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Sample records for scale transcriptome analysis

  1. Large-scale transcriptome analysis reveals arabidopsis metabolic pathways are frequently influenced by different pathogens.

    PubMed

    Jiang, Zhenhong; He, Fei; Zhang, Ziding

    2017-07-01

    Through large-scale transcriptional data analyses, we highlighted the importance of plant metabolism in plant immunity and identified 26 metabolic pathways that were frequently influenced by the infection of 14 different pathogens. Reprogramming of plant metabolism is a common phenomenon in plant defense responses. Currently, a large number of transcriptional profiles of infected tissues in Arabidopsis (Arabidopsis thaliana) have been deposited in public databases, which provides a great opportunity to understand the expression patterns of metabolic pathways during plant defense responses at the systems level. Here, we performed a large-scale transcriptome analysis based on 135 previously published expression samples, including 14 different pathogens, to explore the expression pattern of Arabidopsis metabolic pathways. Overall, metabolic genes are significantly changed in expression during plant defense responses. Upregulated metabolic genes are enriched on defense responses, and downregulated genes are enriched on photosynthesis, fatty acid and lipid metabolic processes. Gene set enrichment analysis (GSEA) identifies 26 frequently differentially expressed metabolic pathways (FreDE_Paths) that are differentially expressed in more than 60% of infected samples. These pathways are involved in the generation of energy, fatty acid and lipid metabolism as well as secondary metabolite biosynthesis. Clustering analysis based on the expression levels of these 26 metabolic pathways clearly distinguishes infected and control samples, further suggesting the importance of these metabolic pathways in plant defense responses. By comparing with FreDE_Paths from abiotic stresses, we find that the expression patterns of 26 FreDE_Paths from biotic stresses are more consistent across different infected samples. By investigating the expression correlation between transcriptional factors (TFs) and FreDE_Paths, we identify several notable relationships. Collectively, the current study

  2. Genome-Scale Transcriptome Analysis in Response to Nitric Oxide in Birch Cells: Implications of the Triterpene Biosynthetic Pathway

    PubMed Central

    Zeng, Fansuo; Sun, Fengkun; Li, Leilei; Liu, Kun; Zhan, Yaguang

    2014-01-01

    Evidence supporting nitric oxide (NO) as a mediator of plant biochemistry continues to grow, but its functions at the molecular level remains poorly understood and, in some cases, controversial. To study the role of NO at the transcriptional level in Betula platyphylla cells, we conducted a genome-scale transcriptome analysis of these cells. The transcriptome of untreated birch cells and those treated by sodium nitroprusside (SNP) were analyzed using the Solexa sequencing. Data were collected by sequencing cDNA libraries of birch cells, which had a long period to adapt to the suspension culture conditions before SNP-treated cells and untreated cells were sampled. Among the 34,100 UniGenes detected, BLASTX search revealed that 20,631 genes showed significant (E-values≤10−5) sequence similarity with proteins from the NR-database. Numerous expressed sequence tags (i.e., 1374) were identified as differentially expressed between the 12 h SNP-treated cells and control cells samples: 403 up-regulated and 971 down-regulated. From this, we specifically examined a core set of NO-related transcripts. The altered expression levels of several transcripts, as determined by transcriptome analysis, was confirmed by qRT-PCR. The results of transcriptome analysis, gene expression quantification, the content of triterpenoid and activities of defensive enzymes elucidated NO has a significant effect on many processes including triterpenoid production, carbohydrate metabolism and cell wall biosynthesis. PMID:25551661

  3. Transcriptome analysis of the Chinese white wax scale Ericerus pela with focus on genes involved in wax biosynthesis.

    PubMed

    Yang, Pu; Zhu, Jia-Ying; Gong, Zhong-Jun; Xu, Dong-Li; Chen, Xiao-Ming; Liu, Wei-Wei; Lin, Xin-Da; Li, Yan-Fei

    2012-01-01

    The Chinese white wax scale, Ericerus pela Chavannes is economically significant for its role in wax production. This insect has been bred in China for over a thousand years. The wax secreted by the male scale insect during the second-instar larval stage has been widespread used in wax candle production, wax printing, engraving, Chinese medicine, and more recently in the chemical, pharmaceutical, food, and cosmetics industries. However, little is known about the mechanisms responsible for white wax biosynthesis. The characterization of its larval transcriptome may promote better understanding of wax biosynthesis. In this study, characterization of the transcriptome of E. pela during peak wax secretion was performed using Illumina sequencing technology. Illumina sequencing produced 41,839 unigenes. These unigenes were annotated by blastx alignment against the NCBI Non-Redundant (NR), Swiss-Prot, KEGG, and COG databases. A total of 104 unigenes related to white wax biosynthesis were identified, and 15 of them were selected for quantitative real-time PCR analysis. We evaluated the variations in gene expression across different development stages, including egg, first/second instar larvae, male pupae, and male and female adults. Then we identified five genes involved in white wax biosynthesis. These genes were expressed most strongly during the second-instar larval stage of male E. pela. The transcriptome analysis of E. pela during peak wax secretion provided an overview of gene expression information at the transcriptional level and a resource for gene mining. Five genes related to white wax biosynthesis were identified.

  4. Large-scale Gene Ontology analysis of plant transcriptome-derived sequences retrieved by AFLP technology

    PubMed Central

    Botton, Alessandro; Galla, Giulio; Conesa, Ana; Bachem, Christian; Ramina, Angelo; Barcaccia, Gianni

    2008-01-01

    Background After 10-year-use of AFLP (Amplified Fragment Length Polymorphism) technology for DNA fingerprinting and mRNA profiling, large repertories of genome- and transcriptome-derived sequences are available in public databases for model, crop and tree species. AFLP marker systems have been and are being extensively exploited for genome scanning and gene mapping, as well as cDNA-AFLP for transcriptome profiling and differentially expressed gene cloning. The evaluation, annotation and classification of genomic markers and expressed transcripts would be of great utility for both functional genomics and systems biology research in plants. This may be achieved by means of the Gene Ontology (GO), consisting in three structured vocabularies (i.e. ontologies) describing genes, transcripts and proteins of any organism in terms of their associated cellular component, biological process and molecular function in a species-independent manner. In this paper, the functional annotation of about 8,000 AFLP-derived ESTs retrieved in the NCBI databases was carried out by using GO terminology. Results Descriptive statistics on the type, size and nature of gene sequences obtained by means of AFLP technology were calculated. The gene products associated with mRNA transcripts were then classified according to the three main GO vocabularies. A comparison of the functional content of cDNA-AFLP records was also performed by splitting the sequence dataset into monocots and dicots and by comparing them to all annotated ESTs of Arabidopsis and rice, respectively. On the whole, the statistical parameters adopted for the in silico AFLP-derived transcriptome-anchored sequence analysis proved to be critical for obtaining reliable GO results. Such an exhaustive annotation may offer a suitable platform for functional genomics, particularly useful in non-model species. Conclusion Reliable GO annotations of AFLP-derived sequences can be gathered through the optimization of the experimental steps

  5. Genome-scale transcriptome analysis of the desert poplar, Populus euphratica.

    PubMed

    Qiu, Qiang; Ma, Tao; Hu, Quanjun; Liu, Bingbing; Wu, Yuxia; Zhou, Haihong; Wang, Qian; Wang, Juan; Liu, Jianquan

    2011-04-01

    Populus euphratica is well-adapted to extreme desert environments and is an important model species for studying the effects of abiotic stresses on trees. Here we present the first deep transcriptomic analysis of this species. To maximize representation of conditional transcripts, mRNA was obtained from living tissues of desert-grown trees and two types of callus (salt-stressed and unstressed). De novo assembly generated 86,777 Unigenes using Solexa sequence data. These sequences covered 92% of previously reported P. euphratica expressed sequence tags (ESTs) and 90% of the TIGR poplar ESTs, and a total of 58,499 high-quality unique sequences were annotated by BLAST similarity searches against public databases. We found that 27% of the total Unigenes were differentially expressed (up- or down-regulated) in response to salt stress in P. euphratica callus. These differentially expressed genes are mainly involved in transport, transcription, cellular communication and metabolism. In addition, we found that numerous putative genes involved in ABA regulation and biosynthesis were also differentially regulated. This study represents the deepest transcriptomic and gene-annotation analysis of P. euphratica to date. The genetic knowledge acquired should be very useful for future studies of the molecular adaptation of this tree species to abiotic stress and facilitate genetic manipulation of other poplar species.

