyses often generate multiple derived datasets such as data subsets or data transformations, which can be difficult to track. Here we present PIVOT, an R-based platform that wraps open source transcriptome analysis packages with a uniform user interface and graphical data management that allows non-programmers to interactively explore transcriptomics data. PIVOT supports more than 40 popular open source packages for transcriptome analysis and provides an extensive set of tools for statistical data manipulations. A graph-based visual interface is used to represent the links between derived datasets, allowing easy tracking of data versions. PIVOT further supports automatic report generation, publication-quality plots, and program/data state saving, such that all analysis can be saved, shared and reproduced. PIVOT will allow researchers with broad background to easily access sophisticated transcriptome analysis tools and interactively explore transcriptome datasets.

Baculovirus induced transcripts in hemocytes from Heliothis virescens

USDA-ARS?s Scientific Manuscript database

Using RNA-sequencing digital difference expression profiling methods we have assessed the gene expression profiles of hemocytes harvested from Heliothis virescens that were challenged with Helicoverpa zea single nucleopolyhedrovirus (HzSNPV). A reference transcriptome of hemocyte-expressed transcri...

Global gene profiling of aging lungs in Atp8b1 mutant mice.

PubMed

Soundararajan, Ramani; Stearns, Timothy M; Czachor, Alexander; Fukumoto, Jutaro; Turn, Christina; Westermann-Clark, Emma; Breitzig, Mason; Tan, Lee; Lockey, Richard F; King, Benjamin L; Kolliputi, Narasaiah

2016-09-29

Recent studies implicate cardiolipin oxidation in several age-related diseases. Atp8b1 encoding Type 4 P-type ATPases is a cardiolipin transporter. Mutation in Atp8b1 gene or inflammation of the lungs impairs the capacity of Atp8b1 to clear cardiolipin from lung fluid. However, the link between Atp8b1 mutation and age-related gene alteration is unknown. Therefore, we investigated how Atp8b1 mutation alters age-related genes. We performed Affymetrix gene profiling of lungs isolated from young (7-9 wks, n=6) and aged (14 months, 14 M, n=6) C57BL/6 and Atp8b1 mutant mice. In addition, Ingenuity Pathway Analysis (IPA) was performed. Differentially expressed genes were validated by quantitative real-time PCR (qRT-PCR). Global transcriptome analysis revealed 532 differentially expressed genes in Atp8b1 lungs, 157 differentially expressed genes in C57BL/6 lungs, and 37 overlapping genes. IPA of age-related genes in Atp8b1 lungs showed enrichment of Xenobiotic metabolism and Nrf2-mediated signaling pathways. The increase in Adamts2 and Mmp13 transcripts in aged Atp8b1 lungs was validated by qRT-PCR. Similarly, the decrease in Col1a1 and increase in Cxcr6 transcripts was confirmed in both Atp8b1 mutant and C57BL/6 lungs. Based on transcriptome profiling, our study indicates that Atp8b1 mutant mice may be susceptible to age-related lung diseases.

Transcriptomic Analysis of Paulownia Infected by Paulownia Witches'-Broom Phytoplasma

PubMed Central

Zhu, Shui-Fang; Lin, Cai-Li; Tian, Guo-Zhong; Xu, Xia; Zhao, Wen-Jun

2013-01-01

Phytoplasmas are plant pathogenic bacteria that have no cell wall and are responsible for major crop losses throughout the world. Phytoplasma-infected plants show a variety of symptoms and the mechanisms they use to physiologically alter the host plants are of considerable interest, but poorly understood. In this study we undertook a detailed analysis of Paulownia infected by Paulownia witches’-broom (PaWB) Phytoplasma using high-throughput mRNA sequencing (RNA-Seq) and digital gene expression (DGE). RNA-Seq analysis identified 74,831 unigenes, which were subsequently used as reference sequences for DGE analysis of diseased and healthy Paulownia in field grown and tissue cultured plants. Our study revealed that dramatic changes occurred in the gene expression profile of Paulownia after PaWB Phytoplasma infection. Genes encoding key enzymes in cytokinin biosynthesis, such as isopentenyl diphosphate isomerase and isopentenyltransferase, were significantly induced in the infected Paulownia. Genes involved in cell wall biosynthesis and degradation were largely up-regulated and genes related to photosynthesis were down-regulated after PaWB Phytoplasma infection. Our systematic analysis provides comprehensive transcriptomic data about plants infected by Phytoplasma. This information will help further our understanding of the detailed interaction mechanisms between plants and Phytoplasma. PMID:24130859

Trinity | Informatics Technology for Cancer Research (ITCR)

Cancer.gov

Trinity Cancer Transcriptome Analysis Toolkit (CTAT) including de novo transcriptome assembly with downstream support for expression analysis and focused analyses on cancer transcriptomes, incorporating mutation and fusion transcript discovery, and single cell analysis.

Improved intra-array and interarray normalization of peptide microarray phosphorylation for phosphorylome and kinome profiling by rational selection of relevant spots

PubMed Central

Scholma, Jetse; Fuhler, Gwenny M.; Joore, Jos; Hulsman, Marc; Schivo, Stefano; List, Alan F.; Reinders, Marcel J. T.; Peppelenbosch, Maikel P.; Post, Janine N.

2016-01-01

Massive parallel analysis using array technology has become the mainstay for analysis of genomes and transcriptomes. Analogously, the predominance of phosphorylation as a regulator of cellular metabolism has fostered the development of peptide arrays of kinase consensus substrates that allow the charting of cellular phosphorylation events (often called kinome profiling). However, whereas the bioinformatical framework for expression array analysis is well-developed, no advanced analysis tools are yet available for kinome profiling. Especially intra-array and interarray normalization of peptide array phosphorylation remain problematic, due to the absence of “housekeeping” kinases and the obvious fallacy of the assumption that different experimental conditions should exhibit equal amounts of kinase activity. Here we describe the development of analysis tools that reliably quantify phosphorylation of peptide arrays and that allow normalization of the signals obtained. We provide a method for intraslide gradient correction and spot quality control. We describe a novel interarray normalization procedure, named repetitive signal enhancement, RSE, which provides a mathematical approach to limit the false negative results occuring with the use of other normalization procedures. Using in silico and biological experiments we show that employing such protocols yields superior insight into cellular physiology as compared to classical analysis tools for kinome profiling. PMID:27225531

Systems analysis of transcriptional data provides insights into muscle's biological response to botulinum toxin.

PubMed

Mukund, Kavitha; Mathewson, Margie; Minamoto, Viviane; Ward, Samuel R; Subramaniam, Shankar; Lieber, Richard L

2014-11-01

This study provides global transcriptomic profiling and analysis of botulinum toxin A (BoNT-A)-treated muscle over a 1-year period. Microarray analysis was performed on rat tibialis anterior muscles from 4 groups (n = 4/group) at 1, 4, 12, and 52 weeks after BoNT-A injection compared with saline-injected rats at 12 weeks. Dramatic transcriptional adaptation occurred at 1 week with a paradoxical increase in expression of slow and immature isoforms, activation of genes in competing pathways of repair and atrophy, impaired mitochondrial biogenesis, and increased metal ion imbalance. Adaptations of the basal lamina and fibrillar extracellular matrix (ECM) occurred by 4 weeks. The muscle transcriptome returned to its unperturbed state 12 weeks after injection. Acute transcriptional adaptations resemble denervated muscle with some subtle differences, but resolved more quickly compared with denervation. Overall, gene expression across time correlates with the generally accepted BoNT-A time course and suggests that the direct action of BoNT-A in skeletal muscle is relatively rapid. © 2014 Wiley Periodicals, Inc.

Microfluidics for genome-wide studies involving next generation sequencing

PubMed Central

Murphy, Travis W.; Lu, Chang

2017-01-01

Next-generation sequencing (NGS) has revolutionized how molecular biology studies are conducted. Its decreasing cost and increasing throughput permit profiling of genomic, transcriptomic, and epigenomic features for a wide range of applications. Microfluidics has been proven to be highly complementary to NGS technology with its unique capabilities for handling small volumes of samples and providing platforms for automation, integration, and multiplexing. In this article, we review recent progress on applying microfluidics to facilitate genome-wide studies. We emphasize on several technical aspects of NGS and how they benefit from coupling with microfluidic technology. We also summarize recent efforts on developing microfluidic technology for genomic, transcriptomic, and epigenomic studies, with emphasis on single cell analysis. We envision rapid growth in these directions, driven by the needs for testing scarce primary cell samples from patients in the context of precision medicine. PMID:28396707

Meta-Analysis of Placental Transcriptome Data Identifies a Novel Molecular Pathway Related to Preeclampsia.

PubMed

van Uitert, Miranda; Moerland, Perry D; Enquobahrie, Daniel A; Laivuori, Hannele; van der Post, Joris A M; Ris-Stalpers, Carrie; Afink, Gijs B

2015-01-01

Studies using the placental transcriptome to identify key molecules relevant for preeclampsia are hampered by a relatively small sample size. In addition, they use a variety of bioinformatics and statistical methods, making comparison of findings challenging. To generate a more robust preeclampsia gene expression signature, we performed a meta-analysis on the original data of 11 placenta RNA microarray experiments, representing 139 normotensive and 116 preeclamptic pregnancies. Microarray data were pre-processed and analyzed using standardized bioinformatics and statistical procedures and the effect sizes were combined using an inverse-variance random-effects model. Interactions between genes in the resulting gene expression signature were identified by pathway analysis (Ingenuity Pathway Analysis, Gene Set Enrichment Analysis, Graphite) and protein-protein associations (STRING). This approach has resulted in a comprehensive list of differentially expressed genes that led to a 388-gene meta-signature of preeclamptic placenta. Pathway analysis highlights the involvement of the previously identified hypoxia/HIF1A pathway in the establishment of the preeclamptic gene expression profile, while analysis of protein interaction networks indicates CREBBP/EP300 as a novel element central to the preeclamptic placental transcriptome. In addition, there is an apparent high incidence of preeclampsia in women carrying a child with a mutation in CREBBP/EP300 (Rubinstein-Taybi Syndrome). The 388-gene preeclampsia meta-signature offers a vital starting point for further studies into the relevance of these genes (in particular CREBBP/EP300) and their concomitant pathways as biomarkers or functional molecules in preeclampsia. This will result in a better understanding of the molecular basis of this disease and opens up the opportunity to develop rational therapies targeting the placental dysfunction causal to preeclampsia.

The Spatial and Temporal Transcriptomic Landscapes of Ginseng, Panax ginseng C. A. Meyer.

PubMed

Wang, Kangyu; Jiang, Shicui; Sun, Chunyu; Lin, Yanping; Yin, Rui; Wang, Yi; Zhang, Meiping

2015-12-11

Ginseng, including Asian ginseng (Panax ginseng C. A. Meyer) and American ginseng (P. quinquefolius L.), is one of the most important medicinal herbs in Asia and North America, but significantly understudied. This study sequenced and characterized the transcriptomes and expression profiles of genes expressed in 14 tissues and four different aged roots of Asian ginseng. A total of 265.2 million 100-bp clean reads were generated using the high-throughput sequencing platform HiSeq 2000, representing >8.3x of the 3.2-Gb ginseng genome. From the sequences, 248,993 unigenes were assembled for whole plant, 61,912-113,456 unigenes for each tissue and 54,444-65,412 unigenes for different year-old roots. We comprehensively analyzed the unigene sets and gene expression profiles. We found that the number of genes allocated to each functional category is stable across tissues or developmental stages, while the expression profiles of different genes of a gene family or involved in ginsenoside biosynthesis dramatically diversified spatially and temporally. These results provide an overall insight into the spatial and temporal transcriptome dynamics and landscapes of Asian ginseng, and comprehensive resources for advanced research and breeding of ginseng and related species.

Single-cell transcriptome conservation in cryopreserved cells and tissues.

PubMed

Guillaumet-Adkins, Amy; Rodríguez-Esteban, Gustavo; Mereu, Elisabetta; Mendez-Lago, Maria; Jaitin, Diego A; Villanueva, Alberto; Vidal, August; Martinez-Marti, Alex; Felip, Enriqueta; Vivancos, Ana; Keren-Shaul, Hadas; Heath, Simon; Gut, Marta; Amit, Ido; Gut, Ivo; Heyn, Holger

2017-03-01

A variety of single-cell RNA preparation procedures have been described. So far, protocols require fresh material, which hinders complex study designs. We describe a sample preservation method that maintains transcripts in viable single cells, allowing one to disconnect time and place of sampling from subsequent processing steps. We sequence single-cell transcriptomes from >1000 fresh and cryopreserved cells using 3'-end and full-length RNA preparation methods. Our results confirm that the conservation process did not alter transcriptional profiles. This substantially broadens the scope of applications in single-cell transcriptomics and could lead to a paradigm shift in future study designs.

Transcription profile analysis of Lycopersicum esculentum leaves, unravels volatile emissions and gene expression under salinity stress.

PubMed

Zhang, Jihong; Zeng, Li; Chen, Shaoyang; Sun, Helong; Ma, Shuang

2018-05-01

Salinity stress can impede development and plant growth adversely. However, there is very little molecular information on NaCl resistance and volatile emissions in Lycopersicum esculentum. In order to investigate the effects of salt stress on the release of volatile compounds, we quantified and compared transcriptome changes by RNA-Seq analysis and volatile constituents with gas chromatography/mass spectrometry (GC/MS) coupled with solid-phase microextraction (SPME) after exposure to continuous salt stress. Chemical analysis by GC-MS analysis revealed that NaCl stress had changed species and quantity of volatile compounds released. In this research, 21,578 unigenes that represented 44,714 assembled unique transcripts were separated from tomato leaves exposed to NaCl stress based on de novo transcriptome assembly. The total number of differentially expressed genes was 7210 after exposure to NaCl, including 6200 down-regulated and 1208 up-regulated genes. Among these differentially expressed genes (DEGs), there were eighteen differentially expressed genes associated with volatile biosynthesis. Of the unigenes, 3454 were mapped to 131 KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways, mainly those are involved in RNA transport, plant-pathogen interactions, and plant hormone signal transduction. qRT-PCR analysis showed that NaCl exposure affected the expression profiles of the biosynthesis genes for eight volatile compounds (IPI, GPS, and TPS, etc.), which corresponded well with the RNA-Seq analysis and GC-MS results. Our results suggest that NaCl stress affects the emission of volatile substances from L. esculentum leaves by regulating the expression of genes that are involved in volatile organic compounds' biosynthesis. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Transcriptomic profiles of human foreskin fibroblast cells in response to orf virus.

PubMed

Chen, Daxiang; Long, Mingjian; Xiao, Bin; Xiong, Yufeng; Chen, Huiqin; Chen, Yu; Kuang, Zhenzhan; Li, Ming; Wu, Yingsong; Rock, Daniel L; Gong, Daoyuan; Wang, Yong; He, Haijian; Liu, Fang; Luo, Shuhong; Hao, Wenbo

2017-08-29

Orf virus has been utilized as a safe and efficient viral vector against not only diverse infectious diseases, but also against tumors. However, the nature of the genes triggered by the vector in human cells is poorly characterized. Using RNA sequencing technology, we compared specific changes in the transcriptomic profiles in human foreskin fibroblast cells following infection by the orf virus. The results indicated that orf virus upregulates or downregulates expression of a variety of genes, including genes involved in antiviral immune response, apoptosis, cell cycle and a series of signaling pathways, such as the IFN and p53-signaling pathways. The orf virus stimulates or inhibits immune gene expression such as chemokines, chemokine receptors, cytokines, cytokine receptors, and molecules involved in antigen uptake and processing after infection. Expression of pro-apoptotic genes increased at 8 hours post-infection. The p53 signaling pathway was activated to induce apoptosis at the same time. However, the cell cycle program was promoted after infection, which may be due to the immunomodulatory genes of the orf virus. This presents the first description of transcription profile changes in human foreskin fibroblast cells after orf virus infection and provides an in-depth analysis of the interaction between the host and orf virus. These data offer new insights into the understanding of the mechanisms of infection by orf virus and identify potential targets for future studies.