  6. A new RNASeq-based reference transcriptome for sugar beet and its application in transcriptome-scale analysis of vernalization and gibberellin responses

    PubMed Central

    2012-01-01

    Background Sugar beet (Beta vulgaris sp. vulgaris) crops account for about 30% of world sugar. Sugar yield is compromised by reproductive growth hence crops must remain vegetative until harvest. Prolonged exposure to cold temperature (vernalization) in the range 6°C to 12°C induces reproductive growth, leading to bolting (rapid elongation of the main stem) and flowering. Spring cultivation of crops in cool temperate climates makes them vulnerable to vernalization and hence bolting, which is initiated in the apical shoot meristem in processes involving interaction between gibberellin (GA) hormones and vernalization. The underlying mechanisms are unknown and genome scale next generation sequencing approaches now offer comprehensive strategies to investigate them; enabling the identification of novel targets for bolting control in sugar beet crops. In this study, we demonstrate the application of an mRNA-Seq based strategy for this purpose. Results There is no sugar beet reference genome, or public expression array platforms. We therefore used RNA-Seq to generate the first reference transcriptome. We next performed digital gene expression profiling using shoot apex mRNA from two sugar beet cultivars with and without applied GA, and also a vernalized cultivar with and without applied GA. Subsequent bioinformatics analyses identified transcriptional changes associated with genotypic difference and experimental treatments. Analysis of expression profiles in response to vernalization and GA treatment suggested previously unsuspected roles for a RAV1-like AP2/B3 domain protein in vernalization and efflux transporters in the GA response. Conclusions Next generation RNA-Seq enabled the generation of the first reference transcriptome for sugar beet and the study of global transcriptional responses in the shoot apex to vernalization and GA treatment, without the need for a reference genome or established array platforms. Comprehensive bioinformatic analysis identified

  7. Transcriptome Analysis of the Chinese White Wax Scale Ericerus pela with Focus on Genes Involved in Wax Biosynthesis

    PubMed Central

    Gong, Zhong-Jun; Xu, Dong-Li; Chen, Xiao-Ming; Liu, Wei-Wei; Lin, Xin-Da; Li, Yan-Fei

    2012-01-01

    Background The Chinese white wax scale, Ericerus pela Chavannes is economically significant for its role in wax production. This insect has been bred in China for over a thousand years. The wax secreted by the male scale insect during the second-instar larval stage has been widespread used in wax candle production, wax printing, engraving, Chinese medicine, and more recently in the chemical, pharmaceutical, food, and cosmetics industries. However, little is known about the mechanisms responsible for white wax biosynthesis. The characterization of its larval transcriptome may promote better understanding of wax biosynthesis. Methodology/Principal Findings In this study, characterization of the transcriptome of E. pela during peak wax secretion was performed using Illumina sequencing technology. Illumina sequencing produced 41,839 unigenes. These unigenes were annotated by blastx alignment against the NCBI Non-Redundant (NR), Swiss-Prot, KEGG, and COG databases. A total of 104 unigenes related to white wax biosynthesis were identified, and 15 of them were selected for quantitative real-time PCR analysis. We evaluated the variations in gene expression across different development stages, including egg, first/second instar larvae, male pupae, and male and female adults. Then we identified five genes involved in white wax biosynthesis. These genes were expressed most strongly during the second-instar larval stage of male E. pela. Conclusion/Significance The transcriptome analysis of E. pela during peak wax secretion provided an overview of gene expression information at the transcriptional level and a resource for gene mining. Five genes related to white wax biosynthesis were identified. PMID:22536429

  8. Genome-Scale Transcriptome Analysis of the Desert Shrub Artemisia sphaerocephala

    PubMed Central

    Zhang, Lijing; Hu, Xiaowei; Miao, Xiumei; Chen, Xiaolong; Nan, Shuzhen; Fu, Hua

    2016-01-01

    Background Artemisia sphaerocephala, a semi-shrub belonging to the Artemisia genus of the Compositae family, is an important pioneer plant that inhabits moving and semi-stable sand dunes in the deserts and steppes of northwest and north-central China. It is very resilient in extreme environments. Additionally, its seeds have excellent nutritional value, and the abundant lipids and polysaccharides in the seeds make this plant a potential valuable source of bio-energy. However, partly due to the scarcity of genetic information, the genetic mechanisms controlling the traits and environmental adaptation capacity of A. sphaerocephala are unknown. Results Here, we present the first in-depth transcriptomic analysis of A. sphaerocephala. To maximize the representation of conditional transcripts, mRNA was obtained from 17 samples, including living tissues of desert-growing A. sphaerocephala, seeds germinated in the laboratory, and calli subjected to no stress (control) and high and low temperature, high and low osmotic, and salt stresses. De novo transcriptome assembly performed using an Illumina HiSeq 2500 platform resulted in the generation of 68,373 unigenes. We analyzed the key genes involved in the unsaturated fatty acid synthesis pathway and identified 26 A. sphaerocephala fad2 genes, which is the largest fad2 gene family reported to date. Furthermore, a set of genes responsible for resistance to extreme temperatures, salt, drought and a combination of stresses was identified. Conclusion The present work provides abundant genomic information for functional dissection of the important traits of A. sphaerocephala and contributes to the current understanding of molecular adaptive mechanisms of A. sphaerocephala in the desert environment. Identification of the key genes in the unsaturated fatty acid synthesis pathway could increase understanding of the biological regulatory mechanisms of fatty acid composition traits in plants and facilitate genetic manipulation of the

  9. Genome-Scale Transcriptome Analysis of the Desert Shrub Artemisia sphaerocephala.

    PubMed

    Zhang, Lijing; Hu, Xiaowei; Miao, Xiumei; Chen, Xiaolong; Nan, Shuzhen; Fu, Hua

    2016-01-01

    Artemisia sphaerocephala, a semi-shrub belonging to the Artemisia genus of the Compositae family, is an important pioneer plant that inhabits moving and semi-stable sand dunes in the deserts and steppes of northwest and north-central China. It is very resilient in extreme environments. Additionally, its seeds have excellent nutritional value, and the abundant lipids and polysaccharides in the seeds make this plant a potential valuable source of bio-energy. However, partly due to the scarcity of genetic information, the genetic mechanisms controlling the traits and environmental adaptation capacity of A. sphaerocephala are unknown. Here, we present the first in-depth transcriptomic analysis of A. sphaerocephala. To maximize the representation of conditional transcripts, mRNA was obtained from 17 samples, including living tissues of desert-growing A. sphaerocephala, seeds germinated in the laboratory, and calli subjected to no stress (control) and high and low temperature, high and low osmotic, and salt stresses. De novo transcriptome assembly performed using an Illumina HiSeq 2500 platform resulted in the generation of 68,373 unigenes. We analyzed the key genes involved in the unsaturated fatty acid synthesis pathway and identified 26 A. sphaerocephala fad2 genes, which is the largest fad2 gene family reported to date. Furthermore, a set of genes responsible for resistance to extreme temperatures, salt, drought and a combination of stresses was identified. The present work provides abundant genomic information for functional dissection of the important traits of A. sphaerocephala and contributes to the current understanding of molecular adaptive mechanisms of A. sphaerocephala in the desert environment. Identification of the key genes in the unsaturated fatty acid synthesis pathway could increase understanding of the biological regulatory mechanisms of fatty acid composition traits in plants and facilitate genetic manipulation of the fatty acid composition of oil

  10. Transcriptome Analysis of Kiwifruit (Actinidia chinensis) Bark in Response to Armoured Scale Insect (Hemiberlesia lataniae) Feeding

    PubMed Central

    Hill, M. Garry; Wurms, Kirstin V.; Davy, Marcus W.; Gould, Elaine; Allan, Andrew; Mauchline, Nicola A.; Luo, Zhiwei; Ah Chee, Annette; Stannard, Kate; Storey, Roy D.; Rikkerink, Erik H.

    2015-01-01

    The kiwifruit cultivar Actinidia chinensis ‘Hort16A’ is resistant to the polyphagous armoured scale insect pest Hemiberlesia lataniae (Hemiptera: Diaspididae). A cDNA microarray consisting of 17,512 unigenes selected from over 132,000 expressed sequence tags (ESTs) was used to measure the transcriptomic profile of the A. chinensis ‘Hort16A’ canes in response to a controlled infestation of H. lataniae. After 2 days, 272 transcripts were differentially expressed. After 7 days, 5,284 (30%) transcripts were differentially expressed. The transcripts were grouped into 22 major functional categories using MapMan software. After 7 days, transcripts associated with photosynthesis (photosystem II) were significantly down-regulated, while those associated with secondary metabolism were significantly up-regulated. A total of 643 transcripts associated with response to stress were differentially expressed. This included biotic stress-related transcripts orthologous with pathogenesis related proteins, the phenylpropanoid pathway, NBS-LRR (R) genes, and receptor-like kinase–leucine rich repeat signalling proteins. While transcriptional studies are not conclusive in their own right, results were suggestive of a defence response involving both ETI and PTI, with predominance of the SA signalling pathway. Exogenous application of an SA-mimic decreased H. lataniae growth on A. chinensis ‘Hort16A’ plants in two laboratory experiments. PMID:26571404

  11. Inferring the choreography of parental genomes during fertilization from ultralarge-scale whole-transcriptome analysis.

    PubMed

    Park, Sung-Joon; Komata, Makiko; Inoue, Fukashi; Yamada, Kaori; Nakai, Kenta; Ohsugi, Miho; Shirahige, Katsuhiko

    2013-12-15

    Fertilization precisely choreographs parental genomes by using gamete-derived cellular factors and activating genome regulatory programs. However, the mechanism remains elusive owing to the technical difficulties of preparing large numbers of high-quality preimplantation cells. Here, we collected >14 × 10(4) high-quality mouse metaphase II oocytes and used these to establish detailed transcriptional profiles for four early embryo stages and parthenogenetic development. By combining these profiles with other public resources, we found evidence that gene silencing appeared to be mediated in part by noncoding RNAs and that this was a prerequisite for post-fertilization development. Notably, we identified 817 genes that were differentially expressed in embryos after fertilization compared with parthenotes. The regulation of these genes was distinctly different from those expressed in parthenotes, suggesting functional specialization of particular transcription factors prior to first cell cleavage. We identified five transcription factors that were potentially necessary for developmental progression: Foxd1, Nkx2-5, Sox18, Myod1, and Runx1. Our very large-scale whole-transcriptome profile of early mouse embryos yielded a novel and valuable resource for studies in developmental biology and stem cell research. The database is available at http://dbtmee.hgc.jp.

  12. Transcriptome Analysis of Kiwifruit (Actinidia chinensis) Bark in Response to Armoured Scale Insect (Hemiberlesia lataniae) Feeding.

    PubMed

    Hill, M Garry; Wurms, Kirstin V; Davy, Marcus W; Gould, Elaine; Allan, Andrew; Mauchline, Nicola A; Luo, Zhiwei; Ah Chee, Annette; Stannard, Kate; Storey, Roy D; Rikkerink, Erik H

    2015-01-01

    The kiwifruit cultivar Actinidia chinensis 'Hort16A' is resistant to the polyphagous armoured scale insect pest Hemiberlesia lataniae (Hemiptera: Diaspididae). A cDNA microarray consisting of 17,512 unigenes selected from over 132,000 expressed sequence tags (ESTs) was used to measure the transcriptomic profile of the A. chinensis 'Hort16A' canes in response to a controlled infestation of H. lataniae. After 2 days, 272 transcripts were differentially expressed. After 7 days, 5,284 (30%) transcripts were differentially expressed. The transcripts were grouped into 22 major functional categories using MapMan software. After 7 days, transcripts associated with photosynthesis (photosystem II) were significantly down-regulated, while those associated with secondary metabolism were significantly up-regulated. A total of 643 transcripts associated with response to stress were differentially expressed. This included biotic stress-related transcripts orthologous with pathogenesis related proteins, the phenylpropanoid pathway, NBS-LRR (R) genes, and receptor-like kinase-leucine rich repeat signalling proteins. While transcriptional studies are not conclusive in their own right, results were suggestive of a defence response involving both ETI and PTI, with predominance of the SA signalling pathway. Exogenous application of an SA-mimic decreased H. lataniae growth on A. chinensis 'Hort16A' plants in two laboratory experiments.

  13. Optimal Scaling of Digital Transcriptomes

    PubMed Central

    Glusman, Gustavo; Caballero, Juan; Robinson, Max; Kutlu, Burak; Hood, Leroy

    2013-01-01

    Deep sequencing of transcriptomes has become an indispensable tool for biology, enabling expression levels for thousands of genes to be compared across multiple samples. Since transcript counts scale with sequencing depth, counts from different samples must be normalized to a common scale prior to comparison. We analyzed fifteen existing and novel algorithms for normalizing transcript counts, and evaluated the effectiveness of the resulting normalizations. For this purpose we defined two novel and mutually independent metrics: (1) the number of “uniform” genes (genes whose normalized expression levels have a sufficiently low coefficient of variation), and (2) low Spearman correlation between normalized expression profiles of gene pairs. We also define four novel algorithms, one of which explicitly maximizes the number of uniform genes, and compared the performance of all fifteen algorithms. The two most commonly used methods (scaling to a fixed total value, or equalizing the expression of certain ‘housekeeping’ genes) yielded particularly poor results, surpassed even by normalization based on randomly selected gene sets. Conversely, seven of the algorithms approached what appears to be optimal normalization. Three of these algorithms rely on the identification of “ubiquitous” genes: genes expressed in all the samples studied, but never at very high or very low levels. We demonstrate that these include a “core” of genes expressed in many tissues in a mutually consistent pattern, which is suitable for use as an internal normalization guide. The new methods yield robustly normalized expression values, which is a prerequisite for the identification of differentially expressed and tissue-specific genes as potential biomarkers. PMID:24223126

  14. Transcriptome analysis of tube foot and large scale marker discovery in sea cucumber, Apostichopus japonicus.

    PubMed

    Zhou, Xiaoxu; Wang, Hongdi; Cui, Jun; Qiu, Xuemei; Chang, Yaqing; Wang, Xiuli

    2016-12-01

    Tube foot as one of the ambulacral appendages types in Aspidochirote holothurioids, is known for their functions in locomotion, feeding, chemoreception, light sensitivity and respiration. In this study, we explored the characteristic of transcriptome in the tube foot of sea cucumber (Apostichopus japonicus). Our results showed that among 390 unigenes which specifically expressed in the tube foot, 190 of them were annotated. Based on the assembly transcriptome, we found 219,860 SNPs from 34,749 unigenes, 97,683, 53,624, 27,767 and 40,786 were located in CDSs, 5'-UTRs, 3'-UTRs and non-CDS separately. Furthermore, 12,114 SSRs were detected from 7394 unigenes. Target genes of four specifically expressed miRNAs (miR-29a, miR-29b, miR-278-3p and miR-2005) in tube foot were also predicted based on the transcriptome, which contain immune-related factors (MBL, VLRA, AjC3, MyD88, CFB), skin pigmentation (MITF), candidate regeneration factor (TRP) and holothurians autolysis-related factor (CL). These results develop a relatively large number of molecular markers and transcriptome resources, and will provide a foundation for further analyses on the function and molecular mechanisms underlying A. japonicas tube foot.

  15. Parallel analysis of Arabidopsis circadian clock mutants reveals different scales of transcriptome and proteome regulation

    PubMed Central

    Graf, Alexander; Coman, Diana; Walsh, Sean; Flis, Anna; Stitt, Mark; Gruissem, Wilhelm

    2017-01-01

    The circadian clock regulates physiological processes central to growth and survival. To date, most plant circadian clock studies have relied on diurnal transcriptome changes to elucidate molecular connections between the circadian clock and observable phenotypes in wild-type plants. Here, we have integrated RNA-sequencing and protein mass spectrometry data to comparatively analyse the lhycca1, prr7prr9, gi and toc1 circadian clock mutant rosette at the end of day and end of night. Each mutant affects specific sets of genes and proteins, suggesting that the circadian clock regulation is modular. Furthermore, each circadian clock mutant maintains its own dynamically fluctuating transcriptome and proteome profile specific to subcellular compartments. Most of the measured protein levels do not correlate with changes in their corresponding transcripts. Transcripts and proteins that have coordinated changes in abundance are enriched for carbohydrate- and cold-responsive genes. Transcriptome changes in all four circadian clock mutants also affect genes encoding starch degradation enzymes, transcription factors and protein kinases. The comprehensive transcriptome and proteome datasets demonstrate that future system-driven research of the circadian clock requires multi-level experimental approaches. Our work also shows that further work is needed to elucidate the roles of post-translational modifications and protein degradation in the regulation of clock-related processes. PMID:28250106

  16. Parallel analysis of Arabidopsis circadian clock mutants reveals different scales of transcriptome and proteome regulation.

    PubMed

    Graf, Alexander; Coman, Diana; Uhrig, R Glen; Walsh, Sean; Flis, Anna; Stitt, Mark; Gruissem, Wilhelm

    2017-03-01

    The circadian clock regulates physiological processes central to growth and survival. To date, most plant circadian clock studies have relied on diurnal transcriptome changes to elucidate molecular connections between the circadian clock and observable phenotypes in wild-type plants. Here, we have integrated RNA-sequencing and protein mass spectrometry data to comparatively analyse the lhycca1, prr7prr9, gi and toc1 circadian clock mutant rosette at the end of day and end of night. Each mutant affects specific sets of genes and proteins, suggesting that the circadian clock regulation is modular. Furthermore, each circadian clock mutant maintains its own dynamically fluctuating transcriptome and proteome profile specific to subcellular compartments. Most of the measured protein levels do not correlate with changes in their corresponding transcripts. Transcripts and proteins that have coordinated changes in abundance are enriched for carbohydrate- and cold-responsive genes. Transcriptome changes in all four circadian clock mutants also affect genes encoding starch degradation enzymes, transcription factors and protein kinases. The comprehensive transcriptome and proteome datasets demonstrate that future system-driven research of the circadian clock requires multi-level experimental approaches. Our work also shows that further work is needed to elucidate the roles of post-translational modifications and protein degradation in the regulation of clock-related processes. © 2017 The Authors.

  17. Large-scale identification of wheat genes resistant to cereal cyst nematode Heterodera avenae using comparative transcriptomic analysis.

    PubMed

    Kong, Ling-An; Wu, Du-Qing; Huang, Wen-Kun; Peng, Huan; Wang, Gao-Feng; Cui, Jiang-Kuan; Liu, Shi-Ming; Li, Zhi-Gang; Yang, Jun; Peng, De-Liang

    2015-10-16

    , phospholipase was involved in many defense-related pathways and localized in the connection position. Furthermore, strong bursts of reactive oxygen species (ROS) within VP1620 roots infected with H. avenae were induced at 24 h and 3 d, and eight ROS-producing genes were significantly upregulated, including three class III peroxidase and five lipoxygenase genes. Large-scale identification of wheat resistance genes were processed by comparative transcriptomic analysis. Functional characterization showed that phospholipases associated with ROS production played vital roles in early defense responses to H. avenae via involvement in diverse defense-related pathways as a hub switch. This study is the first to investigate the early defense responses of wheat against H. avenae, not only provides applicable candidate resistance genes for breeding novel wheat cultivars, but also enables a better understanding of the defense mechanisms of wheat against H. avenae.