RNA-seq Analysis Reveals Unique Transcriptome Signatures in Systemic Lupus Erythematosus Patients with Distinct Autoantibody Specificities

PubMed Central

Rai, Richa; Chauhan, Sudhir Kumar; Singh, Vikas Vikram; Rai, Madhukar; Rai, Geeta

2016-01-01

Systemic lupus erythematosus (SLE) patients exhibit immense heterogeneity which is challenging from the diagnostic perspective. Emerging high throughput sequencing technologies have been proved to be a useful platform to understand the complex and dynamic disease processes. SLE patients categorised based on autoantibody specificities are reported to have differential immuno-regulatory mechanisms. Therefore, we performed RNA-seq analysis to identify transcriptomics of SLE patients with distinguished autoantibody specificities. The SLE patients were segregated into three subsets based on the type of autoantibodies present in their sera (anti-dsDNA+ group with anti-dsDNA autoantibody alone; anti-ENA+ group having autoantibodies against extractable nuclear antigens (ENA) only, and anti-dsDNA+ENA+ group having autoantibodies to both dsDNA and ENA). Global transcriptome profiling for each SLE patients subsets was performed using Illumina® Hiseq-2000 platform. The biological relevance of dysregulated transcripts in each SLE subsets was assessed by ingenuity pathway analysis (IPA) software. We observed that dysregulation in the transcriptome expression pattern was clearly distinct in each SLE patients subsets. IPA analysis of transcripts uniquely expressed in different SLE groups revealed specific biological pathways to be affected in each SLE subsets. Multiple cytokine signaling pathways were specifically dysregulated in anti-dsDNA+ patients whereas Interferon signaling was predominantly dysregulated in anti-ENA+ patients. In anti-dsDNA+ENA+ patients regulation of actin based motility by Rho pathway was significantly affected. The granulocyte gene signature was a common feature to all SLE subsets; however, anti-dsDNA+ group showed relatively predominant expression of these genes. Dysregulation of Plasma cell related transcripts were higher in anti-dsDNA+ and anti-ENA+ patients as compared to anti-dsDNA+ ENA+. Association of specific canonical pathways with the uniquely expressed transcripts in each SLE subgroup indicates that specific immunological disease mechanisms are operative in distinct SLE patients’ subsets. This ‘sub-grouping’ approach could further be useful for clinical evaluation of SLE patients and devising targeted therapeutics. PMID:27835693

Transcriptomic Analysis of Paeonia delavayi Wild Population Flowers to Identify Differentially Expressed Genes Involved in Purple-Red and Yellow Petal Pigmentation

PubMed Central

Wang, Yan; Li, Kui; Zheng, Baoqiang; Miao, Kun

2015-01-01

Tree peony (Paeonia suffruticosa Andrews) is a very famous traditional ornamental plant in China. P. delavayi is a species endemic to Southwest China that has aroused great interest from researchers as a precious genetic resource for flower color breeding. However, the current understanding of the molecular mechanisms of flower pigmentation in this plant is limited, hindering the genetic engineering of novel flower color in tree peonies. In this study, we conducted a large-scale transcriptome analysis based on Illumina HiSeq sequencing of cDNA libraries generated from yellow and purple-red P. delavayi petals. A total of 90,202 unigenes were obtained by de novo assembly, with an average length of 721 nt. Using Blastx, 44,811 unigenes (49.68%) were found to have significant similarity to accessions in the NR, NT, and Swiss-Prot databases. We also examined COG, GO and KEGG annotations to better understand the functions of these unigenes. Further analysis of the two digital transcriptomes revealed that 6,855 unigenes were differentially expressed between yellow and purple-red flower petals, with 3,430 up-regulated and 3,425 down-regulated. According to the RNA-Seq data and qRT-PCR analysis, we proposed that four up-regulated key structural genes, including F3H, DFR, ANS and 3GT, might play an important role in purple-red petal pigmentation, while high co-expression of THC2'GT, CHI and FNS II ensures the accumulation of pigments contributing to the yellow color. We also found 50 differentially expressed transcription factors that might be involved in flavonoid biosynthesis. This study is the first to report genetic information for P. delavayi. The large number of gene sequences produced by transcriptome sequencing and the candidate genes identified using pathway mapping and expression profiles will provide a valuable resource for future association studies aimed at better understanding the molecular mechanisms underlying flower pigmentation in tree peonies. PMID:26267644

Tissue-Specific Transcriptome Profiling of Plutella Xylostella Third Instar Larval Midgut

PubMed Central

Xie, Wen; Lei, Yanyuan; Fu, Wei; Yang, Zhongxia; Zhu, Xun; Guo, Zhaojiang; Wu, Qingjun; Wang, Shaoli; Xu, Baoyun; Zhou, Xuguo; Zhang, Youjun

2012-01-01

The larval midgut of diamondback moth, Plutella xylostella, is a dynamic tissue that interfaces with a diverse array of physiological and toxicological processes, including nutrient digestion and allocation, xenobiotic detoxification, innate and adaptive immune response, and pathogen defense. Despite its enormous agricultural importance, the genomic resources for P. xylostella are surprisingly scarce. In this study, a Bt resistant P. xylostella strain was subjected to the in-depth transcriptome analysis to identify genes and gene networks putatively involved in various physiological and toxicological processes in the P. xylostella larval midgut. Using Illumina deep sequencing, we obtained roughly 40 million reads containing approximately 3.6 gigabases of sequence data. De novo assembly generated 63,312 ESTs with an average read length of 416bp, and approximately half of the P. xylostella sequences (45.4%, 28,768) showed similarity to the non-redundant database in GenBank with a cut-off E-value below 10-5. Among them, 11,092 unigenes were assigned to one or multiple GO terms and 16,732 unigenes were assigned to 226 specific pathways. In-depth analysis indentified genes putatively involved in insecticide resistance, nutrient digestion, and innate immune defense. Besides conventional detoxification enzymes and insecticide targets, novel genes, including 28 chymotrypsins and 53 ABC transporters, have been uncovered in the P. xylostella larval midgut transcriptome; which are potentially linked to the Bt toxicity and resistance. Furthermore, an unexpectedly high number of ESTs, including 46 serpins and 7 lysozymes, were predicted to be involved in the immune defense. As the first tissue-specific transcriptome analysis of P. xylostella, this study sheds light on the molecular understanding of insecticide resistance, especially Bt resistance in an agriculturally important insect pest, and lays the foundation for future functional genomics research. In addition, current sequencing effort greatly enriched the existing P. xylostella EST database, and makes RNAseq a viable option in the future genomic analysis. PMID:23091412

Tissue-specific transcriptome profiling of Plutella xylostella third instar larval midgut.

PubMed

Xie, Wen; Lei, Yanyuan; Fu, Wei; Yang, Zhongxia; Zhu, Xun; Guo, Zhaojiang; Wu, Qingjun; Wang, Shaoli; Xu, Baoyun; Zhou, Xuguo; Zhang, Youjun

2012-01-01

The larval midgut of diamondback moth, Plutella xylostella, is a dynamic tissue that interfaces with a diverse array of physiological and toxicological processes, including nutrient digestion and allocation, xenobiotic detoxification, innate and adaptive immune response, and pathogen defense. Despite its enormous agricultural importance, the genomic resources for P. xylostella are surprisingly scarce. In this study, a Bt resistant P. xylostella strain was subjected to the in-depth transcriptome analysis to identify genes and gene networks putatively involved in various physiological and toxicological processes in the P. xylostella larval midgut. Using Illumina deep sequencing, we obtained roughly 40 million reads containing approximately 3.6 gigabases of sequence data. De novo assembly generated 63,312 ESTs with an average read length of 416 bp, and approximately half of the P. xylostella sequences (45.4%, 28,768) showed similarity to the non-redundant database in GenBank with a cut-off E-value below 10(-5). Among them, 11,092 unigenes were assigned to one or multiple GO terms and 16,732 unigenes were assigned to 226 specific pathways. In-depth analysis identified genes putatively involved in insecticide resistance, nutrient digestion, and innate immune defense. Besides conventional detoxification enzymes and insecticide targets, novel genes, including 28 chymotrypsins and 53 ABC transporters, have been uncovered in the P. xylostella larval midgut transcriptome; which are potentially linked to the Bt toxicity and resistance. Furthermore, an unexpectedly high number of ESTs, including 46 serpins and 7 lysozymes, were predicted to be involved in the immune defense.As the first tissue-specific transcriptome analysis of P. xylostella, this study sheds light on the molecular understanding of insecticide resistance, especially Bt resistance in an agriculturally important insect pest, and lays the foundation for future functional genomics research. In addition, current sequencing effort greatly enriched the existing P. xylostella EST database, and makes RNAseq a viable option in the future genomic analysis.

Comparative description of ten transcriptomes of newly sequenced invertebrates and efficiency estimation of genomic sampling in non-model taxa

PubMed Central

2012-01-01

Introduction Traditionally, genomic or transcriptomic data have been restricted to a few model or emerging model organisms, and to a handful of species of medical and/or environmental importance. Next-generation sequencing techniques have the capability of yielding massive amounts of gene sequence data for virtually any species at a modest cost. Here we provide a comparative analysis of de novo assembled transcriptomic data for ten non-model species of previously understudied animal taxa. Results cDNA libraries of ten species belonging to five animal phyla (2 Annelida [including Sipuncula], 2 Arthropoda, 2 Mollusca, 2 Nemertea, and 2 Porifera) were sequenced in different batches with an Illumina Genome Analyzer II (read length 100 or 150 bp), rendering between ca. 25 and 52 million reads per species. Read thinning, trimming, and de novo assembly were performed under different parameters to optimize output. Between 67,423 and 207,559 contigs were obtained across the ten species, post-optimization. Of those, 9,069 to 25,681 contigs retrieved blast hits against the NCBI non-redundant database, and approximately 50% of these were assigned with Gene Ontology terms, covering all major categories, and with similar percentages in all species. Local blasts against our datasets, using selected genes from major signaling pathways and housekeeping genes, revealed high efficiency in gene recovery compared to available genomes of closely related species. Intriguingly, our transcriptomic datasets detected multiple paralogues in all phyla and in nearly all gene pathways, including housekeeping genes that are traditionally used in phylogenetic applications for their purported single-copy nature. Conclusions We generated the first study of comparative transcriptomics across multiple animal phyla (comparing two species per phylum in most cases), established the first Illumina-based transcriptomic datasets for sponge, nemertean, and sipunculan species, and generated a tractable catalogue of annotated genes (or gene fragments) and protein families for ten newly sequenced non-model organisms, some of commercial importance (i.e., Octopus vulgaris). These comprehensive sets of genes can be readily used for phylogenetic analysis, gene expression profiling, developmental analysis, and can also be a powerful resource for gene discovery. The characterization of the transcriptomes of such a diverse array of animal species permitted the comparison of sequencing depth, functional annotation, and efficiency of genomic sampling using the same pipelines, which proved to be similar for all considered species. In addition, the datasets revealed their potential as a resource for paralogue detection, a recurrent concern in various aspects of biological inquiry, including phylogenetics, molecular evolution, development, and cellular biochemistry. PMID:23190771

Analysis of ripening-related gene expression in papaya using an Arabidopsis-based microarray

PubMed Central

2012-01-01

Background Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals. Despite its importance, little is known about transcriptional modifications during papaya fruit ripening and their control. In this study we report the analysis of ripe papaya transcriptome by using a cross-species (XSpecies) microarray technique based on the phylogenetic proximity between papaya and Arabidopsis thaliana. Results Papaya transcriptome analyses resulted in the identification of 414 ripening-related genes with some having their expression validated by qPCR. The transcription profile was compared with that from ripening tomato and grape. There were many similarities between papaya and tomato especially with respect to the expression of genes encoding proteins involved in primary metabolism, regulation of transcription, biotic and abiotic stress and cell wall metabolism. XSpecies microarray data indicated that transcription factors (TFs) of the MADS-box, NAC and AP2/ERF gene families were involved in the control of papaya ripening and revealed that cell wall-related gene expression in papaya had similarities to the expression profiles seen in Arabidopsis during hypocotyl development. Conclusion The cross-species array experiment identified a ripening-related set of genes in papaya allowing the comparison of transcription control between papaya and other fruit bearing taxa during the ripening process. PMID:23256600

Transcriptome and Differential Expression Profiling Analysis of the Mechanism of Ca2+ Regulation in Peanut (Arachis hypogaea) Pod Development

PubMed Central

Yang, Sha; Li, Lin; Zhang, Jialei; Geng, Yun; Guo, Feng; Wang, Jianguo; Meng, Jingjing; Sui, Na; Wan, Shubo; Li, Xinguo

2017-01-01

Calcium not only serves as a necessary nutrient for plant growth but also acts as a ubiquitous central hub in a large number of signaling pathways. Free Ca2+ deficiency in the soil may cause early embryo abortion, which eventually led to abnormal development of peanut pod during the harvest season. To understand the mechanisms of Ca2+ regulation in pod development, transcriptome analysis of peanut gynophores and pods was performed by comparing the treatments between free Ca2+ sufficiency and free Ca2+ deficiency using Illumina HiSeq™ 2000. 9,903,082,800 nt bases are generated totally. After assembly, the average length of 102,819 unigenes is 999 nt, N50 is 1,782 nt. RNA-seq based gene expression profilings showed a large number of genes at the transcriptional level changed significantly between the aerial pegs and underground swelling pods under free Ca2+ sufficienct or deficiency treatments, respectively. Genes encoding key members of Ca2+ signaling transduction pathway, enzymes for hormone metabolism, cell division and growth, transcriptional factor as well as embryo development were highlighted. This information provides useful information for our further study. The results of digital gene expression (DGE) indicated that exogenous calcium might contribute to the development of peanut pod through its signal transduction pathway, meanwhile, promote the normal transition of the gynophores to the reproductive development. PMID:29033956

Differential expression of lncRNAs during the HIV replication cycle: an underestimated layer in the HIV-host interplay.

PubMed

Trypsteen, Wim; Mohammadi, Pejman; Van Hecke, Clarissa; Mestdagh, Pieter; Lefever, Steve; Saeys, Yvan; De Bleser, Pieter; Vandesompele, Jo; Ciuffi, Angela; Vandekerckhove, Linos; De Spiegelaere, Ward

2016-10-26

Studying the effects of HIV infection on the host transcriptome has typically focused on protein-coding genes. However, recent advances in the field of RNA sequencing revealed that long non-coding RNAs (lncRNAs) add an extensive additional layer to the cell's molecular network. Here, we performed transcriptome profiling throughout a primary HIV infection in vitro to investigate lncRNA expression at the different HIV replication cycle processes (reverse transcription, integration and particle production). Subsequently, guilt-by-association, transcription factor and co-expression analysis were performed to infer biological roles for the lncRNAs identified in the HIV-host interplay. Many lncRNAs were suggested to play a role in mechanisms relying on proteasomal and ubiquitination pathways, apoptosis, DNA damage responses and cell cycle regulation. Through transcription factor binding analysis, we found that lncRNAs display a distinct transcriptional regulation profile as compared to protein coding mRNAs, suggesting that mRNAs and lncRNAs are independently modulated. In addition, we identified five differentially expressed lncRNA-mRNA pairs with mRNA involvement in HIV pathogenesis with possible cis regulatory lncRNAs that control nearby mRNA expression and function. Altogether, the present study demonstrates that lncRNAs add a new dimension to the HIV-host interplay and should be further investigated as they may represent targets for controlling HIV replication.

Transcriptome architecture across tissues in the pig

PubMed Central

Ferraz, André LJ; Ojeda, Ana; López-Béjar, Manel; Fernandes, Lana T; Castelló, Anna; Folch, Josep M; Pérez-Enciso, Miguel

2008-01-01

Background Artificial selection has resulted in animal breeds with extreme phenotypes. As an organism is made up of many different tissues and organs, each with its own genetic programme, it is pertinent to ask: How relevant is tissue in terms of total transcriptome variability? Which are the genes most distinctly expressed between tissues? Does breed or sex equally affect the transcriptome across tissues? Results In order to gain insight on these issues, we conducted microarray expression profiling of 16 different tissues from four animals of two extreme pig breeds, Large White and Iberian, two males and two females. Mixed model analysis and neighbor – joining trees showed that tissues with similar developmental origin clustered closer than those with different embryonic origins. Often a sound biological interpretation was possible for overrepresented gene ontology categories within differentially expressed genes between groups of tissues. For instance, an excess of nervous system or muscle development genes were found among tissues of ectoderm or mesoderm origins, respectively. Tissue accounted for ~11 times more variability than sex or breed. Nevertheless, we were able to confidently identify genes with differential expression across tissues between breeds (33 genes) and between sexes (19 genes). The genes primarily affected by sex were overall different than those affected by breed or tissue. Interaction with tissue can be important for differentially expressed genes between breeds but not so much for genes whose expression differ between sexes. Conclusion Embryonic development leaves an enduring footprint on the transcriptome. The interaction in gene × tissue for differentially expressed genes between breeds suggests that animal breeding has targeted differentially each tissue's transcriptome. PMID:18416811

A comparative transcriptomic approach to understanding the formation of cork.