  18. Large-Scale Transcriptome Analysis in Faba Bean (Vicia faba L.) under Ascochyta fabae Infection

    PubMed Central

    Ocaña, Sara; Seoane, Pedro; Bautista, Rocio; Palomino, Carmen; Claros, Gonzalo M.; Torres, Ana M.; Madrid, Eva

    2015-01-01

    Faba bean is an important food crop worldwide. However, progress in faba bean genomics lags far behind that of model systems due to limited availability of genetic and genomic information. Using the Illumina platform the faba bean transcriptome from leaves of two lines (29H and Vf136) subjected to Ascochyta fabae infection have been characterized. De novo transcriptome assembly provided a total of 39,185 different transcripts that were functionally annotated, and among these, 13,266 were assigned to gene ontology against Arabidopsis. Quality of the assembly was validated by RT-qPCR amplification of selected transcripts differentially expressed. Comparison of faba bean transcripts with those of better-characterized plant genomes such as Arabidopsis thaliana, Medicago truncatula and Cicer arietinum revealed a sequence similarity of 68.3%, 72.8% and 81.27%, respectively. Moreover, 39,060 single nucleotide polymorphism (SNP) and 3,669 InDels were identified for genotyping applications. Mapping of the sequence reads generated onto the assembled transcripts showed that 393 and 457 transcripts were overexpressed in the resistant (29H) and susceptible genotype (Vf136), respectively. Transcripts involved in plant-pathogen interactions such as leucine rich proteins (LRR) or plant growth regulators involved in plant adaptation to abiotic and biotic stresses were found to be differently expressed in the resistant line. The results reported here represent the most comprehensive transcript database developed so far in faba bean, providing valuable information that could be used to gain insight into the pathways involved in the resistance mechanism against A. fabae and to identify potential resistance genes to be further used in marker assisted selection. PMID:26267359

  19. Large-Scale Transcriptome Analysis in Faba Bean (Vicia faba L.) under Ascochyta fabae Infection.

    PubMed

    Ocaña, Sara; Seoane, Pedro; Bautista, Rocio; Palomino, Carmen; Claros, Gonzalo M; Torres, Ana M; Madrid, Eva

    2015-01-01

    Faba bean is an important food crop worldwide. However, progress in faba bean genomics lags far behind that of model systems due to limited availability of genetic and genomic information. Using the Illumina platform the faba bean transcriptome from leaves of two lines (29H and Vf136) subjected to Ascochyta fabae infection have been characterized. De novo transcriptome assembly provided a total of 39,185 different transcripts that were functionally annotated, and among these, 13,266 were assigned to gene ontology against Arabidopsis. Quality of the assembly was validated by RT-qPCR amplification of selected transcripts differentially expressed. Comparison of faba bean transcripts with those of better-characterized plant genomes such as Arabidopsis thaliana, Medicago truncatula and Cicer arietinum revealed a sequence similarity of 68.3%, 72.8% and 81.27%, respectively. Moreover, 39,060 single nucleotide polymorphism (SNP) and 3,669 InDels were identified for genotyping applications. Mapping of the sequence reads generated onto the assembled transcripts showed that 393 and 457 transcripts were overexpressed in the resistant (29H) and susceptible genotype (Vf136), respectively. Transcripts involved in plant-pathogen interactions such as leucine rich proteins (LRR) or plant growth regulators involved in plant adaptation to abiotic and biotic stresses were found to be differently expressed in the resistant line. The results reported here represent the most comprehensive transcript database developed so far in faba bean, providing valuable information that could be used to gain insight into the pathways involved in the resistance mechanism against A. fabae and to identify potential resistance genes to be further used in marker assisted selection.

  20. Large-Scale Transcriptome Analysis of Two Sugarcane Genotypes Contrasting for Lignin Content

    PubMed Central

    Vicentini, Renato; Bottcher, Alexandra; Brito, Michael dos Santos; dos Santos, Adriana Brombini; Creste, Silvana; Landell, Marcos Guimarães de Andrade; Cesarino, Igor; Mazzafera, Paulo

    2015-01-01

    Sugarcane is an important crop worldwide for sugar and first generation ethanol production. Recently, the residue of sugarcane mills, named bagasse, has been considered a promising lignocellulosic biomass to produce the second-generation ethanol. Lignin is a major factor limiting the use of bagasse and other plant lignocellulosic materials to produce second-generation ethanol. Lignin biosynthesis pathway is a complex network and changes in the expression of genes of this pathway have in general led to diverse and undesirable impacts on plant structure and physiology. Despite its economic importance, sugarcane genome was still not sequenced. In this study a high-throughput transcriptome evaluation of two sugarcane genotypes contrasting for lignin content was carried out. We generated a set of 85,151 transcripts of sugarcane using RNA-seq and de novo assembling. More than 2,000 transcripts showed differential expression between the genotypes, including several genes involved in the lignin biosynthetic pathway. This information can give valuable knowledge on the lignin biosynthesis and its interactions with other metabolic pathways in the complex sugarcane genome. PMID:26241317

  1. Genome scale transcriptomics of baculovirus-insect interactions.

    PubMed

    Nguyen, Quan; Nielsen, Lars K; Reid, Steven

    2013-11-12

    Baculovirus-insect cell technologies are applied in the production of complex proteins, veterinary and human vaccines, gene delivery vectors' and biopesticides. Better understanding of how baculoviruses and insect cells interact would facilitate baculovirus-based production. While complete genomic sequences are available for over 58 baculovirus species, little insect genomic information is known. The release of the Bombyx mori and Plutella xylostella genomes, the accumulation of EST sequences for several Lepidopteran species, and especially the availability of two genome-scale analysis tools, namely oligonucleotide microarrays and next generation sequencing (NGS), have facilitated expression studies to generate a rich picture of insect gene responses to baculovirus infections. This review presents current knowledge on the interaction dynamics of the baculovirus-insect system' which is relatively well studied in relation to nucleocapsid transportation, apoptosis, and heat shock responses, but is still poorly understood regarding responses involved in pro-survival pathways, DNA damage pathways, protein degradation, translation, signaling pathways, RNAi pathways, and importantly metabolic pathways for energy, nucleotide and amino acid production. We discuss how the two genome-scale transcriptomic tools can be applied for studying such pathways and suggest that proteomics and metabolomics can produce complementary findings to transcriptomic studies.

  2. Integrated Analysis of Transcriptomic and Proteomic Data

    PubMed Central

    Haider, Saad; Pal, Ranadip

    2013-01-01

    Until recently, understanding the regulatory behavior of cells has been pursued through independent analysis of the transcriptome or the proteome. Based on the central dogma, it was generally assumed that there exist a direct correspondence between mRNA transcripts and generated protein expressions. However, recent studies have shown that the correlation between mRNA and Protein expressions can be low due to various factors such as different half lives and post transcription machinery. Thus, a joint analysis of the transcriptomic and proteomic data can provide useful insights that may not be deciphered from individual analysis of mRNA or protein expressions. This article reviews the existing major approaches for joint analysis of transcriptomic and proteomic data. We categorize the different approaches into eight main categories based on the initial algorithm and final analysis goal. We further present analogies with other domains and discuss the existing research problems in this area. PMID:24082820

  3. Transcriptome Analysis of Sexually Dimorphic Chinese White Wax Scale Insects Reveals Key Differences in Developmental Programs and Transcription Factor Expression

    PubMed Central

    Yang, Pu; Chen, Xiao-Ming; Liu, Wei-Wei; Feng, Ying; Sun, Tao

    2015-01-01

    The Chinese white wax scale insect, Ericerus pela, represents one of the most dramatic examples of sexual dimorphism in any insect species. In this study, we showed that although E. pela males display complete metamorphosis similar to holometabolous insects, the species forms the sister group to Acyrthosiphon pisum and cluster with hemimetabolous insects. The gene expression profile and Gene Ontology (GO) analyses revealed that the two sexes engaged in distinct developmental programs. In particular, female development appeared to prioritize the expression of genes related to cellular, metabolic, and developmental processes and to anatomical structure formation in nymphs. By contrast, male nymphal development is characterized by the significant down-regulation of genes involved in chitin, the respiratory system, and neurons. The wing and appendage morphogenesis, anatomical and tissue structure morphogenesis programs activated after male nymphal development. Transcription factors (that convey juvenile hormone or ecdysone signals, and Hox genes) and DNA methyltransferase were also differentially expressed between females and males. These results may indicate the roles that these differentially expressed genes play in regulating sexual dimorphism through orchestrating complex genetic programs. This differential expression was particularly prominent for processes linked to female development and wing development in males. PMID:25634031

  4. Transcriptome analysis of sexually dimorphic Chinese white wax scale insects reveals key differences in developmental programs and transcription factor expression.

    PubMed

    Yang, Pu; Chen, Xiao-Ming; Liu, Wei-Wei; Feng, Ying; Sun, Tao

    2015-01-30

    The Chinese white wax scale insect, Ericerus pela, represents one of the most dramatic examples of sexual dimorphism in any insect species. In this study, we showed that although E. pela males display complete metamorphosis similar to holometabolous insects, the species forms the sister group to Acyrthosiphon pisum and cluster with hemimetabolous insects. The gene expression profile and Gene Ontology (GO) analyses revealed that the two sexes engaged in distinct developmental programs. In particular, female development appeared to prioritize the expression of genes related to cellular, metabolic, and developmental processes and to anatomical structure formation in nymphs. By contrast, male nymphal development is characterized by the significant down-regulation of genes involved in chitin, the respiratory system, and neurons. The wing and appendage morphogenesis, anatomical and tissue structure morphogenesis programs activated after male nymphal development. Transcription factors (that convey juvenile hormone or ecdysone signals, and Hox genes) and DNA methyltransferase were also differentially expressed between females and males. These results may indicate the roles that these differentially expressed genes play in regulating sexual dimorphism through orchestrating complex genetic programs. This differential expression was particularly prominent for processes linked to female development and wing development in males.