PubMed

Boher, Pau; Soler, Marçal; Sánchez, Anna; Hoede, Claire; Noirot, Céline; Paiva, Jorge Almiro Pinto; Serra, Olga; Figueras, Mercè

2018-01-01

The transcriptome comparison of two oak species reveals possible candidates accounting for the exceptionally thick and pure cork oak phellem, such as those involved in secondary metabolism and phellogen activity. Cork oak, Quercus suber, differs from other Mediterranean oaks such as holm oak (Quercus ilex) by the thickness and organization of the external bark. While holm oak outer bark contains sequential periderms interspersed with dead secondary phloem (rhytidome), the cork oak outer bark only contains thick layers of phellem (cork rings) that accumulate until reaching a thickness that allows industrial uses. Here we compare the cork oak outer bark transcriptome with that of holm oak. Both transcriptomes present similitudes in their complexity, but whereas cork oak external bark is enriched with upregulated genes related to suberin, which is the main polymer responsible for the protective function of periderm, the upregulated categories of holm oak are enriched in abiotic stress and chromatin assembly. Concomitantly with the upregulation of suberin-related genes, there is also induction of regulatory and meristematic genes, whose predicted activities agree with the increased number of phellem layers found in the cork oak sample. Further transcript profiling among different cork oak tissues and conditions suggests that cork and wood share many regulatory mechanisms, probably reflecting similar ontogeny. Moreover, the analysis of transcripts accumulation during the cork growth season showed that most regulatory genes are upregulated early in the season when the cork cambium becomes active. Altogether our work provides the first transcriptome comparison between cork oak and holm oak outer bark, which unveils new regulatory candidate genes of phellem development.

A draft of the genome and four transcriptomes of a medicinal and pesticidal angiosperm Azadirachta indica

PubMed Central

2012-01-01

Background The Azadirachta indica (neem) tree is a source of a wide number of natural products, including the potent biopesticide azadirachtin. In spite of its widespread applications in agriculture and medicine, the molecular aspects of the biosynthesis of neem terpenoids remain largely unexplored. The current report describes the draft genome and four transcriptomes of A. indica and attempts to contextualise the sequence information in terms of its molecular phylogeny, transcript expression and terpenoid biosynthesis pathways. A. indica is the first member of the family Meliaceae to be sequenced using next generation sequencing approach. Results The genome and transcriptomes of A. indica were sequenced using multiple sequencing platforms and libraries. The A. indica genome is AT-rich, bears few repetitive DNA elements and comprises about 20,000 genes. The molecular phylogenetic analyses grouped A. indica together with Citrus sinensis from the Rutaceae family validating its conventional taxonomic classification. Comparative transcript expression analysis showed either exclusive or enhanced expression of known genes involved in neem terpenoid biosynthesis pathways compared to other sequenced angiosperms. Genome and transcriptome analyses in A. indica led to the identification of repeat elements, nucleotide composition and expression profiles of genes in various organs. Conclusions This study on A. indica genome and transcriptomes will provide a model for characterization of metabolic pathways involved in synthesis of bioactive compounds, comparative evolutionary studies among various Meliaceae family members and help annotate their genomes. A better understanding of molecular pathways involved in the azadirachtin synthesis in A. indica will pave ways for bulk production of environment friendly biopesticides. PMID:22958331

De novo transcriptome assembly of the calanoid copepod Neocalanus flemingeri: A new resource for emergence from diapause.

PubMed

Roncalli, Vittoria; Cieslak, Matthew C; Sommer, Stephanie A; Hopcroft, Russell R; Lenz, Petra H

2018-02-01

Copepods, small planktonic crustaceans, are key links between primary producers and upper trophic levels, including many economically important fishes. In the subarctic North Pacific, the life cycle of copepods like Neocalanus flemingeri includes an ontogenetic migration to depth followed by a period of diapause (a type of dormancy) characterized by arrested development and low metabolic activity. The end of diapause is marked by the production of the first brood of eggs. Recent temperature anomalies in the North Pacific have raised concerns about potential negative effects on N. flemingeri. Since diapause is a developmental program, its progress can be tracked using through global gene expression. Thus, a reference transcriptome was developed as a first step towards physiological profiling of diapausing females using high-throughput Illumina sequencing. The de novo transcriptome, the first for this species was designed to investigate the diapause period. RNA-Seq reads were obtained for dormant to reproductive N. flemingeri females. A high quality de novo transcriptome was obtained by first assembling reads from each individual using Trinity software followed by clustering with CAP3 Assembly Program. This assembly consisted of 140,841transcripts (contigs). Bench-marking universal single-copy orthologs analysis identified 85% of core eukaryotic genes, with 79% predicted to be complete. Comparison with other calanoid transcriptomes confirmed its quality and degree of completeness. Trinity assembly of reads originating from multiple individuals led to fragmentation. Thus, the workflow applied here differed from the one recommended by Trinity, but was required to obtain a good assembly. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

In Silico Comparative Transcriptome Analysis of Two Color Morphs of the Common Coral Trout (Plectropomus Leopardus)

PubMed Central

Wang, Le; Yu, Cuiping; Guo, Liang; Lin, Haoran; Meng, Zining

2015-01-01

The common coral trout is one species of major importance in commercial fisheries and aquaculture. Recently, two different color morphs of Plectropomus leopardus were discovered and the biological importance of the color difference is unknown. Since coral trout species are poorly characterized at the molecular level, we undertook the transcriptomic characterization of the two color morphs, one black and one red coral trout, using Illumina next generation sequencing technologies. The study produced 55162966 and 54588952 paired-end reads, for black and red trout, respectively. De novo transcriptome assembly generated 95367 and 99424 unique sequences in black and red trout, respectively, with 88813 sequences shared between them. Approximately 50% of both trancriptomes were functionally annotated by BLAST searches against protein databases. The two trancriptomes were enriched into 25 functional categories and showed similar profiles of Gene Ontology category compositions. 34110 unigenes were grouped into 259 KEGG pathways. Moreover, we identified 14649 simple sequence repeats (SSRs) and designed primers for potential application. We also discovered 130524 putative single nucleotide polymorphisms (SNPs) in the two transcriptomes, supplying potential genomic resources for the coral trout species. In addition, we identified 936 fast-evolving genes and 165 candidate genes under positive selection between the two color morphs. Finally, 38 candidate genes underlying the mechanism of color and pigmentation were also isolated. This study presents the first transcriptome resources for the common coral trout and provides basic information for the development of genomic tools for the identification, conservation, and understanding of the speciation and local adaptation of coral reef fish species. PMID:26713756

Plant body weight-induced secondary growth in Arabidopsis and its transcription phenotype revealed by whole-transcriptome profiling.

PubMed

Ko, Jae-Heung; Han, Kyung-Hwan; Park, Sunchung; Yang, Jaemo

2004-06-01

Wood is an important raw material and environmentally cost-effective renewable source of energy. However, the molecular biology of wood formation (i.e. secondary growth) is surprisingly understudied. A novel experimental system was employed to study the molecular regulation of secondary xylem formation in Arabidopsis. First, we demonstrate that the weight carried by the stem is a primary signal for the induction of cambium differentiation and the plant hormone, auxin, is a downstream carrier of the signal for this process. We used Arabidopsis whole-transcriptome (23 K) GeneChip analysis to examine gene expression profile changes in the inflorescent stems treated for wood formation by cultural manipulation or artificial weight application. Many of the genes up-regulated in wood-forming stems had auxin responsive cis-acting elements in their promoter region, indicating auxin-mediated regulation of secondary growth. We identified 700 genes that were differentially expressed during the transition from primary growth to secondary growth. More than 40% of the genes that were up-regulated (>5x) were associated with signal transduction and transcriptional regulation. Biological significance of these regulatory genes is discussed in light of the induction and development of secondary xylem.

Multivariate inference of pathway activity in host immunity and response to therapeutics

PubMed Central

Goel, Gautam; Conway, Kara L.; Jaeger, Martin; Netea, Mihai G.; Xavier, Ramnik J.

2014-01-01

Developing a quantitative view of how biological pathways are regulated in response to environmental factors is central for understanding of disease phenotypes. We present a computational framework, named Multivariate Inference of Pathway Activity (MIPA), which quantifies degree of activity induced in a biological pathway by computing five distinct measures from transcriptomic profiles of its member genes. Statistical significance of inferred activity is examined using multiple independent self-contained tests followed by a competitive analysis. The method incorporates a new algorithm to identify a subset of genes that may regulate the extent of activity induced in a pathway. We present an in-depth evaluation of specificity, robustness, and reproducibility of our method. We benchmarked MIPA's false positive rate at less than 1%. Using transcriptomic profiles representing distinct physiological and disease states, we illustrate applicability of our method in (i) identifying gene–gene interactions in autophagy-dependent response to Salmonella infection, (ii) uncovering gene–environment interactions in host response to bacterial and viral pathogens and (iii) identifying driver genes and processes that contribute to wound healing and response to anti-TNFα therapy. We provide relevant experimental validation that corroborates the accuracy and advantage of our method. PMID:25147207

Transcriptome profiling of sleeping, waking, and sleep deprived adult heterozygous Aldh1L1 - eGFP-L10a mice.

PubMed

Bellesi, Michele; de Vivo, Luisa; Tononi, Giulio; Cirelli, Chiara

2015-12-01

Transcriptomic studies revealed that hundreds of mRNAs show differential expression in the brains of sleeping relative to awake rats, mice, flies, and sparrows. Although these results have offered clues regarding the molecular consequences of sleep and sleep loss, their functional significance thus far has been limited. This is probably because the previous studies pooled transcripts from all brain cells, including neurons and glia. In Bellesi et al. 2015 [1], we used the translating ribosome affinity purification technology (TRAP) and microarray analysis to obtain a genome-wide mRNA profiling of astrocytes as a function of sleep and wake. We used bacterial artificial chromosome (BAC) transgenic mice expressing eGFP tagged ribosomal protein L10a under the promoter of the Aldh1L1 gene, a highly expressed astrocytic gene. Using this approach, we could extract only the astrocytic mRNAs, and only those already committed to be translated into proteins (L10a is part of the translational machinery). Here, we report a detailed description of the protocol used in the study [1]. Array data have been submitted to NCBI GEO under accession number (GSE69079).

Quantitative radiomic profiling of glioblastoma represents transcriptomic expression.

PubMed

Kong, Doo-Sik; Kim, Junhyung; Ryu, Gyuha; You, Hye-Jin; Sung, Joon Kyung; Han, Yong Hee; Shin, Hye-Mi; Lee, In-Hee; Kim, Sung-Tae; Park, Chul-Kee; Choi, Seung Hong; Choi, Jeong Won; Seol, Ho Jun; Lee, Jung-Il; Nam, Do-Hyun

2018-01-19

Quantitative imaging biomarkers have increasingly emerged in the field of research utilizing available imaging modalities. We aimed to identify good surrogate radiomic features that can represent genetic changes of tumors, thereby establishing noninvasive means for predicting treatment outcome. From May 2012 to June 2014, we retrospectively identified 65 patients with treatment-naïve glioblastoma with available clinical information from the Samsung Medical Center data registry. Preoperative MR imaging data were obtained for all 65 patients with primary glioblastoma. A total of 82 imaging features including first-order statistics, volume, and size features, were semi-automatically extracted from structural and physiologic images such as apparent diffusion coefficient and perfusion images. Using commercially available software, NordicICE, we performed quantitative imaging analysis and collected the dataset composed of radiophenotypic parameters. Unsupervised clustering methods revealed that the radiophenotypic dataset was composed of three clusters. Each cluster represented a distinct molecular classification of glioblastoma; classical type, proneural and neural types, and mesenchymal type. These clusters also reflected differential clinical outcomes. We found that extracted imaging signatures does not represent copy number variation and somatic mutation. Quantitative radiomic features provide a potential evidence to predict molecular phenotype and treatment outcome. Radiomic profiles represents transcriptomic phenotypes more well.

Comparative transcriptomics of Atlantic Salmo salar, chum Oncorhynchus keta and pink salmon O. gorbuscha during infections with salmon lice Lepeophtheirus salmonis.

PubMed

Sutherland, Ben J G; Koczka, Kim W; Yasuike, Motoshige; Jantzen, Stuart G; Yazawa, Ryosuke; Koop, Ben F; Jones, Simon R M

2014-03-15

Salmon species vary in susceptibility to infections with the salmon louse (Lepeophtheirus salmonis). Comparing mechanisms underlying responses in susceptible and resistant species is important for estimating impacts of infections on wild salmon, selective breeding of farmed salmon, and expanding our knowledge of fish immune responses to ectoparasites. Herein we report three L. salmonis experimental infection trials of co-habited Atlantic Salmo salar, chum Oncorhynchus keta and pink salmon O. gorbuscha, profiling hematocrit, blood cortisol concentrations, and transcriptomic responses of the anterior kidney and skin to the infection. In all trials, infection densities (lice per host weight (g)) were consistently highest on chum salmon, followed by Atlantic salmon, and lowest in pink salmon. At 43 days post-exposure, all lice had developed to motile stages, and infection density was uniformly low among species. Hematocrit was reduced in infected Atlantic and chum salmon, and cortisol was elevated in infected chum salmon. Systemic transcriptomic responses were profiled in all species and large differences in response functions were identified between Atlantic and Pacific (chum and pink) salmon. Pink and chum salmon up-regulated acute phase response genes, including complement and coagulation components, and down-regulated antiviral immune genes. The pink salmon response involved the largest and most diverse iron sequestration and homeostasis mechanisms. Pattern recognition receptors were up-regulated in all species but the active components were often species-specific. C-type lectin domain family 4 member M and acidic mammalian chitinase were specifically up-regulated in the resistant pink salmon. Experimental exposures consistently indicated increased susceptibility in chum and Atlantic salmon, and resistance in pink salmon, with differences in infection density occurring within the first three days of infection. Transcriptomic analysis suggested candidate resistance functions including local inflammation with cytokines, specific innate pattern recognition receptors, and iron homeostasis. Suppressed antiviral immunity in both susceptible and resistant species indicates the importance of future work investigating co-infections of viral pathogens and lice.

Complementary transcriptome and proteome profiling in cabbage buds of a recessive male sterile mutant provides new insights into male reproductive development.

PubMed

Ji, Jialei; Yang, Limei; Fang, Zhiyuan; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao; Liu, Yumei; Li, Zhansheng

2018-05-15

Plant male reproductive development is a very complex biological process that involves multiple metabolic pathways. To reveal novel insights into male reproductive development, we conducted an integrated profiling of gene activity in the developing buds of a cabbage recessive genetic male sterile mutant. Using RNA-Seq and label-free quantitative proteomics, 2881 transcripts and 1245 protein species were identified with significant differential abundance between the male sterile line 83121A and its isogenic maintainer line 83121B. Analyses of function annotations and correlations between transcriptome and proteome and protein interaction networks were also conducted, which suggested that the male sterility involves a complex regulatory pattern. Moreover, several key biological processes, such as fatty acid metabolism, tapetosome biosynthesis, amino acid metabolism and protein synthesis and degradation were identified as being of relevance to male reproductive development. A large number of protein species involved in sporopollenin synthesis, amino acid synthesis, ribosome assembly, protein processing in endoplasmic reticulum and lipid transfer were observed to be significantly down-accumulated in 83121A buds, indicating their potential roles in the regulation of cabbage microspore abortion. In summary, the conjoint analysis of the transcriptome and proteome provided a global picture regarding the molecular dynamics in male sterile buds of 83121A. Male sterile mutants are excellent materials for the study of plant male reproductive development. This study revealed the molecular dynamics of recessive male sterility in cabbage at the transcriptome and proteome levels, which deepens our understanding of the metabolic pathways involved in male development. Moreover, the male sterility-related genes identified in this study could provide a reference for the artificial regulation of cabbage fertility by using genetic engineering technology, which may result in potential applications in agriculture such as production of hybrid seeds using male sterility. Copyright © 2018 Elsevier B.V. All rights reserved.