  5. Multi-Scale Genomic, Transcriptomic and Proteomic Analysis of Colorectal Cancer Cell Lines to Identify Novel Biomarkers

    PubMed Central

    Briffa, Romina; Um, Inhwa; Faratian, Dana; Zhou, Ying; Turnbull, Arran K.; Langdon, Simon P.; Harrison, David J.

    2015-01-01

    Selecting colorectal cancer (CRC) patients likely to respond to therapy remains a clinical challenge. The objectives of this study were to establish which genes were differentially expressed with respect to treatment sensitivity and relate this to copy number in a panel of 15 CRC cell lines. Copy number variations of the identified genes were assessed in a cohort of CRCs. IC50’s were measured for 5-fluorouracil, oxaliplatin, and BEZ-235, a PI3K/mTOR inhibitor. Cell lines were profiled using array comparative genomic hybridisation, Illumina gene expression analysis, reverse phase protein arrays, and targeted sequencing of KRAS hotspot mutations. Frequent gains were observed at 2p, 3q, 5p, 7p, 7q, 8q, 12p, 13q, 14q, and 17q and losses at 2q, 3p, 5q, 8p, 9p, 9q, 14q, 18q, and 20p. Frequently gained regions contained EGFR, PIK3CA, MYC, SMO, TRIB1, FZD1, and BRCA2, while frequently lost regions contained FHIT and MACROD2. TRIB1 was selected for further study. Gene enrichment analysis showed that differentially expressed genes with respect to treatment response were involved in Wnt signalling, EGF receptor signalling, apoptosis, cell cycle, and angiogenesis. Stepwise integration of copy number and gene expression data yielded 47 candidate genes that were significantly correlated. PDCD6 was differentially expressed in all three treatment responses. Tissue microarrays were constructed for a cohort of 118 CRC patients and TRIB1 and MYC amplifications were measured using fluorescence in situ hybridisation. TRIB1 and MYC were amplified in 14.5% and 7.4% of the cohort, respectively, and these amplifications were significantly correlated (p≤0.0001). TRIB1 protein expression in the patient cohort was significantly correlated with pERK, Akt, and Caspase 3 expression. In conclusion, a set of candidate predictive biomarkers for 5-fluorouracil, oxaliplatin, and BEZ235 are described that warrant further study. Amplification of the putative oncogene TRIB1 has been described for

  6. Domestication causes large-scale effects on gene expression in rainbow trout: analysis of muscle, liver and brain transcriptomes.

    PubMed

    Tymchuk, Wendy; Sakhrani, Dionne; Devlin, Robert

    2009-01-01

    Domestication has produced faster-growing strains of animals for use in agriculture, but selection has been applied with little knowledge of the underlying genetic changes that arose throughout the process. Mammals and birds have been domesticated for thousands of years whereas fish have been domesticated only recently; therefore, wild progenitor strains remain for comparison. Rainbow trout (Oncorhynchus mykiss) have undergone intensive selection and domesticated strains grow more rapidly than extant wild strains. To assess physiological pathways altered by domestication, whole-genome mRNA expression was measured in brain, muscle and liver of size-matched domestic and wild trout using a 16K (cGRASP) salmonid microarray. A large number of genes differed between strains, ranging from 3% of genes in brain to 9% in muscle. Domestic fish had more down-regulated genes in the brain relative to wild fish, whereas more genes were up-regulated in domestic liver and muscle. Relative to wild fish, there was a down-regulation of cell division and an up-regulation of structural genes in the brain of domestic fish. In liver from domestic fish, there was an up-regulation of genes related to transport with a down-regulation of lipid binding. Analysis of the functional categories for muscle indicated that most pathways, including pathways related to metabolism and catabolism, were up-regulated in domestic fish. Comparison of these results to other genomic studies on transgenic, domestic and wild salmonids suggests that similar physiological pathways are altered systemically to support faster rates of growth, regardless of the underlying genetic alteration that has caused the altered growth.

  7. Comparative Transcriptome Analysis of Four Prymnesiophyte Algae

    PubMed Central

    Koid, Amy E.; Liu, Zhenfeng; Terrado, Ramon; Jones, Adriane C.; Caron, David A.; Heidelberg, Karla B.

    2014-01-01

    Genomic studies of bacteria, archaea and viruses have provided insights into the microbial world by unveiling potential functional capabilities and molecular pathways. However, the rate of discovery has been slower among microbial eukaryotes, whose genomes are larger and more complex. Transcriptomic approaches provide a cost-effective alternative for examining genetic potential and physiological responses of microbial eukaryotes to environmental stimuli. In this study, we generated and compared the transcriptomes of four globally-distributed, bloom-forming prymnesiophyte algae: Prymnesium parvum, Chrysochromulina brevifilum, Chrysochromulina ericina and Phaeocystis antarctica. Our results revealed that the four transcriptomes possess a set of core genes that are similar in number and shared across all four organisms. The functional classifications of these core genes using the euKaryotic Orthologous Genes (KOG) database were also similar among the four study organisms. More broadly, when the frequencies of different cellular and physiological functions were compared with other protists, the species clustered by both phylogeny and nutritional modes. Thus, these clustering patterns provide insight into genomic factors relating to both evolutionary relationships as well as trophic ecology. This paper provides a novel comparative analysis of the transcriptomes of ecologically important and closely related prymnesiophyte protists and advances an emerging field of study that uses transcriptomics to reveal ecology and function in protists. PMID:24926657

  8. 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.

  9. Transcriptome analysis of Ginkgo biloba kernels

    PubMed Central

    He, Bing; Gu, Yincong; Xu, Meng; Wang, Jianwen; Cao, Fuliang; Xu, Li-an

    2015-01-01

    Ginkgo biloba is a dioecious species native to China with medicinally and phylogenetically important characteristics; however, genomic resources for this species are limited. In this study, we performed the first transcriptome sequencing for Ginkgo kernels at five time points using Illumina paired-end sequencing. Approximately 25.08-Gb clean reads were obtained, and 68,547 unigenes with an average length of 870 bp were generated by de novo assembly. Of these unigenes, 29,987 (43.74%) were annotated in publicly available plant protein database. A total of 3,869 genes were identified as significantly differentially expressed, and enrichment analysis was conducted at different time points. Furthermore, metabolic pathway analysis revealed that 66 unigenes were responsible for terpenoid backbone biosynthesis, with up to 12 up-regulated unigenes involved in the biosynthesis of ginkgolide and bilobalide. Differential gene expression analysis together with real-time PCR experiments indicated that the synthesis of bilobalide may have interfered with the ginkgolide synthesis process in the kernel. These data can remarkably expand the existing transcriptome resources of Ginkgo, and provide a valuable platform to reveal more on developmental and metabolic mechanisms of this species. PMID:26500663

  10. Comparative analysis of de novo transcriptome assembly.

    PubMed

    Clarke, Kaitlin; Yang, Yi; Marsh, Ronald; Xie, Linglin; Zhang, Ke K

    2013-02-01

    The fast development of next-generation sequencing technology presents a major computational challenge for data processing and analysis. A fast algorithm, de Bruijn graph has been successfully used for genome DNA de novo assembly; nevertheless, its performance for transcriptome assembly is unclear. In this study, we used both simulated and real RNA-Seq data, from either artificial RNA templates or human transcripts, to evaluate five de novo assemblers, ABySS, Mira, Trinity, Velvet and Oases. Of these assemblers, ABySS, Trinity, Velvet and Oases are all based on de Bruijn graph, and Mira uses an overlap graph algorithm. Various numbers of RNA short reads were selected from the External RNA Control Consortium (ERCC) data and human chromosome 22. A number of statistics were then calculated for the resulting contigs from each assembler. Each experiment was repeated multiple times to obtain the mean statistics and standard error estimate. Trinity had relative good performance for both ERCC and human data, but it may not consistently generate full length transcripts. ABySS was the fastest method but its assembly quality was low. Mira gave a good rate for mapping its contigs onto human chromosome 22, but its computational speed is not satisfactory. Our results suggest that transcript assembly remains a challenge problem for bioinformatics society. Therefore, a novel assembler is in need for assembling transcriptome data generated by next generation sequencing technique.

  11. Integrative analysis of the mouse embryonic transcriptome

    PubMed Central

    Singh, Amar V; Knudsen, Kenneth B; Knudsen, Thomas B

    2007-01-01

    Monitoring global gene expression provides insight into how genes and regulatory signals work together to guide embryo development. The fields of developmental biology and teratology are now confronted with the need for automated access to a reference library of gene-expression signatures that benchmark programmed (genetic) and adaptive (environmental) regulation of the embryonic transcriptome. Such a library must be constructed from highly-distributed microarray data. Birth Defects Systems Manager (BDSM), an open access knowledge management system, provides custom software to mine public microarray data focused on developmental health and disease. The present study describes tools for seamless data integration in the BDSM library (MetaSample, MetaChip, CIAeasy) using the QueryBDSM module. A field test of the prototype was run using published microarray data series derived from a variety of laboratories, experiments, microarray platforms, organ systems, and developmental stages. The datasets focused on several developing systems in the mouse embryo, including preimplantation stages, heart and nerve development, testis and ovary development, and craniofacial development. Using BDSM data integration tools, a gene-expression signature for 346 genes was resolved that accurately classified samples by organ system and developmental sequence. The module builds a potential for the BDSM approach to decipher a large number developmental processes through comparative bioinformatics analysis of embryological systems at-risk for specific defects, using multiple scenarios to define the range of probabilities leading from molecular phenotype to clinical phenotype. We conclude that an integrative analysis of global gene-expression of the developing embryo can form the foundation for constructing a reference library of signaling pathways and networks for normal and abnormal regulation of the embryonic transcriptome. These tools are available free of charge from the web-site http

  12. Integrative analysis of the mouse embryonic transcriptome.

    PubMed

    Singh, Amar V; Knudsen, Kenneth B; Knudsen, Thomas B

    2007-04-10

    Monitoring global gene expression provides insight into how genes and regulatory signals work together to guide embryo development. The fields of developmental biology and teratology are now confronted with the need for automated access to a reference library of gene-expression signatures that benchmark programmed (genetic) and adaptive (environmental) regulation of the embryonic transcriptome. Such a library must be constructed from highly-distributed microarray data. Birth Defects Systems Manager (BDSM), an open access knowledge management system, provides custom software to mine public microarray data focused on developmental health and disease. The present study describes tools for seamless data integration in the BDSM library (MetaSample, MetaChip, CIAeasy) using the QueryBDSM module. A field test of the prototype was run using published microarray data series derived from a variety of laboratories, experiments, microarray platforms, organ systems, and developmental stages. The datasets focused on several developing systems in the mouse embryo, including preimplantation stages, heart and nerve development, testis and ovary development, and craniofacial development. Using BDSM data integration tools, a gene-expression signature for 346 genes was resolved that accurately classified samples by organ system and developmental sequence. The module builds a potential for the BDSM approach to decipher a large number developmental processes through comparative bioinformatics analysis of embryological systems at-risk for specific defects, using multiple scenarios to define the range of probabilities leading from molecular phenotype to clinical phenotype. We conclude that an integrative analysis of global gene-expression of the developing embryo can form the foundation for constructing a reference library of signaling pathways and networks for normal and abnormal regulation of the embryonic transcriptome. These tools are available free of charge from the web-site http

  13. Large-scale RNA-Seq Transcriptome Analysis of 4043 Cancers and 548 Normal Tissue Controls across 12 TCGA Cancer Types

    PubMed Central

    Peng, Li; Bian, Xiu Wu; Li, Di Kang; Xu, Chuan; Wang, Guang Ming; Xia, Qing You; Xiong, Qing

    2015-01-01

    The Cancer Genome Atlas (TCGA) has accrued RNA-Seq-based transcriptome data for more than 4000 cancer tissue samples across 12 cancer types, translating these data into biological insights remains a major challenge. We analyzed and compared the transcriptomes of 4043 cancer and 548 normal tissue samples from 21 TCGA cancer types, and created a comprehensive catalog of gene expression alterations for each cancer type. By clustering genes into co-regulated gene sets, we identified seven cross-cancer gene signatures altered across a diverse panel of primary human cancer samples. A 14-gene signature extracted from these seven cross-cancer gene signatures precisely differentiated between cancerous and normal samples, the predictive accuracy of leave-one-out cross-validation (LOOCV) were 92.04%, 96.23%, 91.76%, 90.05%, 88.17%, 94.29%, and 99.10% for BLCA, BRCA, COAD, HNSC, LIHC, LUAD, and LUSC, respectively. A lung cancer-specific gene signature, containing SFTPA1 and SFTPA2 genes, accurately distinguished lung cancer from other cancer samples, the predictive accuracy of LOOCV for TCGA and GSE5364 data were 95.68% and 100%, respectively. These gene signatures provide rich insights into the transcriptional programs that trigger tumorigenesis and metastasis, and many genes in the signature gene panels may be of significant value to the diagnosis and treatment of cancer. PMID:26292924

  14. Applying thiouracil tagging to mouse transcriptome analysis.

    PubMed

    Gay, Leslie; Karfilis, Kate V; Miller, Michael R; Doe, Chris Q; Stankunas, Kryn

    2014-02-01

    Transcriptional profiling is a powerful approach for studying mouse development, physiology and disease models. Here we describe a protocol for mouse thiouracil tagging (TU tagging), a transcriptome analysis technology that includes in vivo covalent labeling, purification and analysis of cell type-specific RNA. TU tagging enables the isolation of RNA from a given cell population of a complex tissue, avoiding transcriptional changes induced by cell isolation trauma, as well as the identification of actively transcribed RNAs and not preexisting transcripts. Therefore, in contrast to other cell-specific transcriptional profiling methods based on the purification of tagged ribosomes or nuclei, TU tagging provides a direct examination of transcriptional regulation. We describe how to (i) deliver 4-thiouracil to transgenic mice to thio-label cell lineage-specific transcripts, (ii) purify TU-tagged RNA and prepare libraries for Illumina sequencing and (iii) follow a straightforward bioinformatics workflow to identify cell type-enriched or differentially expressed genes. Tissue containing TU-tagged RNA can be obtained in 1 d, RNA-seq libraries can be generated within 2 d and, after sequencing, an initial bioinformatics analysis can be completed in 1 additional day.

  15. Differential transcriptomic analysis reveals hidden light response in Streptomyces lividans.

    PubMed

    Koepff, Joachim; Morschett, Holger; Busche, Tobias; Winkler, Anika; Kalinowski, Jörn; Wiechert, Wolfgang; Oldiges, Marco

    2017-09-27

    Recently, a comprehensive screening workflow for the filamentous bacterium Streptomyces lividans, a highly performant source for pharmaceutically active agents was introduced. This framework used parallelized cultivation in microtiter plates to efficiently accelerate early upstream process development. Focusing on growth performance, cultivation was successfully scaled-up to 1 liter stirred tank reactors. However, metabolic adaptation was observed on the transcriptomic level as among others, several genes incorporated in light response were up-regulated during bioreactor cultivation. Despite it was assumed that this was attributed to the fact that reactor cultivations were carried out in glass vessels exposed to daylight and artificial room light, this setup did not allow distinguishing exclusively between light and other effects. Upon that, the present study directly investigates the influence of light by defined illumination of microtiter plate cultures. Almost identical growth performance was observed for cultures grown in the dark or with illumination. Transcriptomics revealed the up-regulation of seven genes of which 6 have previously been described to be relevant for carotenoid synthesis and its regulation. These pigments are effective quenchers of reactive oxygen species. The seventh transcript coded for a photo-lyase incorporated in UV-damage repair of DNA further confirming induced light response. However, this was fully compensated by metabolic adaptation on the transcriptomic level and overall process performance was maintained. Consequently, environmental conditions need extremely careful control and evaluation during in-depth omics analysis of bioprocesses. Otherwise metabolic adaptation induced by such issues can easily be misinterpreted, especially during studies addressing cultivation system comparisons. This article is protected by copyright. All rights reserved. © 2017 American Institute of Chemical Engineers.

  16. Transcriptomic analysis of degraded forensic body fluids.

    PubMed

    Lin, Meng-Han; Jones, Daniel F; Fleming, Rachel

    2015-07-01

    Massively parallel sequencing (MPS) has facilitated a significant increase in transcriptomic studies in all biological disciplines. However, the analysis of degraded RNA remains a genuine challenge in practice. In forensic science the biological samples encountered are often extensively degraded and of low abundance. RNA from these compromised samples is used for body fluid identification through the detection of body fluid-specific transcripts. Here we demonstrate the sequencing of four forensically relevant body fluids: oral mucosa/saliva (buccal), circulatory blood, menstrual blood and vaginal fluid. RNA was extracted from fresh, two and six week aged samples. Despite the extensive degradation of most body fluids, significant high quality sequencing output (>80% sequence above Q30) was generated. An average of over 80% of reads from all but one sample aligned successfully to the reference human genome. Furthermore, FPKMs (fragments per kilobase of exon per million fragments mapped) generated indicate the accurate detection of known body fluid markers in respective body fluids. Assessment of global gene expression levels over degradation time enabled the characterisation of differential RNA degradation in different body fluids. This study demonstrates the practical application of MPS technology for the accurate analysis of degraded RNA from minimal samples.