Transcriptome profiling of two olive cultivars in response to infection by the CoDiRO strain of Xylella fastidiosa subsp. pauca.

PubMed

Giampetruzzi, Annalisa; Morelli, Massimiliano; Saponari, Maria; Loconsole, Giuliana; Chiumenti, Michela; Boscia, Donato; Savino, Vito N; Martelli, Giovanni P; Saldarelli, Pasquale

2016-06-27

The recent Xylella fastidiosa subsp. pauca (Xfp) outbreak in olive (Olea europaea) groves in southern Italy is causing a destructive disease denoted Olive Quick Decline Syndrome (OQDS). Field observations disclosed that Xfp-infected plants of cv. Leccino show much milder symptoms, than the more widely grown and highly susceptible cv. Ogliarola salentina. To determine whether these field observations underlie a tolerant condition of cv. Leccino, which could be exploited for lessening the economic impact of the disease on the local olive industry, transcriptional changes occurring in plants of the two cultivars affected by Xfp were investigated. A global quantitative transcriptome profiling comparing susceptible (Ogliarola salentina) and tolerant (Leccino) olive cultivars, infected or not by Xfp, was done on messenger RNA (mRNAs) extracted from xylem tissues. The study revealed that 659 and 447 genes were differentially regulated in cvs Leccino and Ogliarola upon Xfp infection, respectively, whereas 512 genes were altered when the transcriptome of both infected cultivars was compared. Analysis of these differentially expressed genes (DEGs) shows that the presence of Xfp is perceived by the plants of both cultivars, in which it triggers a differential response strongly involving the cell wall. Up-regulation of genes encoding receptor-like kinases (RLK) and receptor-like proteins (RLP) is the predominant response of cv. Leccino, which is missing in cv. Ogliarola salentina. Moreover, both cultivars react with a strong re-modelling of cell wall proteins. These data suggest that Xfp elicits a different transcriptome response in the two cultivars, which determines a lower pathogen concentration in cv. Leccino and indicates that this cultivar may harbor genetic constituents and/or regulatory elements which counteract Xfp infection. Collectively these findings suggest that cv. Leccino is endowed with an intrinsic tolerance to Xfp, which makes it eligible for further studies aiming at investigating molecular basis and pathways modulating its different defense response.

Benchmark Dose Modeling Estimates of the Concentrations of Inorganic Arsenic That Induce Changes to the Neonatal Transcriptome, Proteome, and Epigenome in a Pregnancy Cohort.

PubMed

Rager, Julia E; Auerbach, Scott S; Chappell, Grace A; Martin, Elizabeth; Thompson, Chad M; Fry, Rebecca C

2017-10-16

Prenatal inorganic arsenic (iAs) exposure influences the expression of critical genes and proteins associated with adverse outcomes in newborns, in part through epigenetic mediators. The doses at which these genomic and epigenomic changes occur have yet to be evaluated in the context of dose-response modeling. The goal of the present study was to estimate iAs doses that correspond to changes in transcriptomic, proteomic, epigenomic, and integrated multi-omic signatures in human cord blood through benchmark dose (BMD) modeling. Genome-wide DNA methylation, microRNA expression, mRNA expression, and protein expression levels in cord blood were modeled against total urinary arsenic (U-tAs) levels from pregnant women exposed to varying levels of iAs. Dose-response relationships were modeled in BMDExpress, and BMDs representing 10% response levels were estimated. Overall, DNA methylation changes were estimated to occur at lower exposure concentrations in comparison to other molecular endpoints. Multi-omic module eigengenes were derived through weighted gene co-expression network analysis, representing co-modulated signatures across transcriptomic, proteomic, and epigenomic profiles. One module eigengene was associated with decreased gestational age occurring alongside increased iAs exposure. Genes/proteins within this module eigengene showed enrichment for organismal development, including potassium voltage-gated channel subfamily Q member 1 (KCNQ1), an imprinted gene showing differential methylation and expression in response to iAs. Modeling of this prioritized multi-omic module eigengene resulted in a BMD(BMDL) of 58(45) μg/L U-tAs, which was estimated to correspond to drinking water arsenic concentrations of 51(40) μg/L. Results are in line with epidemiological evidence supporting effects of prenatal iAs occurring at levels <100 μg As/L urine. Together, findings present a variety of BMD measures to estimate doses at which prenatal iAs exposure influences neonatal outcome-relevant transcriptomic, proteomic, and epigenomic profiles.

Cardiac transcriptome profiling of diabetic Akita mice using microarray and next generation sequencing

PubMed Central

Kesherwani, Varun; Shahshahan, Hamid R.

2017-01-01

Although diabetes mellitus (DM) causes cardiomyopathy and exacerbates heart failure, the underlying molecular mechanisms for diabetic cardiomyopathy/heart failure are poorly understood. Insulin2 mutant (Ins2+/-) Akita is a mouse model of T1DM, which manifests cardiac dysfunction. However, molecular changes at cardiac transcriptome level that lead to cardiomyopathy remain unclear. To understand the molecular changes in the heart of diabetic Akita mice, we profiled cardiac transcriptome of Ins2+/- Akita and Ins2+/+ control mice using next generation sequencing (NGS) and microarray, and determined the implications of differentially expressed genes on various heart failure signaling pathways using Ingenuity pathway (IPA) analysis. First, we validated hyperglycemia, increased cardiac fibrosis, and cardiac dysfunction in twelve-week male diabetic Akita. Then, we analyzed the transcriptome levels in the heart. NGS analyses on Akita heart revealed 137 differentially expressed transcripts, where Bone Morphogenic Protein-10 (BMP10) was the most upregulated and hairy and enhancer of split-related (HELT) was the most downregulated gene. Moreover, twelve long non-coding RNAs (lncRNAs) were upregulated. The microarray analyses on Akita heart showed 351 differentially expressed transcripts, where vomeronasal-1 receptor-180 (Vmn1r180) was the most upregulated and WD Repeat Domain 83 Opposite Strand (WDR83OS) was the most downregulated gene. Further, miR-101c and H19 lncRNA were upregulated but Neat1 lncRNA was downregulated in Akita heart. Eleven common genes were upregulated in Akita heart in both NGS and microarray analyses. IPA analyses revealed the role of these differentially expressed genes in key signaling pathways involved in diabetic cardiomyopathy. Our results provide a platform to initiate focused future studies by targeting these genes and/or non-coding RNAs, which are differentially expressed in Akita hearts and are involved in diabetic cardiomyopathy. PMID:28837672

Transcriptome Profiling of Neurosensory Perception Genes in Wing Tissue of Two Evolutionary Distant Insect Orders: Diptera (Drosophila melanogaster) and Hemiptera (Acyrthosiphon pisum).

PubMed

Agnel, Sandra; da Rocha, Martine; Robichon, Alain

2017-12-01

The neurogenesis and neuronal functions in insect wing have been understudied mainly due to technical hindrances that have prevented electrophysiology studies for decades. The reason is that the nano-architecture of the wing chemosensory bristles hampers the receptors accessibility of odorants/tastants to receptors in fixed setup, whereas in nature, the wing flapping mixes these molecules in bristle lymph. In this report, we analyzed the transcriptome of the wing tissue of two species phylogenetically strongly divergent: Drosophila melanogaster a generic model for diptera order (complete metamorphosis) and the aphid acyrthosiphon pisum, representative of hemiptera order (incomplete metamorphosis) for which a conditional winged/wingless polyphenism is under control of population density and resources. The transcriptome shows that extensive gene networks involved in chemosensory perception are active in adult wing for both species. Surprisingly, the specific transcripts of genes that are commonly found in eye were present in Drosophila wing but not in aphid. The analysis reveals that in the aphid conditional wing, expressed genes show strong similarities with those in the gut epithelia. This suggests that the epithelial cell layer between the cuticle sheets is persistent at least in young aphid adult, whereas it disappears after emergence in Drosophila. Despite marked differences between the two transcriptomes, the results highlight the probable universalism of wing chemosensory function in the holometabolous and hemimetabolous orders of winged insects.

CXCL4 induces a unique transcriptome in monocyte-derived macrophages

PubMed Central

Gleissner, Christian A.; Shaked, Iftach; Little, Kristina M.; Ley, Klaus

2012-01-01

In atherosclerotic arteries, blood monocytes differentiate to macrophages in the presence of growth factors like macrophage colony-stimulation factor (MCSF) and chemokines like platelet factor 4 (CXCL4). To compare the gene expression signature of CXCL4-induced macrophages with MCSF-induced macrophages or macrophages polarized with IFN-γ/LPS (M1) or IL-4 (M2), we cultured primary human peripheral blood monocytes for six days. mRNA expression was measured by Affymetrix gene chips and differences were analyzed by Local Pooled Error test, Profile of Complex Functionality and Gene Set Enrichment Analysis. 375 genes were differentially expressed between MCSF- and CXCL4-induced macrophages, 206 of them overexpressed in CXCL4 macrophages coding for genes implicated in the inflammatory/immune response, antigen processing/presentation, and lipid metabolism. CXCL4-induced macrophages overexpressed some M1 and M2 genes and the corresponding cytokines at the protein level, however, their transcriptome clustered with neither M1 nor M2 transcriptomes. They almost completely lost the ability to phagocytose zymosan beads. Genes linked to atherosclerosis were not consistently up- or downregulated. Scavenger receptors showed lower and cholesterol efflux transporters higher expression in CXCL4- than MCSF-induced macrophages, resulting in lower LDL content. We conclude that CXCL4 induces a unique macrophage transcriptome distinct from known macrophage types, defining a new macrophage differentiation that we propose to call M4. PMID:20335529

Alterations in the Aedes aegypti Transcriptome during Infection with West Nile, Dengue and Yellow Fever Viruses

PubMed Central

Colpitts, Tonya M.; Cox, Jonathan; Vanlandingham, Dana L.; Feitosa, Fabiana M.; Cheng, Gong; Kurscheid, Sebastian; Wang, Penghua; Krishnan, Manoj N.; Higgs, Stephen; Fikrig, Erol

2011-01-01

West Nile (WNV), dengue (DENV) and yellow fever (YFV) viruses are (re)emerging, mosquito-borne flaviviruses that cause human disease and mortality worldwide. Alterations in mosquito gene expression common and unique to individual flaviviral infections are poorly understood. Here, we present a microarray analysis of the Aedes aegypti transcriptome over time during infection with DENV, WNV or YFV. We identified 203 mosquito genes that were ≥5-fold differentially up-regulated (DUR) and 202 genes that were ≥10-fold differentially down-regulated (DDR) during infection with one of the three flaviviruses. Comparative analysis revealed that the expression profile of 20 DUR genes and 15 DDR genes was quite similar between the three flaviviruses on D1 of infection, indicating a potentially conserved transcriptomic signature of flaviviral infection. Bioinformatics analysis revealed changes in expression of genes from diverse cellular processes, including ion binding, transport, metabolic processes and peptidase activity. We also demonstrate that virally-regulated gene expression is tissue-specific. The overexpression of several virally down-regulated genes decreased WNV infection in mosquito cells and Aedes aegypti mosquitoes. Among these, a pupal cuticle protein was shown to bind WNV envelope protein, leading to inhibition of infection in vitro and the prevention of lethal WNV encephalitis in mice. This work provides an extensive list of targets for controlling flaviviral infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses. PMID:21909258

Alterations in the Aedes aegypti transcriptome during infection with West Nile, dengue and yellow fever viruses.

PubMed

Colpitts, Tonya M; Cox, Jonathan; Vanlandingham, Dana L; Feitosa, Fabiana M; Cheng, Gong; Kurscheid, Sebastian; Wang, Penghua; Krishnan, Manoj N; Higgs, Stephen; Fikrig, Erol

2011-09-01

West Nile (WNV), dengue (DENV) and yellow fever (YFV) viruses are (re)emerging, mosquito-borne flaviviruses that cause human disease and mortality worldwide. Alterations in mosquito gene expression common and unique to individual flaviviral infections are poorly understood. Here, we present a microarray analysis of the Aedes aegypti transcriptome over time during infection with DENV, WNV or YFV. We identified 203 mosquito genes that were ≥ 5-fold differentially up-regulated (DUR) and 202 genes that were ≥ 10-fold differentially down-regulated (DDR) during infection with one of the three flaviviruses. Comparative analysis revealed that the expression profile of 20 DUR genes and 15 DDR genes was quite similar between the three flaviviruses on D1 of infection, indicating a potentially conserved transcriptomic signature of flaviviral infection. Bioinformatics analysis revealed changes in expression of genes from diverse cellular processes, including ion binding, transport, metabolic processes and peptidase activity. We also demonstrate that virally-regulated gene expression is tissue-specific. The overexpression of several virally down-regulated genes decreased WNV infection in mosquito cells and Aedes aegypti mosquitoes. Among these, a pupal cuticle protein was shown to bind WNV envelope protein, leading to inhibition of infection in vitro and the prevention of lethal WNV encephalitis in mice. This work provides an extensive list of targets for controlling flaviviral infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses.

Transcriptomic Analysis and the Expression of Disease-Resistant Genes in Oryza meyeriana under Native Condition

PubMed Central

He, Bin; Tao, Xiang; Gu, Yinghong; Wei, Changhe; Cheng, Xiaojie; Xiao, Suqin; Cheng, Zaiquan; Zhang, Yizheng

2015-01-01

Oryza meyeriana (O. meyeriana), with a GG genome type (2n = 24), accumulated plentiful excellent characteristics with respect to resistance to many diseases such as rice shade and blast, even immunity to bacterial blight. It is very important to know if the diseases-resistant genes exist and express in this wild rice under native conditions. However, limited genomic or transcriptomic data of O. meyeriana are currently available. In this study, we present the first comprehensive characterization of the O. meyeriana transcriptome using RNA-seq and obtained 185,323 contigs with an average length of 1,692 bp and an N50 of 2,391 bp. Through differential expression analysis, it was found that there were most tissue-specifically expressed genes in roots, and next to stems and leaves. By similarity search against protein databases, 146,450 had at least a significant alignment to existed gene models. Comparison with the Oryza sativa (japonica-type Nipponbare and indica-type 93–11) genomes revealed that 13% of the O. meyeriana contigs had not been detected in O. sativa. Many diseases-resistant genes, such as bacterial blight resistant, blast resistant, rust resistant, fusarium resistant, cyst nematode resistant and downy mildew gene, were mined from the transcriptomic database. There are two kinds of rice bacterial blight-resistant genes (Xa1 and Xa26) differentially or specifically expressed in O. meyeriana. The 4 Xa1 contigs were all only expressed in root, while three of Xa26 contigs have the highest expression level in leaves, two of Xa26 contigs have the highest expression profile in stems and one of Xa26 contigs was expressed dominantly in roots. The transcriptomic database of O. meyeriana has been constructed and many diseases-resistant genes were found to express under native condition, which provides a foundation for future discovery of a number of novel genes and provides a basis for studying the molecular mechanisms associated with disease resistance in O. meyeriana. PMID:26640944

Integrated transcriptome catalogue and organ-specific profiling of gene expression in fertile garlic (Allium sativum L.).

PubMed

Kamenetsky, Rina; Faigenboim, Adi; Shemesh Mayer, Einat; Ben Michael, Tomer; Gershberg, Chen; Kimhi, Sagie; Esquira, Itzhak; Rohkin Shalom, Sarit; Eshel, Dani; Rabinowitch, Haim D; Sherman, Amir

2015-01-22

Garlic is cultivated and consumed worldwide as a popular condiment and green vegetable with medicinal and neutraceutical properties. Garlic cultivars do not produce seeds, and therefore, this plant has not been the subject of either classical breeding or genetic studies. However, recent achievements in fertility restoration in a number of genotypes have led to flowering and seed production, thus enabling genetic studies and breeding in garlic. A transcriptome catalogue of fertile garlic was produced from multiplexed gene libraries, using RNA collected from various plant organs, including inflorescences and flowers. Over 32 million 250-bp paired-end reads were assembled into an extensive transcriptome of 240,000 contigs. An abundant transcriptome assembled separately from 102,000 highly expressed contigs was annotated and analyzed for gene ontology and metabolic pathways. Organ-specific analysis showed significant variation of gene expression between plant organs, with the highest number of specific reads in inflorescences and flowers. Analysis of the enriched biological processes and molecular functions revealed characteristic patterns for stress response, flower development and photosynthetic activity. Orthologues of key flowering genes were differentially expressed, not only in reproductive tissues, but also in leaves and bulbs, suggesting their role in flower-signal transduction and the bulbing process. More than 100 variants and isoforms of enzymes involved in organosulfur metabolism were differentially expressed and had organ-specific patterns. In addition to plant genes, viral RNA of at least four garlic viruses was detected, mostly in the roots and cloves, whereas only 1-4% of the reads were found in the foliage leaves. The de novo transcriptome of fertile garlic represents a new resource for research and breeding of this important crop, as well as for the development of effective molecular markers for useful traits, including fertility and seed production, resistance to pests and neutraceutical characteristics.