  17. Transcriptome Analysis of Zebrafish Embryogenesis Using Microarrays

    PubMed Central

    Mathavan, Sinnakaruppan; Lee, Serene G. P; Mak, Alicia; Miller, Lance D; Murthy, Karuturi Radha Krishna; Govindarajan, Kunde R; Tong, Yan; Wu, Yi Lian; Lam, Siew Hong; Yang, Henry; Ruan, Yijun; Korzh, Vladimir; Gong, Zhiyuan; Liu, Edison T; Lufkin, Thomas

    2005-01-01

    Zebrafish (Danio rerio) is a well-recognized model for the study of vertebrate developmental genetics, yet at the same time little is known about the transcriptional events that underlie zebrafish embryogenesis. Here we have employed microarray analysis to study the temporal activity of developmentally regulated genes during zebrafish embryogenesis. Transcriptome analysis at 12 different embryonic time points covering five different developmental stages (maternal, blastula, gastrula, segmentation, and pharyngula) revealed a highly dynamic transcriptional profile. Hierarchical clustering, stage-specific clustering, and algorithms to detect onset and peak of gene expression revealed clearly demarcated transcript clusters with maximum gene activity at distinct developmental stages as well as co-regulated expression of gene groups involved in dedicated functions such as organogenesis. Our study also revealed a previously unidentified cohort of genes that are transcribed prior to the mid-blastula transition, a time point earlier than when the zygotic genome was traditionally thought to become active. Here we provide, for the first time to our knowledge, a comprehensive list of developmentally regulated zebrafish genes and their expression profiles during embryogenesis, including novel information on the temporal expression of several thousand previously uncharacterized genes. The expression data generated from this study are accessible to all interested scientists from our institute resource database (http://giscompute.gis.a-star.edu.sg/~govind/zebrafish/data_download.html). PMID:16132083

  18. Transcriptome Analysis of Encystation in Entamoeba invadens

    PubMed Central

    Nakada-Tsukui, Kumiko; Caler, Elisabet; Nozaki, Tomoyoshi

    2013-01-01

    Encystation is an essential differentiation process for the completion of the life cycle of a group of intestinal protozoa including Entamoeba histolytica, the causative agent of intestinal and extraintestinal amebiasis. However, regulation of gene expression during encystation is poorly understood. To comprehensively understand the process at the molecular level, the transcriptomic profiles of E. invadens, which is a related reptilian species that causes an invasive disease similar to that of E. histolytica, was investigated during encystation. Using a custom-generated Affymetrix platform microarray, we performed time course (0.5, 2, 8, 24, 48, and 120 h) gene expression analysis of encysting E. invadens. ANOVA analysis revealed that a total of 1,528 genes showed ≥3 fold up-regulation at one or more time points, relative to the trophozoite stage. Of these modulated genes, 8% (116 genes) were up-regulated at the early time points (0.5, 2 and 8h), while 63% (962 genes) were up-regulated at the later time points (24, 48, and 120 h). Twenty nine percent (450 genes) are either up-regulated at 2 to 5 time points or constitutively up-regulated in both early and late stages. Among the up-regulated genes are the genes encoding transporters, cytoskeletal proteins, proteins involved in vesicular trafficking (small GTPases), Myb transcription factors, cysteine proteases, components of the proteasome, and enzymes for chitin biosynthesis. This study represents the first kinetic analysis of gene expression during differentiation from the invasive trophozoite to the dormant, infective cyst stage in Entamoeba. Functional analysis on individual genes and their encoded products that are modulated during encystation may lead to the discovery of targets for the development of new chemotherapeutics that interfere with stage conversion of the parasite. PMID:24040350

  19. Transcriptome analysis of encystation in Entamoeba invadens.

    PubMed

    De Cádiz, Aleyla Escueta; Jeelani, Ghulam; Nakada-Tsukui, Kumiko; Caler, Elisabet; Nozaki, Tomoyoshi

    2013-01-01

    Encystation is an essential differentiation process for the completion of the life cycle of a group of intestinal protozoa including Entamoeba histolytica, the causative agent of intestinal and extraintestinal amebiasis. However, regulation of gene expression during encystation is poorly understood. To comprehensively understand the process at the molecular level, the transcriptomic profiles of E. invadens, which is a related reptilian species that causes an invasive disease similar to that of E. histolytica, was investigated during encystation. Using a custom-generated Affymetrix platform microarray, we performed time course (0.5, 2, 8, 24, 48, and 120 h) gene expression analysis of encysting E. invadens. ANOVA analysis revealed that a total of 1,528 genes showed ≥3 fold up-regulation at one or more time points, relative to the trophozoite stage. Of these modulated genes, 8% (116 genes) were up-regulated at the early time points (0.5, 2 and 8h), while 63% (962 genes) were up-regulated at the later time points (24, 48, and 120 h). Twenty nine percent (450 genes) are either up-regulated at 2 to 5 time points or constitutively up-regulated in both early and late stages. Among the up-regulated genes are the genes encoding transporters, cytoskeletal proteins, proteins involved in vesicular trafficking (small GTPases), Myb transcription factors, cysteine proteases, components of the proteasome, and enzymes for chitin biosynthesis. This study represents the first kinetic analysis of gene expression during differentiation from the invasive trophozoite to the dormant, infective cyst stage in Entamoeba. Functional analysis on individual genes and their encoded products that are modulated during encystation may lead to the discovery of targets for the development of new chemotherapeutics that interfere with stage conversion of the parasite.

  20. Integrated analysis of the Plasmodium species transcriptome.

    PubMed

    Hoo, Regina; Zhu, Lei; Amaladoss, Anburaj; Mok, Sachel; Natalang, Onguma; Lapp, Stacey A; Hu, Guangan; Liew, Kingsley; Galinski, Mary R; Bozdech, Zbynek; Preiser, Peter R

    2016-05-01

    The genome sequence available for different Plasmodium species is a valuable resource for understanding malaria parasite biology. However, comparative genomics on its own cannot fully explain all the species-specific differences which suggests that other genomic aspects such as regulation of gene expression play an important role in defining species-specific characteristics. Here, we developed a comprehensive approach to measure transcriptional changes of the evolutionary conserved syntenic orthologs during the intraerythrocytic developmental cycle across six Plasmodium species. We show significant transcriptional constraint at the mid-developmental stage of Plasmodium species while the earliest stages of parasite development display the greatest transcriptional variation associated with critical functional processes. Modeling of the evolutionary relationship based on changes in transcriptional profile reveal a phylogeny pattern of the Plasmodium species that strictly follows its mammalian hosts. In addition, the work shows that transcriptional conserved orthologs represent potential future targets for anti-malaria intervention as they would be expected to carry out key essential functions within the parasites. This work provides an integrated analysis of orthologous transcriptome, which aims to provide insights into the Plasmodium evolution thereby establishing a framework to explore complex pathways and drug discovery in Plasmodium species with broad host range. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Transcriptomic Analysis of Mouse Cochlear Supporting Cell Maturation Reveals Large-Scale Changes in Notch Responsiveness Prior to the Onset of Hearing

    PubMed Central

    Maass, Juan C.; Gu, Rende; Cai, Tiantian; Wan, Ying-Wooi; Cantellano, Silvia C.; Asprer, Joanna S. T.; Zhang, Hongyuan; Jen, Hsin-I; Edlund, Renée K.; Liu, Zhandong; Groves, Andrew K.

    2016-01-01

    Neonatal mouse cochlear supporting cells have a limited ability to divide and trans-differentiate into hair cells, but this ability declines rapidly in the two weeks after birth. This decline is concomitant with the morphological and functional maturation of the organ of Corti prior to the onset of hearing. However, despite this association between maturation and loss of regenerative potential, little is known of the molecular changes that underlie these events. To identify these changes, we used RNA-seq to generate transcriptional profiles of purified cochlear supporting cells from 1- and 6-day-old mice. We found many significant changes in gene expression during this period, many of which were related to regulation of proliferation, differentiation of inner ear components and the maturation of the organ of Corti prior to the onset of hearing. One example of a change in regenerative potential of supporting cells is their robust production of hair cells in response to a blockade of the Notch signaling pathway at the time of birth, but a complete lack of response to such blockade just a few days later. By comparing our supporting cell transcriptomes to those of supporting cells cultured in the presence of Notch pathway inhibitors, we show that the transcriptional response to Notch blockade disappears almost completely in the first postnatal week. Our results offer some of the first molecular insights into the failure of hair cell regeneration in the mammalian cochlea. PMID:27918591

  2. Transcriptomic Analysis of Grazing by Marine Nanoflagellates

    NASA Astrophysics Data System (ADS)

    Palenik, B.; Shi, X.; Paz-Yepes, J.

    2016-02-01

    Synechococcus is an important contributor to coastal and open ocean primary productivity. It is thought to be consumed by heterotrophic nanoflagellates and ciliates. To better understand if such predators adapt to the consumption of different prey, we obtained transcriptomes from five nanoflagellate grazers provided with or without Synechococcus prey. Transcriptomes were sequenced as part of the Moore Foundation Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) program. All transcriptomes showed differential gene expression in the presence of Synechococcus CC9311. No genes were found to be up-regulated in all five grazers, but 18 genes were up-regulated more than two fold in at least three grazers grown with CC9311, and two of the 18 genes were up-regulated in four grazers. These were a c-type lectin-like gene and an ankyrin repeat gene and the respective proteins would be likely candidates for Synechococcus binding/recognition by grazers. These results will likely help to interpret metatranscriptome studies in marine environments.