Comprehensive RNA-Seq profiling to evaluate lactating sheep mammary gland transcriptome

PubMed Central

Suárez-Vega, Aroa; Gutiérrez-Gil, Beatriz; Klopp, Christophe; Tosser-Klopp, Gwenola; Arranz, Juan-José

2016-01-01

RNA-Seq enables the generation of extensive transcriptome information providing the capability to characterize transcripts (including alternative isoforms and polymorphism), to quantify expression and to identify differential regulation in a single experiment. Our aim in this study was to take advantage of using RNA-Seq high-throughput technology to provide a comprehensive transcriptome profiling of the sheep lactating mammary gland. Eight ewes of two dairy sheep breeds with differences in milk production traits were used in this experiment (four Churra and four Assaf ewes). Milk samples from these animals were collected on days 10, 50, 120 and 150 after lambing to cover the various physiological stages of the mammary gland across the complete lactation. RNA samples were extracted from milk somatic cells. The RNA-Seq dataset was generated using an Illumina HiSeq 2000 sequencer. The information reported here will be useful to understand the biology of lactation in sheep, providing also an opportunity to characterize their different patterns on milk production aptitude. PMID:27377755

Comprehensive RNA-Seq profiling to evaluate lactating sheep mammary gland transcriptome.

PubMed

Suárez-Vega, Aroa; Gutiérrez-Gil, Beatriz; Klopp, Christophe; Tosser-Klopp, Gwenola; Arranz, Juan-José

2016-07-05

RNA-Seq enables the generation of extensive transcriptome information providing the capability to characterize transcripts (including alternative isoforms and polymorphism), to quantify expression and to identify differential regulation in a single experiment. Our aim in this study was to take advantage of using RNA-Seq high-throughput technology to provide a comprehensive transcriptome profiling of the sheep lactating mammary gland. Eight ewes of two dairy sheep breeds with differences in milk production traits were used in this experiment (four Churra and four Assaf ewes). Milk samples from these animals were collected on days 10, 50, 120 and 150 after lambing to cover the various physiological stages of the mammary gland across the complete lactation. RNA samples were extracted from milk somatic cells. The RNA-Seq dataset was generated using an Illumina HiSeq 2000 sequencer. The information reported here will be useful to understand the biology of lactation in sheep, providing also an opportunity to characterize their different patterns on milk production aptitude.

Genome-wide comparative transcriptome analysis of CMS-D2 and its maintainer and restorer lines in upland cotton.

PubMed

Wu, Jianyong; Zhang, Meng; Zhang, Bingbing; Zhang, Xuexian; Guo, Liping; Qi, Tingxiang; Wang, Hailin; Zhang, Jinfa; Xing, Chaozhu

2017-06-08

Cytoplasmic male sterility (CMS) conferred by the cytoplasm from Gossypium harknessii (D2) is an important system for hybrid seed production in Upland cotton (G. hirsutum). The male sterility of CMS-D2 (i.e., A line) can be restored to fertility by a restorer (i.e., R line) carrying the restorer gene Rf1 transferred from the D2 nuclear genome. However, the molecular mechanisms of CMS-D2 and its restoration are poorly understood. In this study, a genome-wide comparative transcriptome analysis was performed to identify differentially expressed genes (DEGs) in flower buds among the isogenic fertile R line and sterile A line derived from a backcross population (BC 8 F 1 ) and the recurrent parent, i.e., the maintainer (B line). A total of 1464 DEGs were identified among the three isogenic lines, and the Rf1-carrying Chr_D05 and its homeologous Chr_A05 had more DEGs than other chromosomes. The results of GO and KEGG enrichment analysis showed differences in circadian rhythm between the fertile and sterile lines. Eleven DEGs were selected for validation using qRT-PCR, confirming the accuracy of the RNA-seq results. Through genome-wide comparative transcriptome analysis, the differential expression profiles of CMS-D2 and its maintainer and restorer lines in Upland cotton were identified. Our results provide an important foundation for further studies into the molecular mechanisms of the interactions between the restorer gene Rf1 and the CMS-D2 cytoplasm.

Comparative analysis of chrysanthemum transcriptome in response to three RNA viruses: Cucumber mosaic virus, Tomato spotted wilt virus and Potato virus X.

PubMed

Choi, Hoseong; Jo, Yeonhwa; Lian, Sen; Jo, Kyoung-Min; Chu, Hyosub; Yoon, Ju-Yeon; Choi, Seung-Kook; Kim, Kook-Hyung; Cho, Won Kyong

2015-06-01

The chrysanthemum is one of popular flowers in the world and a host for several viruses. So far, molecular interaction studies between the chrysanthemum and viruses are limited. In this study, we carried out a transcriptome analysis of chrysanthemum in response to three different viruses including Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV) and Potato virus X (PVX). A chrysanthemum 135K microarray derived from expressed sequence tags was successfully applied for the expression profiles of the chrysanthemum at early stage of virus infection. Finally, we identified a total of 125, 70 and 124 differentially expressed genes (DEGs) for CMV, TSWV and PVX, respectively. Many DEGs were virus specific; however, 33 DEGs were commonly regulated by three viruses. Gene ontology (GO) enrichment analysis identified a total of 132 GO terms, and of them, six GO terms related stress response and MCM complex were commonly identified for three viruses. Several genes functioning in stress response such as chitin response and ethylene mediated signaling pathway were up-regulated indicating their involvement in establishment of host immune system. In particular, TSWV infection significantly down-regulated genes related to DNA metabolic process including DNA replication, chromatin organization, histone modification and cytokinesis, and they are mostly targeted to nucleosome and MCM complex. Taken together, our comparative transcriptome analysis revealed several genes related to hormone mediated viral stress response and DNA modification. The identified chrysanthemums genes could be good candidates for further functional study associated with resistant to various plant viruses.

Transcriptome analysis of root response to citrus blight based on the newly assembled Swingle citrumelo draft genome.

PubMed

Zhang, Yunzeng; Barthe, Gary; Grosser, Jude W; Wang, Nian

2016-07-08

Citrus blight is a citrus tree overall decline disease and causes serious losses in the citrus industry worldwide. Although it was described more than one hundred years ago, its causal agent remains unknown and its pathophysiology is not well determined, which hampers our understanding of the disease and design of suitable disease management. In this study, we sequenced and assembled the draft genome for Swingle citrumelo, one important citrus rootstock. The draft genome is approximately 280 Mb, which covers 74 % of the estimated Swingle citrumelo genome and the average coverage is around 15X. The draft genome of Swingle citrumelo enabled us to conduct transcriptome analysis of roots of blight and healthy Swingle citrumelo using RNA-seq. The RNA-seq was reliable as evidenced by the high consistence of RNA-seq analysis and quantitative reverse transcription PCR results (R(2) = 0.966). Comparison of the gene expression profiles between blight and healthy root samples revealed the molecular mechanism underneath the characteristic blight phenotypes including decline, starch accumulation, and drought stress. The JA and ET biosynthesis and signaling pathways showed decreased transcript abundance, whereas SA-mediated defense-related genes showed increased transcript abundance in blight trees, suggesting unclassified biotrophic pathogen was involved in this disease. Overall, the Swingle citrumelo draft genome generated in this study will advance our understanding of plant biology and contribute to the citrus breeding. Transcriptome analysis of blight and healthy trees deepened our understanding of the pathophysiology of citrus blight.

Comparative transcriptome analysis of pepper (Capsicum annuum) revealed common regulons in multiple stress conditions and hormone treatments.

PubMed

Lee, Sanghyeob; Choi, Doil

2013-09-01

Global transcriptome analysis revealed common regulons for biotic/abiotic stresses, and some of these regulons encoding signaling components in both stresses were newly identified in this study. In this study, we aimed to identify plant responses to multiple stress conditions and discover the common regulons activated under a variety of stress conditions. Global transcriptome analysis revealed that salicylic acid (SA) may affect the activation of abiotic stress-responsive genes in pepper. Our data indicate that methyl jasmonate (MeJA) and ethylene (ET)-responsive genes were primarily activated by biotic stress, while abscisic acid (ABA)-responsive genes were activated under both types of stresses. We also identified differentially expressed gene (DEG) responses to specific stress conditions. Biotic stress induces more DEGs than those induced by abiotic and hormone applications. The clustering analysis using DEGs indicates that there are common regulons for biotic or abiotic stress conditions. Although SA and MeJA have an antagonistic effect on gene expression levels, SA and MeJA show a largely common regulation as compared to the regulation at the DEG expression level induced by other hormones. We also monitored the expression profiles of DEG encoding signaling components. Twenty-two percent of these were commonly expressed in both stress conditions. The importance of this study is that several genes commonly regulated by both stress conditions may have future applications for creating broadly stress-tolerant pepper plants. This study revealed that there are complex regulons in pepper plant to both biotic and abiotic stress conditions.

Transcriptomics of coping strategies in free-swimming Lepeophtheirus salmonis (Copepoda) larvae responding to abiotic stress.

PubMed

Sutherland, Ben J G; Jantzen, Stuart G; Yasuike, Motoshige; Sanderson, Dan S; Koop, Ben F; Jones, Simon R M

2012-12-01

The salmon louse Lepeophtheirus salmonis is a marine ectoparasite of wild and farmed salmon in the Northern Hemisphere. Infections of farmed salmon are of economic and ecological concern. Nauplius and copepodid salmon lice larvae are free-swimming and disperse in the water column until they encounter a host. In this study, we characterized the sublethal stress responses of L. salmonis copepodid larvae by applying a 38K oligonucleotide microarray to profile transcriptomes following 24 h exposures to suboptimal salinity (30-10 parts per thousand (‰)) or temperature (16-4 °C) environments. Hyposalinity exposure resulted in large-scale gene expression changes relative to those elicited by a thermal gradient. Subsequently, transcriptome responses to a more finely resolved salinity gradient between 30 ‰ and 25 ‰ were profiled. Minimal changes occurred at 29 ‰ or 28 ‰, a threshold of response was identified at 27 ‰, and the largest response was at 25 ‰. Differentially expressed genes were clustered by pattern of expression, and clusters were characterized by functional enrichment analysis. Results indicate larval copepods adopt two distinct coping strategies in response to short-term hyposaline stress: a primary response using molecular chaperones and catabolic processes at 27 ‰; and a secondary response up-regulating ion pumps, transporters, a different suite of chaperones and apoptosis-related transcripts at 26 ‰ and 25 ‰. The results further our understanding of the tolerances of L. salmonis copepodids to salinity and temperature gradients and may assist in the development of salmon louse management strategies. © 2012 Blackwell Publishing Ltd.

Arabidopsis whole-transcriptome profiling defines the features of coordinated regulations that occur during secondary growth.

PubMed

Ko, Jae-Heung; Han, Kyung-Hwan

2004-05-01

Secondary growth in the inflorescence stems of Arabidopsis plants was induced by a combination of short-day and long-day treatments. The induced stems were divided into three different stem developmental stages (i.e., immature, intermediate, and mature) with regard to secondary growth. Whole transcriptome microarrays were used to examine the changes in global gene expression occurring at the different stem developmental stages. Over 70% of the Arabidopsis transcriptome was expressed in the stem tissues. In the mature stems with secondary growth, 567 genes were upregulated 5-fold or higher and 530 were downregulated, when compared to immature stems (with no secondary growth) and 10-day old seedlings (with no inflorescence stem). The transcription phenotypes obtained from the stems at different developmental stages largely confirm the existing insights into the biochemical processes involved in the sequential events that lead to wood formation. The major difference found between the stems undergoing secondary growth and only primary growth was in the expression profiles of transcriptional regulation-and signal transduction-related genes. An analysis of several shoot apical meristem (SAM) activity-related gene expression patterns in the stems indicated that the genetic control of secondary meristem activity might be governed by a different mechanism from that of SAM. The current study established the expression patterns of many unknown genes and identified candidate genes that are involved in the genetic regulation of secondary growth. The findings described in this report should improve our understanding of the molecular mechanisms that regulate the growth and development of the stem.

Comparative transcriptome profiling of upland (VS16) and lowland (AP13) ecotypes of switchgrass.

PubMed

Ayyappan, Vasudevan; Saha, Malay C; Thimmapuram, Jyothi; Sripathi, Venkateswara R; Bhide, Ketaki P; Fiedler, Elizabeth; Hayford, Rita K; Kalavacharla, Venu Kal

2017-01-01

Transcriptomes of two switchgrass genotypes representing the upland and lowland ecotypes will be key tools in switchgrass genome annotation and biotic and abiotic stress functional genomics. Switchgrass (Panicum virgatum L.) is an important bioenergy feedstock for cellulosic ethanol production. We report genome-wide transcriptome profiling of two contrasting tetraploid switchgrass genotypes, VS16 and AP13, representing the upland and lowland ecotypes, respectively. A total of 268 million Illumina short reads (50 nt) were generated, of which, 133 million were obtained in AP13 and the rest 135 million in VS16. More than 90% of these reads were mapped to the switchgrass reference genome (V1.1). We identified 6619 and 5369 differentially expressed genes in VS16 and AP13, respectively. Gene ontology and KEGG pathway analysis identified key genes that regulate important pathways including C4 photosynthesis, photorespiration and phenylpropanoid metabolism. A series of genes (33) involved in photosynthetic pathway were up-regulated in AP13 but only two genes showed higher expression in VS16. We identified three dicarboxylate transporter homologs that were highly expressed in AP13. Additionally, genes that mediate drought, heat, and salinity tolerance were also identified. Vesicular transport proteins, syntaxin and signal recognition particles were seen to be up-regulated in VS16. Analyses of selected genes involved in biosynthesis of secondary metabolites, plant-pathogen interaction, membrane transporters, heat, drought and salinity stress responses confirmed significant variation in the relative expression reflected in RNA-Seq data between VS16 and AP13 genotypes. The phenylpropanoid pathway genes identified here are potential targets for biofuel conversion.

The Long Noncoding RNA Transcriptome of Dictyostelium discoideum Development.

PubMed

Rosengarten, Rafael D; Santhanam, Balaji; Kokosar, Janez; Shaulsky, Gad

2017-02-09

Dictyostelium discoideum live in the soil as single cells, engulfing bacteria and growing vegetatively. Upon starvation, tens of thousands of amoebae enter a developmental program that includes aggregation, multicellular differentiation, and sporulation. Major shifts across the protein-coding transcriptome accompany these developmental changes. However, no study has presented a global survey of long noncoding RNAs (ncRNAs) in D. discoideum To characterize the antisense and long intergenic noncoding RNA (lncRNA) transcriptome, we analyzed previously published developmental time course samples using an RNA-sequencing (RNA-seq) library preparation method that selectively depletes ribosomal RNAs (rRNAs). We detected the accumulation of transcripts for 9833 protein-coding messenger RNAs (mRNAs), 621 lncRNAs, and 162 putative antisense RNAs (asRNAs). The noncoding RNAs were interspersed throughout the genome, and were distinct in expression level, length, and nucleotide composition. The noncoding transcriptome displayed a temporal profile similar to the coding transcriptome, with stages of gradual change interspersed with larger leaps. The transcription profiles of some noncoding RNAs were strongly correlated with known differentially expressed coding RNAs, hinting at a functional role for these molecules during development. Examining the mitochondrial transcriptome, we modeled two novel antisense transcripts. We applied yet another ribosomal depletion method to a subset of the samples to better retain transfer RNA (tRNA) transcripts. We observed polymorphisms in tRNA anticodons that suggested a post-transcriptional means by which D. discoideum compensates for codons missing in the genomic complement of tRNAs. We concluded that the prevalence and characteristics of long ncRNAs indicate that these molecules are relevant to the progression of molecular and cellular phenotypes during development. Copyright © 2017 Rosengarten et al.

Dual RNA-seq reveals no plastic transcriptional response of the coccidian parasite Eimeria falciformis to host immune defenses.

PubMed

Ehret, Totta; Spork, Simone; Dieterich, Christoph; Lucius, Richard; Heitlinger, Emanuel

2017-09-05

Parasites can either respond to differences in immune defenses that exist between individual hosts plastically or, alternatively, follow a genetically canalized ("hard wired") program of infection. Assuming that large-scale functional plasticity would be discernible in the parasite transcriptome we have performed a dual RNA-seq study of the lifecycle of Eimeria falciformis using infected mice with different immune status as models for coccidian infections. We compared parasite and host transcriptomes (dual transcriptome) between naïve and challenge infected mice, as well as between immune competent and immune deficient ones. Mice with different immune competence show transcriptional differences as well as differences in parasite reproduction (oocyst shedding). Broad gene categories represented by differently abundant host genes indicate enrichments for immune reaction and tissue repair functions. More specifically, TGF-beta, EGF, TNF and IL-1 and IL-6 are examples of functional annotations represented differently depending on host immune status. Much in contrast, parasite transcriptomes were neither different between Coccidia isolated from immune competent and immune deficient mice, nor between those harvested from naïve and challenge infected mice. Instead, parasite transcriptomes have distinct profiles early and late in infection, characterized largely by biosynthesis or motility associated functional gene groups, respectively. Extracellular sporozoite and oocyst stages showed distinct transcriptional profiles and sporozoite transcriptomes were found enriched for species specific genes and likely pathogenicity factors. We propose that the niche and host-specific parasite E. falciformis uses a genetically canalized program of infection. This program is likely fixed in an evolutionary process rather than employing phenotypic plasticity to interact with its host. This in turn might limit the potential of the parasite to adapt to new host species or niches, forcing it to coevolve with its host.

Surviving in a toxic world: transcriptomics and gene expression profiling in response to environmental pollution in the critically endangered European eel.

PubMed

Pujolar, Jose Martin; Marino, Ilaria A M; Milan, Massimo; Coppe, Alessandro; Maes, Gregory E; Capoccioni, Fabrizio; Ciccotti, Eleonora; Bervoets, Lieven; Covaci, Adrian; Belpaire, Claude; Cramb, Gordon; Patarnello, Tomaso; Bargelloni, Luca; Bortoluzzi, Stefania; Zane, Lorenzo

2012-09-25

Genomic and transcriptomic approaches have the potential for unveiling the genome-wide response to environmental perturbations. The abundance of the catadromous European eel (Anguilla anguilla) stock has been declining since the 1980s probably due to a combination of anthropogenic and climatic factors. In this paper, we explore the transcriptomic dynamics between individuals from high (river Tiber, Italy) and low pollution (lake Bolsena, Italy) environments, which were measured for 36 PCBs, several organochlorine pesticides and brominated flame retardants and nine metals. To this end, we first (i) updated the European eel transcriptome using deep sequencing data with a total of 640,040 reads assembled into 44,896 contigs (Eeelbase release 2.0), and (ii) developed a transcriptomic platform for global gene expression profiling in the critically endangered European eel of about 15,000 annotated contigs, which was applied to detect differentially expressed genes between polluted sites. Several detoxification genes related to metabolism of pollutants were upregulated in the highly polluted site, including genes that take part in phase I of the xenobiotic metabolism (CYP3A), phase II (glutathione-S-transferase) and oxidative stress (glutathione peroxidase). In addition, key genes in the mitochondrial respiratory chain and oxidative phosphorylation were down-regulated at the Tiber site relative to the Bolsena site. Together with the induced high expression of detoxification genes, the suggested lowered expression of genes supposedly involved in metabolism suggests that pollution may also be associated with decreased respiratory and energy production.

A combination of LongSAGE with Solexa sequencing is well suited to explore the depth and the complexity of transcriptome

PubMed Central

Hanriot, Lucie; Keime, Céline; Gay, Nadine; Faure, Claudine; Dossat, Carole; Wincker, Patrick; Scoté-Blachon, Céline; Peyron, Christelle; Gandrillon, Olivier

2008-01-01

Background "Open" transcriptome analysis methods allow to study gene expression without a priori knowledge of the transcript sequences. As of now, SAGE (Serial Analysis of Gene Expression), LongSAGE and MPSS (Massively Parallel Signature Sequencing) are the mostly used methods for "open" transcriptome analysis. Both LongSAGE and MPSS rely on the isolation of 21 pb tag sequences from each transcript. In contrast to LongSAGE, the high throughput sequencing method used in MPSS enables the rapid sequencing of very large libraries containing several millions of tags, allowing deep transcriptome analysis. However, a bias in the complexity of the transcriptome representation obtained by MPSS was recently uncovered. Results In order to make a deep analysis of mouse hypothalamus transcriptome avoiding the limitation introduced by MPSS, we combined LongSAGE with the Solexa sequencing technology and obtained a library of more than 11 millions of tags. We then compared it to a LongSAGE library of mouse hypothalamus sequenced with the Sanger method. Conclusion We found that Solexa sequencing technology combined with LongSAGE is perfectly suited for deep transcriptome analysis. In contrast to MPSS, it gives a complex representation of transcriptome as reliable as a LongSAGE library sequenced by the Sanger method. PMID:18796152

Genome wide transcriptional profiling of Herbaspirillum seropedicae SmR1 grown in the presence of naringenin

PubMed Central

Tadra-Sfeir, Michelle Z.; Faoro, Helisson; Camilios-Neto, Doumit; Brusamarello-Santos, Liziane; Balsanelli, Eduardo; Weiss, Vinicius; Baura, Valter A.; Wassem, Roseli; Cruz, Leonardo M.; De Oliveira Pedrosa, Fábio; Souza, Emanuel M.; Monteiro, Rose A.

2015-01-01

Herbaspirillum seropedicae is a diazotrophic bacterium which associates endophytically with economically important gramineae. Flavonoids such as naringenin have been shown to have an effect on the interaction between H. seropedicae and its host plants. We used a high-throughput sequencing based method (RNA-Seq) to access the influence of naringenin on the whole transcriptome profile of H. seropedicae. Three hundred and four genes were downregulated and seventy seven were upregulated by naringenin. Data analysis revealed that genes related to bacterial flagella biosynthesis, chemotaxis and biosynthesis of peptidoglycan were repressed by naringenin. Moreover, genes involved in aromatic metabolism and multidrug transport efllux were actived. PMID:26052319

Integrated analysis of rice transcriptomic and metabolomic responses to elevated night temperatures identifies sensitivity- and tolerance-related profiles.

PubMed

Glaubitz, Ulrike; Li, Xia; Schaedel, Sandra; Erban, Alexander; Sulpice, Ronan; Kopka, Joachim; Hincha, Dirk K; Zuther, Ellen

2017-01-01

Transcript and metabolite profiling were performed on leaves from six rice cultivars under high night temperature (HNT) condition. Six genes were identified as central for HNT response encoding proteins involved in transcription regulation, signal transduction, protein-protein interactions, jasmonate response and the biosynthesis of secondary metabolites. Sensitive cultivars showed specific changes in transcript abundance including abiotic stress responses, changes of cell wall-related genes, of ABA signaling and secondary metabolism. Additionally, metabolite profiles revealed a highly activated TCA cycle under HNT and concomitantly increased levels in pathways branching off that could be corroborated by enzyme activity measurements. Integrated data analysis using clustering based on one-dimensional self-organizing maps identified two profiles highly correlated with HNT sensitivity. The sensitivity profile included genes of the functional bins abiotic stress, hormone metabolism, cell wall, signaling, redox state, transcription factors, secondary metabolites and defence genes. In the tolerance profile, similar bins were affected with slight differences in hormone metabolism and transcription factor responses. Metabolites of the two profiles revealed involvement of GABA signaling, thus providing a link to the TCA cycle status in sensitive cultivars and of myo-inositol as precursor for inositol phosphates linking jasmonate signaling to the HNT response specifically in tolerant cultivars. © 2016 John Wiley & Sons Ltd.

Transcriptome analysis of carbohydrate metabolism during bulblet formation and development in Lilium davidii var. unicolor.

PubMed

Li, XueYan; Wang, ChunXia; Cheng, JinYun; Zhang, Jing; da Silva, Jaime A Teixeira; Liu, XiaoYu; Duan, Xin; Li, TianLai; Sun, HongMei

2014-12-19

The formation and development of bulblets are crucial to the Lilium genus since these processes are closely related to carbohydrate metabolism, especially to starch and sucrose metabolism. However, little is known about the transcriptional regulation of both processes. To gain insight into carbohydrate-related genes involved in bulblet formation and development, we conducted comparative transcriptome profiling of Lilium davidii var. unicolor bulblets at 0 d, 15 d (bulblets emerged) and 35 d (bulblets formed a basic shape with three or four scales) after scale propagation. Analysis of the transcriptome revealed that a total of 52,901 unigenes with an average sequence size of 630 bp were generated. Based on Clusters of Orthologous Groups (COG) analysis, 8% of the sequences were attributed to carbohydrate transport and metabolism. The results of KEGG pathway enrichment analysis showed that starch and sucrose metabolism constituted the predominant pathway among the three library pairs. The starch content in mother scales and bulblets decreased and increased, respectively, with almost the same trend as sucrose content. Gene expression analysis of the key enzymes in starch and sucrose metabolism suggested that sucrose synthase (SuSy) and invertase (INV), mainly hydrolyzing sucrose, presented higher gene expression in mother scales and bulblets at stages of bulblet appearance and enlargement, while sucrose phosphate synthase (SPS) showed higher expression in bulblets at morphogenesis. The enzymes involved in the starch synthetic direction such as ADPG pyrophosphorylase (AGPase), soluble starch synthase (SSS), starch branching enzyme (SBE) and granule-bound starch synthase (GBSS) showed a decreasing trend in mother scales and higher gene expression in bulblets at bulblet appearance and enlargement stages while the enzyme in the cleavage direction, starch de-branching enzyme (SDBE), showed higher gene expression in mother scales than in bulblets. An extensive transcriptome analysis of three bulblet development stages contributes considerable novel information to our understanding of carbohydrate metabolism-related genes in Lilium at the transcriptional level, and demonstrates the fundamentality of carbohydrate metabolism in bulblet emergence and development at the molecular level. This could facilitate further investigation into the molecular mechanisms underlying these processes in lily and other related species.

Transcriptome sequencing reveals e-cigarette vapor and mainstream-smoke from tobacco cigarettes activate different gene expression profiles in human bronchial epithelial cells

PubMed Central

Shen, Yifei; Wolkowicz, Michael J.; Kotova, Tatyana; Fan, Lonjiang; Timko, Michael P.

2016-01-01

Electronic cigarettes (e-cigarettes) generate an aerosol vapor (e-vapor) thought to represent a less risky alternative to main stream smoke (MSS) of conventional tobacco cigarettes. RNA-seq analysis was used to examine the transcriptomes of differentiated human bronchial epithelial (HBE) cells exposed to air, MSS from 1R5F tobacco reference cigarettes, and e-vapor with and without added nicotine in an in vitro air-liquid interface model for cellular exposure. Our results indicate that while e-vapor does not elicit many of the cell toxicity responses observed in MSS-exposed HBE cells, e-vapor exposure is not benign, but elicits discrete transcriptomic signatures with and without added nicotine. Among the cellular pathways with the most significantly enriched gene expression following e-vapor exposure are the phospholipid and fatty acid triacylglycerol metabolism pathways. Our data suggest that alterations in cellular glycerophopholipid biosynthesis are an important consequences of e-vapor exposure. Moreover, the presence of nicotine in e-vapor elicits a cellular response distinct from e-vapor alone including alterations of cytochrome P450 function, retinoid metabolism, and nicotine catabolism. These studies establish a baseline for future analysis of e-vapor and e-vapor additives that will better inform the FDA and other governmental bodies in discussions of the risks and future regulation of these products. PMID:27041137

The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups

PubMed Central

Curtis, Christina; Shah, Sohrab P.; Chin, Suet-Feung; Turashvili, Gulisa; Rueda, Oscar M.; Dunning, Mark J.; Speed, Doug; Lynch, Andy G.; Samarajiwa, Shamith; Yuan, Yinyin; Gräf, Stefan; Ha, Gavin; Haffari, Gholamreza; Bashashati, Ali; Russell, Roslin; McKinney, Steven; Langerød, Anita; Green, Andrew; Provenzano, Elena; Wishart, Gordon; Pinder, Sarah; Watson, Peter; Markowetz, Florian; Murphy, Leigh; Ellis, Ian; Purushotham, Arnie; Børresen-Dale, Anne-Lise; Brenton, James D.; Tavaré, Simon; Caldas, Carlos; Aparicio, Samuel

2012-01-01

The elucidation of breast cancer subgroups and their molecular drivers requires integrated views of the genome and transcriptome from representative numbers of patients. We present an integrated analysis of copy number and gene expression in a discovery and validation set of 997 and 995 primary breast tumours, respectively, with long-term clinical follow-up. Inherited variants (copy number variants and single nucleotide polymorphisms) and acquired somatic copy number aberrations (CNAs) were associated with expression in ~40% of genes, with the landscape dominated by cis- and trans-acting CNAs. By delineating expression outlier genes driven in cis by CNAs, we identified putative cancer genes, including deletions in PPP2R2A, MTAP and MAP2K4. Unsupervised analysis of paired DNA–RNA profiles revealed novel subgroups with distinct clinical outcomes, which reproduced in the validation cohort. These include a high-risk, oestrogen-receptor-positive 11q13/14 cis-acting subgroup and a favourable prognosis subgroup devoid of CNAs. Trans-acting aberration hotspots were found to modulate subgroup-specific gene networks, including a TCR deletion-mediated adaptive immune response in the ‘CNA-devoid’ subgroup and a basal-specific chromosome 5 deletion-associated mitotic network. Our results provide a novel molecular stratification of the breast cancer population, derived from the impact of somatic CNAs on the transcriptome. PMID:22522925

Single-nucleus RNA-seq of differentiating human myoblasts reveals the extent of fate heterogeneity

PubMed Central

Zeng, Weihua; Jiang, Shan; Kong, Xiangduo; El-Ali, Nicole; Ball, Alexander R.; Ma, Christopher I-Hsing; Hashimoto, Naohiro; Yokomori, Kyoko; Mortazavi, Ali

2016-01-01

Myoblasts are precursor skeletal muscle cells that differentiate into fused, multinucleated myotubes. Current single-cell microfluidic methods are not optimized for capturing very large, multinucleated cells such as myotubes. To circumvent the problem, we performed single-nucleus transcriptome analysis. Using immortalized human myoblasts, we performed RNA-seq analysis of single cells (scRNA-seq) and single nuclei (snRNA-seq) and found them comparable, with a distinct enrichment for long non-coding RNAs (lncRNAs) in snRNA-seq. We then compared snRNA-seq of myoblasts before and after differentiation. We observed the presence of mononucleated cells (MNCs) that remained unfused and analyzed separately from multi-nucleated myotubes. We found that while the transcriptome profiles of myoblast and myotube nuclei are relatively homogeneous, MNC nuclei exhibited significant heterogeneity, with the majority of them adopting a distinct mesenchymal state. Primary transcripts for microRNAs (miRNAs) that participate in skeletal muscle differentiation were among the most differentially expressed lncRNAs, which we validated using NanoString. Our study demonstrates that snRNA-seq provides reliable transcriptome quantification for cells that are otherwise not amenable to current single-cell platforms. Our results further indicate that snRNA-seq has unique advantage in capturing nucleus-enriched lncRNAs and miRNA precursors that are useful in mapping and monitoring differential miRNA expression during cellular differentiation. PMID:27566152

Integrated network analysis identifies fight-club nodes as a class of hubs encompassing key putative switch genes that induce major transcriptome reprogramming during grapevine development.

PubMed

Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

2014-12-01

We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named "fight-club hubs" characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named "switch genes" was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. © 2014 American Society of Plant Biologists. All rights reserved.

Integrated Network Analysis Identifies Fight-Club Nodes as a Class of Hubs Encompassing Key Putative Switch Genes That Induce Major Transcriptome Reprogramming during Grapevine Development[W][OPEN

PubMed Central

Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

2014-01-01

We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named “fight-club hubs” characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named “switch genes” was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops. PMID:25490918

Metastatic breast carcinomas display genomic and transcriptomic heterogeneity

PubMed Central

Weigelt, Britta; Ng, Charlotte KY; Shen, Ronglai; Popova, Tatiana; Schizas, Michail; Natrajan, Rachael; Mariani, Odette; Stern, Marc-Henri; Norton, Larry; Vincent-Salomon, Anne; Reis-Filho, Jorge S

2015-01-01

Metaplastic breast carcinoma is a rare and aggressive histologic type of breast cancer, preferentially displaying a triple-negative phenotype. We sought to define the transcriptomic heterogeneity of metaplastic breast cancers on the basis of current gene expression microarray-based classifiers, and to determine whether these tumors display gene copy number profiles consistent with those of BRCA1-associated breast cancers. Twenty-eight consecutive triple-negative metaplastic breast carcinomas were reviewed, and the metaplastic component present in each frozen specimen was defined (ie, spindle cell, squamous, chondroid metaplasia). RNA and DNA extracted from frozen sections with tumor cell content >60% were subjected to gene expression (Illumina HumanHT-12 v4) and copy number profiling (Affymetrix SNP 6.0), respectively. Using the best practice PAM50/claudin-low microarray-based classifier, all metaplastic breast carcinomas with spindle cell metaplasia were of claudin-low subtype, whereas those with squamous or chondroid metaplasia were preferentially of basal-like subtype. Triple-negative breast cancer subtyping using a dedicated website (http://cbc.mc.vanderbilt.edu/tnbc/) revealed that all metaplastic breast carcinomas with chondroid metaplasia were of mesenchymal-like subtype, spindle cell carcinomas preferentially of unstable or mesenchymal stem-like subtype, and those with squamous metaplasia were of multiple subtypes. None of the cases was classified as immunomodulatory or luminal androgen receptor subtype. Integrative clustering, combining gene expression and gene copy number data, revealed that metaplastic breast carcinomas with spindle cell and chondroid metaplasia were preferentially classified as of integrative clusters 4 and 9, respectively, whereas those with squamous metaplasia were classified into six different clusters. Eight of the 26 metaplastic breast cancers subjected to SNP6 analysis were classified as BRCA1-like. The diversity of histologic features of metaplastic breast carcinomas is reflected at the transcriptomic level, and an association between molecular subtypes and histology was observed. BRCA1-like genomic profiles were found only in a subset (31%) of metaplastic breast cancers, and were not associated with a specific molecular or histologic subtype. PMID:25412848

Decreased hepatotoxic bile acid composition and altered synthesis in progressive human nonalcoholic fatty liver disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lake, April D.; Novak, Petr; Shipkova, Petia

2013-04-15

Bile acids (BAs) have many physiological roles and exhibit both toxic and protective influences within the liver. Alterations in the BA profile may be the result of disease induced liver injury. Nonalcoholic fatty liver disease (NAFLD) is a prevalent form of chronic liver disease characterized by the pathophysiological progression from simple steatosis to nonalcoholic steatohepatitis (NASH). The hypothesis of this study is that the ‘classical’ (neutral) and ‘alternative’ (acidic) BA synthesis pathways are altered together with hepatic BA composition during progression of human NAFLD. This study employed the use of transcriptomic and metabolomic assays to study the hepatic toxicologic BAmore » profile in progressive human NAFLD. Individual human liver samples diagnosed as normal, steatosis, and NASH were utilized in the assays. The transcriptomic analysis of 70 BA genes revealed an enrichment of downregulated BA metabolism and transcription factor/receptor genes in livers diagnosed as NASH. Increased mRNA expression of BAAT and CYP7B1 was observed in contrast to decreased CYP8B1 expression in NASH samples. The BA metabolomic profile of NASH livers exhibited an increase in taurine together with elevated levels of conjugated BA species, taurocholic acid (TCA) and taurodeoxycholic acid (TDCA). Conversely, cholic acid (CA) and glycodeoxycholic acid (GDCA) were decreased in NASH liver. These findings reveal a potential shift toward the alternative pathway of BA synthesis during NASH, mediated by increased mRNA and protein expression of CYP7B1. Overall, the transcriptomic changes of BA synthesis pathway enzymes together with altered hepatic BA composition signify an attempt by the liver to reduce hepatotoxicity during disease progression to NASH. - Highlights: ► Altered hepatic bile acid composition is observed in progressive NAFLD. ► Bile acid synthesis enzymes are transcriptionally altered in NASH livers. ► Increased levels of taurine and conjugated bile acids are observed in NASH. ► Hepatic bile acid synthesis shifts toward the alternative pathway in NASH.« less

A remarkable synergistic effect at the transcriptomic level in peach fruits doubly infected by prunus necrotic ringspot virus and peach latent mosaic viroid.

PubMed

Herranz, Mari Carmen; Niehl, Annette; Rosales, Marlene; Fiore, Nicola; Zamorano, Alan; Granell, Antonio; Pallas, Vicente

2013-05-28

Microarray profiling is a powerful technique to investigate expression changes of large amounts of genes in response to specific environmental conditions. The majority of the studies investigating gene expression changes in virus-infected plants are limited to interactions between a virus and a model host plant, which usually is Arabidopsis thaliana or Nicotiana benthamiana. In the present work, we performed microarray profiling to explore changes in the expression profile of field-grown Prunus persica (peach) originating from Chile upon single and double infection with Prunus necrotic ringspot virus (PNRSV) and Peach latent mosaic viroid (PLMVd), worldwide natural pathogens of peach trees. Upon single PLMVd or PNRSV infection, the number of statistically significant gene expression changes was relatively low. By contrast, doubly-infected fruits presented a high number of differentially regulated genes. Among these, down-regulated genes were prevalent. Functional categorization of the gene expression changes upon double PLMVd and PNRSV infection revealed protein modification and degradation as the functional category with the highest percentage of repressed genes whereas induced genes encoded mainly proteins related to phosphate, C-compound and carbohydrate metabolism and also protein modification. Overrepresentation analysis upon double infection with PLMVd and PNRSV revealed specific functional categories over- and underrepresented among the repressed genes indicating active counter-defense mechanisms of the pathogens during infection. Our results identify a novel synergistic effect of PLMVd and PNRSV on the transcriptome of peach fruits. We demonstrate that mixed infections, which occur frequently in field conditions, result in a more complex transcriptional response than that observed in single infections. Thus, our data demonstrate for the first time that the simultaneous infection of a viroid and a plant virus synergistically affect the host transcriptome in infected peach fruits. These field studies can help to fully understand plant-pathogen interactions and to develop appropriate crop protection strategies.

A remarkable synergistic effect at the transcriptomic level in peach fruits doubly infected by prunus necrotic ringspot virus and peach latent mosaic viroid

PubMed Central

2013-01-01

Background Microarray profiling is a powerful technique to investigate expression changes of large amounts of genes in response to specific environmental conditions. The majority of the studies investigating gene expression changes in virus-infected plants are limited to interactions between a virus and a model host plant, which usually is Arabidopsis thaliana or Nicotiana benthamiana. In the present work, we performed microarray profiling to explore changes in the expression profile of field-grown Prunus persica (peach) originating from Chile upon single and double infection with Prunus necrotic ringspot virus (PNRSV) and Peach latent mosaic viroid (PLMVd), worldwide natural pathogens of peach trees. Results Upon single PLMVd or PNRSV infection, the number of statistically significant gene expression changes was relatively low. By contrast, doubly-infected fruits presented a high number of differentially regulated genes. Among these, down-regulated genes were prevalent. Functional categorization of the gene expression changes upon double PLMVd and PNRSV infection revealed protein modification and degradation as the functional category with the highest percentage of repressed genes whereas induced genes encoded mainly proteins related to phosphate, C-compound and carbohydrate metabolism and also protein modification. Overrepresentation analysis upon double infection with PLMVd and PNRSV revealed specific functional categories over- and underrepresented among the repressed genes indicating active counter-defense mechanisms of the pathogens during infection. Conclusions Our results identify a novel synergistic effect of PLMVd and PNRSV on the transcriptome of peach fruits. We demonstrate that mixed infections, which occur frequently in field conditions, result in a more complex transcriptional response than that observed in single infections. Thus, our data demonstrate for the first time that the simultaneous infection of a viroid and a plant virus synergistically affect the host transcriptome in infected peach fruits. These field studies can help to fully understand plant-pathogen interactions and to develop appropriate crop protection strategies. PMID:23710752

Differential Expression Patterns in Chemosensory and Non-Chemosensory Tissues of Putative Chemosensory Genes Identified by Transcriptome Analysis of Insect Pest the Purple Stem Borer Sesamia inferens (Walker)

PubMed Central

Zhang, Ya-Nan; Jin, Jun-Yan; Jin, Rong; Xia, Yi-Han; Zhou, Jing-Jiang; Deng, Jian-Yu; Dong, Shuang-Lin

2013-01-01

Background A large number of insect chemosensory genes from different gene subfamilies have been identified and annotated, but their functional diversity and complexity are largely unknown. A systemic examination of expression patterns in chemosensory organs could provide important information. Methodology/Principal Findings We identified 92 putative chemosensory genes by analysing the transcriptome of the antennae and female sex pheromone gland of the purple stem borer Sesamia inferens, among them 87 are novel in this species, including 24 transcripts encoding for odorant binding proteins (OBPs), 24 for chemosensory proteins (CSPs), 2 for sensory neuron membrane proteins (SNMPs), 39 for odorant receptors (ORs) and 3 for ionotropic receptors (IRs). The transcriptome analyses were validated and quantified with a detailed global expression profiling by Reverse Transcription-PCR for all 92 transcripts and by Quantitative Real Time RT-PCR for selected 16 ones. Among the chemosensory gene subfamilies, CSP transcripts are most widely and evenly expressed in different tissues and stages, OBP transcripts showed a clear antenna bias and most of OR transcripts are only detected in adult antennae. Our results also revealed that some OR transcripts, such as the transcripts of SNMP2 and 2 IRs were expressed in non-chemosensory tissues, and some CSP transcripts were antenna-biased expression. Furthermore, no chemosensory transcript is specific to female sex pheromone gland and very few are found in the heads. Conclusion Our study revealed that there are a large number of chemosensory genes expressed in S. inferens, and some of them displayed unusual expression profile in non-chemosensory tissues. The identification of a large set of putative chemosensory genes of each subfamily from a single insect species, together with their different expression profiles provide further information in understanding the functions of these chemosensory genes in S. inferens as well as other insects. PMID:23894529

Differential expression patterns in chemosensory and non-chemosensory tissues of putative chemosensory genes identified by transcriptome analysis of insect pest the purple stem borer Sesamia inferens (Walker).

PubMed

Zhang, Ya-Nan; Jin, Jun-Yan; Jin, Rong; Xia, Yi-Han; Zhou, Jing-Jiang; Deng, Jian-Yu; Dong, Shuang-Lin

2013-01-01

A large number of insect chemosensory genes from different gene subfamilies have been identified and annotated, but their functional diversity and complexity are largely unknown. A systemic examination of expression patterns in chemosensory organs could provide important information. We identified 92 putative chemosensory genes by analysing the transcriptome of the antennae and female sex pheromone gland of the purple stem borer Sesamia inferens, among them 87 are novel in this species, including 24 transcripts encoding for odorant binding proteins (OBPs), 24 for chemosensory proteins (CSPs), 2 for sensory neuron membrane proteins (SNMPs), 39 for odorant receptors (ORs) and 3 for ionotropic receptors (IRs). The transcriptome analyses were validated and quantified with a detailed global expression profiling by Reverse Transcription-PCR for all 92 transcripts and by Quantitative Real Time RT-PCR for selected 16 ones. Among the chemosensory gene subfamilies, CSP transcripts are most widely and evenly expressed in different tissues and stages, OBP transcripts showed a clear antenna bias and most of OR transcripts are only detected in adult antennae. Our results also revealed that some OR transcripts, such as the transcripts of SNMP2 and 2 IRs were expressed in non-chemosensory tissues, and some CSP transcripts were antenna-biased expression. Furthermore, no chemosensory transcript is specific to female sex pheromone gland and very few are found in the heads. Our study revealed that there are a large number of chemosensory genes expressed in S. inferens, and some of them displayed unusual expression profile in non-chemosensory tissues. The identification of a large set of putative chemosensory genes of each subfamily from a single insect species, together with their different expression profiles provide further information in understanding the functions of these chemosensory genes in S. inferens as well as other insects.

Disclosure of the differences of Mesorhizobium loti under the free-living and symbiotic conditions by comparative proteome analysis without bacteroid isolation.

PubMed

Tatsukami, Yohei; Nambu, Mami; Morisaka, Hironobu; Kuroda, Kouichi; Ueda, Mitsuyoshi

2013-07-31

Rhizobia are symbiotic nitrogen-fixing soil bacteria that show a symbiotic relationship with their host legume. Rhizobia have 2 different physiological conditions: a free-living condition in soil, and a symbiotic nitrogen-fixing condition in the nodule. The lifestyle of rhizobia remains largely unknown, although genome and transcriptome analyses have been carried out. To clarify the lifestyle of bacteria, proteome analysis is necessary because the protein profile directly reflects in vivo reactions of the organisms. In proteome analysis, high separation performance is required to analyze complex biological samples. Therefore, we used a liquid chromatography-tandem mass spectrometry system, equipped with a long monolithic silica capillary column, which is superior to conventional columns. In this study, we compared the protein profile of Mesorhizobium loti MAFF303099 under free-living condition to that of symbiotic conditions by using small amounts of crude extracts. We identified 1,533 and 847 proteins for M. loti under free-living and symbiotic conditions, respectively. Pathway analysis by Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that many of the enzymes involved in the central carbon metabolic pathway were commonly detected under both conditions. The proteins encoded in the symbiosis island, the transmissible chromosomal region that includes the genes that are highly upregulated under the symbiotic condition, were uniquely detected under the symbiotic condition. The features of the symbiotic condition that have been reported by transcriptome analysis were confirmed at the protein level by proteome analysis. In addition, the genes of the proteins involved in cell surface structure were repressed under the symbiotic nitrogen-fixing condition. Furthermore, farnesyl pyrophosphate (FPP) was found to be biosynthesized only in rhizobia under the symbiotic condition. The obtained protein profile appeared to reflect the difference in phenotypes under the free-living and symbiotic conditions. In addition, KEGG pathway analysis revealed that the cell surface structure of rhizobia was largely different under each condition, and surprisingly, rhizobia might provided FPP to the host as a source of secondary metabolism. M. loti changed its metabolism and cell surface structure in accordance with the surrounding conditions.

Disclosure of the differences of Mesorhizobium loti under the free-living and symbiotic conditions by comparative proteome analysis without bacteroid isolation

PubMed Central

2013-01-01

Background Rhizobia are symbiotic nitrogen-fixing soil bacteria that show a symbiotic relationship with their host legume. Rhizobia have 2 different physiological conditions: a free-living condition in soil, and a symbiotic nitrogen-fixing condition in the nodule. The lifestyle of rhizobia remains largely unknown, although genome and transcriptome analyses have been carried out. To clarify the lifestyle of bacteria, proteome analysis is necessary because the protein profile directly reflects in vivo reactions of the organisms. In proteome analysis, high separation performance is required to analyze complex biological samples. Therefore, we used a liquid chromatography-tandem mass spectrometry system, equipped with a long monolithic silica capillary column, which is superior to conventional columns. In this study, we compared the protein profile of Mesorhizobium loti MAFF303099 under free-living condition to that of symbiotic conditions by using small amounts of crude extracts. Result We identified 1,533 and 847 proteins for M. loti under free-living and symbiotic conditions, respectively. Pathway analysis by Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that many of the enzymes involved in the central carbon metabolic pathway were commonly detected under both conditions. The proteins encoded in the symbiosis island, the transmissible chromosomal region that includes the genes that are highly upregulated under the symbiotic condition, were uniquely detected under the symbiotic condition. The features of the symbiotic condition that have been reported by transcriptome analysis were confirmed at the protein level by proteome analysis. In addition, the genes of the proteins involved in cell surface structure were repressed under the symbiotic nitrogen-fixing condition. Furthermore, farnesyl pyrophosphate (FPP) was found to be biosynthesized only in rhizobia under the symbiotic condition. Conclusion The obtained protein profile appeared to reflect the difference in phenotypes under the free-living and symbiotic conditions. In addition, KEGG pathway analysis revealed that the cell surface structure of rhizobia was largely different under each condition, and surprisingly, rhizobia might provided FPP to the host as a source of secondary metabolism. M. loti changed its metabolism and cell surface structure in accordance with the surrounding conditions. PMID:23898917

Current genetic methodologies in the identification of disaster victims and in forensic analysis.

PubMed

Ziętkiewicz, Ewa; Witt, Magdalena; Daca, Patrycja; Zebracka-Gala, Jadwiga; Goniewicz, Mariusz; Jarząb, Barbara; Witt, Michał

2012-02-01

This review presents the basic problems and currently available molecular techniques used for genetic profiling in disaster victim identification (DVI). The environmental conditions of a mass disaster often result in severe fragmentation, decomposition and intermixing of the remains of victims. In such cases, traditional identification based on the anthropological and physical characteristics of the victims is frequently inconclusive. This is the reason why DNA profiling became the gold standard for victim identification in mass-casualty incidents (MCIs) or any forensic cases where human remains are highly fragmented and/or degraded beyond recognition. The review provides general information about the sources of genetic material for DNA profiling, the genetic markers routinely used during genetic profiling (STR markers, mtDNA and single-nucleotide polymorphisms [SNP]) and the basic statistical approaches used in DNA-based disaster victim identification. Automated technological platforms that allow the simultaneous analysis of a multitude of genetic markers used in genetic identification (oligonucleotide microarray techniques and next-generation sequencing) are also presented. Forensic and population databases containing information on human variability, routinely used for statistical analyses, are discussed. The final part of this review is focused on recent developments, which offer particularly promising tools for forensic applications (mRNA analysis, transcriptome variation in individuals/populations and genetic profiling of specific cells separated from mixtures).

Selective 2′-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) for direct, versatile, and accurate RNA structure analysis

PubMed Central

Smola, Matthew J.; Rice, Greggory M.; Busan, Steven; Siegfried, Nathan A.; Weeks, Kevin M.

2016-01-01

SHAPE chemistries exploit small electrophilic reagents that react with the 2′-hydroxyl group to interrogate RNA structure at single-nucleotide resolution. Mutational profiling (MaP) identifies modified residues based on the ability of reverse transcriptase to misread a SHAPE-modified nucleotide and then counting the resulting mutations by massively parallel sequencing. The SHAPE-MaP approach measures the structure of large and transcriptome-wide systems as accurately as for simple model RNAs. This protocol describes the experimental steps, implemented over three days, required to perform SHAPE probing and construct multiplexed SHAPE-MaP libraries suitable for deep sequencing. These steps include RNA folding and SHAPE structure probing, mutational profiling by reverse transcription, library construction, and sequencing. Automated processing of MaP sequencing data is accomplished using two software packages. ShapeMapper converts raw sequencing files into mutational profiles, creates SHAPE reactivity plots, and provides useful troubleshooting information, often within an hour. SuperFold uses these data to model RNA secondary structures, identify regions with well-defined structures, and visualize probable and alternative helices, often in under a day. We illustrate these algorithms with the E. coli thiamine pyrophosphate riboswitch, E. coli 16S rRNA, and HIV-1 genomic RNAs. SHAPE-MaP can be used to make nucleotide-resolution biophysical measurements of individual RNA motifs, rare components of complex RNA ensembles, and entire transcriptomes. The straightforward MaP strategy greatly expands the number, length, and complexity of analyzable RNA structures. PMID:26426499

Multi-omic profiles of hepatic metabolism in TPN-fed preterm pigs

USDA-ARS?s Scientific Manuscript database

New generation lipid emulsions comprised of fish oil or blends of soybean/fish/medium chain triglyceride/olive oil are emerging that result in favorable clinical metabolic outcomes in pediatric populations. Our aim was to characterize the lipidodomic, metabolomic, and transcriptomic profiles these ...

Gene expression profiles of auxin metabolism in maturing apple fruit

USDA-ARS?s Scientific Manuscript database

Variation exists among apple genotypes in fruit maturation and ripening patterns that influences at-harvest fruit firmness and postharvest storability. Based on the results from our previous large-scale transcriptome profiling on apple fruit maturation and well-documented auxin-ethylene crosstalk, t...

Biochemical and transcriptomic analyses reveal different metabolite biosynthesis profiles among three color and developmental stages in 'Anji Baicha' (Camellia sinensis).

PubMed

Li, Chun-Fang; Xu, Yan-Xia; Ma, Jian-Qiang; Jin, Ji-Qiang; Huang, Dan-Juan; Yao, Ming-Zhe; Ma, Chun-Lei; Chen, Liang

2016-09-08

The new shoots of the albino tea cultivar 'Anji Baicha' are yellow or white at low temperatures and turn green as the environmental temperatures increase during the early spring. 'Anji Baicha' metabolite profiles exhibit considerable variability over three color and developmental stages, especially regarding the carotenoid, chlorophyll, and theanine concentrations. Previous studies focused on physiological characteristics, gene expression differences, and variations in metabolite abundances in albino tea plant leaves at specific growth stages. However, the molecular mechanisms regulating metabolite biosynthesis in various color and developmental stages in albino tea leaves have not been fully characterized. We used RNA-sequencing to analyze 'Anji Baicha' leaves at the yellow-green, albescent, and re-greening stages. The leaf transcriptomes differed considerably among the three stages. Functional classifications based on Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes enrichment analyses revealed that differentially expressed unigenes were mainly related to metabolic pathways, biosynthesis of secondary metabolites, phenylpropanoid biosynthesis, and carbon fixation in photosynthetic organisms. Chemical analyses revealed higher β-carotene and theanine levels, but lower chlorophyll a levels, in the albescent stage than in the green stage. Furthermore, unigenes involved in carotenoid, chlorophyll, and theanine biosyntheses were identified, and the expression patterns of the differentially expressed unigenes in these biosynthesis pathways were characterized. Through co-expression analyses, we identified the key genes in these pathways. These genes may be responsible for the metabolite biosynthesis differences among the different leaf color and developmental stages of 'Anji Baicha' tea plants. Our study presents the results of transcriptomic and biochemical analyses of 'Anji Baicha' tea plants at various stages. The distinct transcriptome profiles for each color and developmental stage enabled us to identify changes to biosynthesis pathways and revealed the contributions of such variations to the albino phenotype of tea plants. Furthermore, comparisons of the transcriptomes and related metabolites helped clarify the molecular regulatory mechanisms underlying the secondary metabolic pathways in different stages.

RBFox1-mediated RNA splicing regulates cardiac hypertrophy and heart failure.

PubMed

Gao, Chen; Ren, Shuxun; Lee, Jae-Hyung; Qiu, Jinsong; Chapski, Douglas J; Rau, Christoph D; Zhou, Yu; Abdellatif, Maha; Nakano, Astushi; Vondriska, Thomas M; Xiao, Xinshu; Fu, Xiang-Dong; Chen, Jau-Nian; Wang, Yibin

2016-01-01

RNA splicing is a major contributor to total transcriptome complexity; however, the functional role and regulation of splicing in heart failure remain poorly understood. Here, we used a total transcriptome profiling and bioinformatic analysis approach and identified a muscle-specific isoform of an RNA splicing regulator, RBFox1 (also known as A2BP1), as a prominent regulator of alternative RNA splicing during heart failure. Evaluation of developing murine and zebrafish hearts revealed that RBFox1 is induced during postnatal cardiac maturation. However, we found that RBFox1 is markedly diminished in failing human and mouse hearts. In a mouse model, RBFox1 deficiency in the heart promoted pressure overload-induced heart failure. We determined that RBFox1 is a potent regulator of RNA splicing and is required for a conserved splicing process of transcription factor MEF2 family members that yields different MEF2 isoforms with differential effects on cardiac hypertrophic gene expression. Finally, induction of RBFox1 expression in murine pressure overload models substantially attenuated cardiac hypertrophy and pathological manifestations. Together, this study identifies regulation of RNA splicing by RBFox1 as an important player in transcriptome reprogramming during heart failure that influence pathogenesis of the disease.

RBFox1-mediated RNA splicing regulates cardiac hypertrophy and heart failure

PubMed Central

Gao, Chen; Ren, Shuxun; Lee, Jae-Hyung; Qiu, Jinsong; Chapski, Douglas J.; Rau, Christoph D.; Zhou, Yu; Abdellatif, Maha; Nakano, Astushi; Vondriska, Thomas M.; Xiao, Xinshu; Fu, Xiang-Dong; Chen, Jau-Nian; Wang, Yibin

2015-01-01

RNA splicing is a major contributor to total transcriptome complexity; however, the functional role and regulation of splicing in heart failure remain poorly understood. Here, we used a total transcriptome profiling and bioinformatic analysis approach and identified a muscle-specific isoform of an RNA splicing regulator, RBFox1 (also known as A2BP1), as a prominent regulator of alternative RNA splicing during heart failure. Evaluation of developing murine and zebrafish hearts revealed that RBFox1 is induced during postnatal cardiac maturation. However, we found that RBFox1 is markedly diminished in failing human and mouse hearts. In a mouse model, RBFox1 deficiency in the heart promoted pressure overload–induced heart failure. We determined that RBFox1 is a potent regulator of RNA splicing and is required for a conserved splicing process of transcription factor MEF2 family members that yields different MEF2 isoforms with differential effects on cardiac hypertrophic gene expression. Finally, induction of RBFox1 expression in murine pressure overload models substantially attenuated cardiac hypertrophy and pathological manifestations. Together, this study identifies regulation of RNA splicing by RBFox1 as an important player in transcriptome reprogramming during heart failure that influence pathogenesis of the disease. PMID:26619120

The transcriptome of nitrofen-induced pulmonary hypoplasia in the rat model of congenital diaphragmatic hernia.

PubMed

Mahood, Thomas H; Johar, Dina R; Iwasiow, Barbara M; Xu, Wayne; Keijzer, Richard

2016-05-01

We currently do not know how the herbicide nitrofen induces lung hypoplasia and congenital diaphragmatic hernia in rats. Our aim was to compare the differentially expressed transcriptome of nitrofen-induced hypoplastic lungs to control lungs in embryonic day 13 rat embryos before the development of embryonic diaphragmatic defects. Using next-generation sequencing technology, we identified the expression profile of microRNA (miRNA) and mRNA genes. Once the dataset was validated by both RT-qPCR and digital-PCR, we conducted gene ontology, miRNA target analysis, and orthologous miRNA sequence matching for the deregulated miRNAs in silico. Our study identified 186 known mRNA and 100 miRNAs which were differentially expressed in nitrofen-induced hypoplastic lungs. Sixty-four rat miRNAs homologous to known human miRNAs were identified. A subset of these genes may promote lung hypoplasia in rat and/or human, and we discuss their associations. Potential miRNA pathways relevant to nitrofen-induced lung hypoplasia include PI3K, TGF-β, and cell cycle kinases. Nitrofen-induced hypoplastic lungs have an abnormal transcriptome that may lead to impaired development.

Integrative Transcriptome Profiling of Cognitive Aging and Its Preservation through Ser/Thr Protein Phosphatase Regulation.

PubMed

Park, C Sehwan; Valomon, Amandine; Welzl, Hans

2015-01-01

Environmental enrichment has been reported to delay or restore age-related cognitive deficits, however, a mechanism to account for the cause and progression of normal cognitive decline and its preservation by environmental enrichment is lacking. Using genome-wide SAGE-Seq, we provide a global assessment of differentially expressed genes altered with age and environmental enrichment in the hippocampus. Qualitative and quantitative proteomics in naïve young and aged mice was used to further identify phosphorylated proteins differentially expressed with age. We found that increased expression of endogenous protein phosphatase-1 inhibitors in aged mice may be characteristic of long-term environmental enrichment and improved cognitive status. As such, hippocampus-dependent performances in spatial, recognition, and associative memories, which are sensitive to aging, were preserved by environmental enrichment and accompanied by decreased protein phosphatase activity. Age-associated phosphorylated proteins were also found to correspond to the functional categories of age-associated genes identified through transcriptome analysis. Together, this study provides a comprehensive map of the transcriptome and proteome in the aging brain, and elucidates endogenous protein phosphatase-1 inhibition as a potential means through which environmental enrichment may ameliorate age-related cognitive deficits.

Transcriptome profiles of embryos before and after cleavage in Eriocheir sinensis: identification of developmental genes at the earliest stages

NASA Astrophysics Data System (ADS)

Hui, Min; Cui, Zhaoxia; Liu, Yuan; Song, Chengwen

2017-07-01

In crab, embryogenesis is a complicated developmental program marked by a series of critical events. RNA-Sequencing technology offers developmental biologists a way to identify many more developmental genes than ever before. Here, we present a comprehensive analysis of the transcriptomes of Eriocheir sinensis oosperms (Os) and embryos at the 2-4 cell stage (Cs), which are separated by a cleavage event. A total of 18 923 unigenes were identified, and 403 genes matched with gene ontology (GO) terms related to developmental processes. In total, 432 differentially expressed genes (DEGs) were detected between the two stages. Nine DEGs were specifically expressed at only one stage. These DEGs may be relevant to stage-specific molecular events during development. A number of DEGs related to `hedgehog signaling pathway', `Wnt signaling pathway' `germplasm', `nervous system', `sensory perception' and `segment polarity' were identified as being up-regulated at the Cs stage. The results suggest that these embryonic developmental events begin before the early cleavage event in crabs, and that many of the genes expressed in the two transcriptomes might be maternal genes. Our study provides ample information for further research on the molecular mechanisms underlying crab development.

Spliced synthetic genes as internal controls in RNA sequencing experiments.

PubMed

Hardwick, Simon A; Chen, Wendy Y; Wong, Ted; Deveson, Ira W; Blackburn, James; Andersen, Stacey B; Nielsen, Lars K; Mattick, John S; Mercer, Tim R

2016-09-01

RNA sequencing (RNA-seq) can be used to assemble spliced isoforms, quantify expressed genes and provide a global profile of the transcriptome. However, the size and diversity of the transcriptome, the wide dynamic range in gene expression and inherent technical biases confound RNA-seq analysis. We have developed a set of spike-in RNA standards, termed 'sequins' (sequencing spike-ins), that represent full-length spliced mRNA isoforms. Sequins have an entirely artificial sequence with no homology to natural reference genomes, but they align to gene loci encoded on an artificial in silico chromosome. The combination of multiple sequins across a range of concentrations emulates alternative splicing and differential gene expression, and it provides scaling factors for normalization between samples. We demonstrate the use of sequins in RNA-seq experiments to measure sample-specific biases and determine the limits of reliable transcript assembly and quantification in accompanying human RNA samples. In addition, we have designed a complementary set of sequins that represent fusion genes arising from rearrangements of the in silico chromosome to aid in cancer diagnosis. RNA sequins provide a qualitative and quantitative reference with which to navigate the complexity of the human transcriptome.

Cross-omics comparison of stress responses in mesothelial cells exposed to heat- versus filter-sterilized peritoneal dialysis fluids.

PubMed

Kratochwill, Klaus; Bender, Thorsten O; Lichtenauer, Anton M; Herzog, Rebecca; Tarantino, Silvia; Bialas, Katarzyna; Jörres, Achim; Aufricht, Christoph

2015-01-01

Recent research suggests that cytoprotective responses, such as expression of heat-shock proteins, might be inadequately induced in mesothelial cells by heat-sterilized peritoneal dialysis (PD) fluids. This study compares transcriptome data and multiple protein expression profiles for providing new insight into regulatory mechanisms. Two-dimensional difference gel electrophoresis (2D-DIGE) based proteomics and topic defined gene expression microarray-based transcriptomics techniques were used to evaluate stress responses in human omental peritoneal mesothelial cells in response to heat- or filter-sterilized PD fluids. Data from selected heat-shock proteins were validated by 2D western-blot analysis. Comparison of proteomics and transcriptomics data discriminated differentially regulated protein abundance into groups depending on correlating or noncorrelating transcripts. Inadequate abundance of several heat-shock proteins following exposure to heat-sterilized PD fluids is not reflected on the mRNA level indicating interference beyond transcriptional regulation. For the first time, this study describes evidence for posttranscriptional inadequacy of heat-shock protein expression by heat-sterilized PD fluids as a novel cytotoxic property. Cross-omics technologies introduce a novel way of understanding PDF bioincompatibility and searching for new interventions to reestablish adequate cytoprotective responses.