  3. Novel software package for cross-platform transcriptome analysis (CPTRA)

    PubMed Central

    2009-01-01

    Background Next-generation sequencing techniques enable several novel transcriptome profiling approaches. Recent studies indicated that digital gene expression profiling based on short sequence tags has superior performance as compared to other transcriptome analysis platforms including microarrays. However, the transcriptomic analysis with tag-based methods often depends on available genome sequence. The use of tag-based methods in species without genome sequence should be complemented by other methods such as cDNA library sequencing. The combination of different next generation sequencing techniques like 454 pyrosequencing and Illumina Genome Analyzer (Solexa) will enable high-throughput and accurate global gene expression profiling in species with limited genome information. The combination of transcriptome data acquisition methods requires cross-platform transcriptome data analysis platforms, including a new software package for data processing. Results Here we presented a software package, CPTRA: Cross-Platform TRanscriptome Analysis, to analyze transcriptome profiling data from separate methods. The software package is available at http://people.tamu.edu/~syuan/cptra/cptra.html. It was applied to the case study of non-target site glyphosate resistance in horseweed; and the data was mined to discover resistance target gene(s). For the software, the input data included a long-read sequence dataset with proper annotation, and a short-read sequence tag dataset for the quantification of transcripts. By combining the two datasets, the software carries out the unique sequence tag identification, tag counting for transcript quantification, and cross-platform sequence matching functions, whereby the short sequence tags can be annotated with a function, level of expression, and Gene Ontology (GO) classification. Multiple sequence search algorithms were implemented and compared. The analysis highlighted the importance of transport genes in glyphosate resistance and

  4. Transcriptome sequencing and comparative transcriptome analysis of the scleroglucan producer Sclerotium rolfsii

    PubMed Central

    2010-01-01

    Background The plant pathogenic basidiomycete Sclerotium rolfsii produces the industrially exploited exopolysaccharide scleroglucan, a polymer that consists of (1 → 3)-β-linked glucose with a (1 → 6)-β-glycosyl branch on every third unit. Although the physicochemical properties of scleroglucan are well understood, almost nothing is known about the genetics of scleroglucan biosynthesis. Similarly, the biosynthetic pathway of oxalate, the main by-product during scleroglucan production, has not been elucidated yet. In order to provide a basis for genetic and metabolic engineering approaches, we studied scleroglucan and oxalate biosynthesis in S. rolfsii using different transcriptomic approaches. Results Two S. rolfsii transcriptomes obtained from scleroglucan-producing and scleroglucan-nonproducing conditions were pooled and sequenced using the 454 pyrosequencing technique yielding ~350,000 reads. These could be assembled into 21,937 contigs and 171,833 singletons, for which 6,951 had significant matches in public protein data bases. Sequence data were used to obtain first insights into the genomics of scleroglucan and oxalate production and to predict putative proteins involved in the synthesis of both metabolites. Using comparative transcriptomics, namely Agilent microarray hybridization and suppression subtractive hybridization, we identified ~800 unigenes which are differently expressed under scleroglucan-producing and non-producing conditions. From these, candidate genes were identified which could represent potential leads for targeted modification of the S. rolfsii metabolism for increased scleroglucan yields. Conclusions The results presented in this paper provide for the first time genomic and transcriptomic data about S. rolfsii and demonstrate the power and usefulness of combined transcriptome sequencing and comparative microarray analysis. The data obtained allowed us to predict the biosynthetic pathways of scleroglucan and oxalate synthesis and to

  5. Analysis of the Citrullus colocynthis Transcriptome during Water Deficit Stress

    PubMed Central

    Wang, Zhuoyu; Hu, Hongtao; Goertzen, Leslie R.; McElroy, J. Scott; Dane, Fenny

    2014-01-01

    Citrullus colocynthis is a very drought tolerant species, closely related to watermelon (C. lanatus var. lanatus), an economically important cucurbit crop. Drought is a threat to plant growth and development, and the discovery of drought inducible genes with various functions is of great importance. We used high throughput mRNA Illumina sequencing technology and bioinformatic strategies to analyze the C. colocynthis leaf transcriptome under drought treatment. Leaf samples at four different time points (0, 24, 36, or 48 hours of withholding water) were used for RNA extraction and Illumina sequencing. qRT-PCR of several drought responsive genes was performed to confirm the accuracy of RNA sequencing. Leaf transcriptome analysis provided the first glimpse of the drought responsive transcriptome of this unique cucurbit species. A total of 5038 full-length cDNAs were detected, with 2545 genes showing significant changes during drought stress. Principle component analysis indicated that drought was the major contributing factor regulating transcriptome changes. Up regulation of many transcription factors, stress signaling factors, detoxification genes, and genes involved in phytohormone signaling and citrulline metabolism occurred under the water deficit conditions. The C. colocynthis transcriptome data highlight the activation of a large set of drought related genes in this species, thus providing a valuable resource for future functional analysis of candidate genes in defense of drought stress. PMID:25118696

  6. Analysis of the Citrullus colocynthis transcriptome during water deficit stress.

    PubMed

    Wang, Zhuoyu; Hu, Hongtao; Goertzen, Leslie R; McElroy, J Scott; Dane, Fenny

    2014-01-01

    Citrullus colocynthis is a very drought tolerant species, closely related to watermelon (C. lanatus var. lanatus), an economically important cucurbit crop. Drought is a threat to plant growth and development, and the discovery of drought inducible genes with various functions is of great importance. We used high throughput mRNA Illumina sequencing technology and bioinformatic strategies to analyze the C. colocynthis leaf transcriptome under drought treatment. Leaf samples at four different time points (0, 24, 36, or 48 hours of withholding water) were used for RNA extraction and Illumina sequencing. qRT-PCR of several drought responsive genes was performed to confirm the accuracy of RNA sequencing. Leaf transcriptome analysis provided the first glimpse of the drought responsive transcriptome of this unique cucurbit species. A total of 5038 full-length cDNAs were detected, with 2545 genes showing significant changes during drought stress. Principle component analysis indicated that drought was the major contributing factor regulating transcriptome changes. Up regulation of many transcription factors, stress signaling factors, detoxification genes, and genes involved in phytohormone signaling and citrulline metabolism occurred under the water deficit conditions. The C. colocynthis transcriptome data highlight the activation of a large set of drought related genes in this species, thus providing a valuable resource for future functional analysis of candidate genes in defense of drought stress.

  7. Transcriptomics analysis and human preimplantation development.

    PubMed

    Freour, Thomas; Vassena, Rita

    2016-10-17

    The study of oocyte and preimplantation embryo biology has been regarded with great curiosity throughout scientific history, but it is not until the development of robust methods for in vitro observation and manipulation of animal gametes that developmental biology has flourished as a discipline. By far the biggest technical challenge in studying transcription in oocytes and early embryo has been the necessity of developing techniques that retain a high level of accuracy when starting from small amount of material. The objective of this narrative review is to summarize the knowledge gained about the embryonic preimplantation period in the human species from transcriptomics experiments, and to discuss technical limitations and solutions to the study of transcriptomics in these samples.

  8. Transcriptome Analysis of Mouse Stem Cells and Early Embryos

    PubMed Central

    Sharov, Alexei A; Piao, Yulan; Matoba, Ryo; Dudekula, Dawood B; Qian, Yong; VanBuren, Vincent; Falco, Geppino; Martin, Patrick R; Stagg, Carole A; Bassey, Uwem C; Wang, Yuxia; Carter, Mark G; Hamatani, Toshio; Aiba, Kazuhiro; Akutsu, Hidenori; Sharova, Lioudmila; Tanaka, Tetsuya S; Kimber, Wendy L; Yoshikawa, Toshiyuki; Jaradat, Saied A; Pantano, Serafino; Nagaraja, Ramaiah; Boheler, Kenneth R; Taub, Dennis; Hodes, Richard J; Longo, Dan L; Schlessinger, David; Keller, Jonathan; Klotz, Emily; Kelsoe, Garnett; Umezawa, Akihiro; Vescovi, Angelo L; Rossant, Janet; Kunath, Tilo; Hogan, Brigid L. M; Curci, Anna; D'Urso, Michele; Kelso, Janet; Hide, Winston

    2003-01-01

    Understanding and harnessing cellular potency are fundamental in biology and are also critical to the future therapeutic use of stem cells. Transcriptome analysis of these pluripotent cells is a first step towards such goals. Starting with sources that include oocytes, blastocysts, and embryonic and adult stem cells, we obtained 249,200 high-quality EST sequences and clustered them with public sequences to produce an index of approximately 30,000 total mouse genes that includes 977 previously unidentified genes. Analysis of gene expression levels by EST frequency identifies genes that characterize preimplantation embryos, embryonic stem cells, and adult stem cells, thus providing potential markers as well as clues to the functional features of these cells. Principal component analysis identified a set of 88 genes whose average expression levels decrease from oocytes to blastocysts, stem cells, postimplantation embryos, and finally to newborn tissues. This can be a first step towards a possible definition of a molecular scale of cellular potency. The sequences and cDNA clones recovered in this work provide a comprehensive resource for genes functioning in early mouse embryos and stem cells. The nonrestricted community access to the resource can accelerate a wide range of research, particularly in reproductive and regenerative medicine. PMID:14691545

  9. Transcriptome analysis by strand-specific sequencing of complementary DNA.

    PubMed

    Parkhomchuk, Dmitri; Borodina, Tatiana; Amstislavskiy, Vyacheslav; Banaru, Maria; Hallen, Linda; Krobitsch, Sylvia; Lehrach, Hans; Soldatov, Alexey

    2009-10-01

    High-throughput complementary DNA sequencing (RNA-Seq) is a powerful tool for whole-transcriptome analysis, supplying information about a transcript's expression level and structure. However, it is difficult to determine the polarity of transcripts, and therefore identify which strand is transcribed. Here, we present a simple cDNA sequencing protocol that preserves information about a transcript's direction. Using Saccharomyces cerevisiae and mouse brain transcriptomes as models, we demonstrate that knowing the transcript's orientation allows more accurate determination of the structure and expression of genes. It also helps to identify new genes and enables studying promoter-associated and antisense transcription. The transcriptional landscapes we obtained are available online.

  10. Transcriptomic analysis of phenotypic changes in birch (Betula platyphylla) autotetraploids.

    PubMed

    Mu, Huai-Zhi; Liu, Zi-Jia; Lin, Lin; Li, Hui-Yu; Jiang, Jing; Liu, Gui-Feng

    2012-10-11

    Plant breeders have focused much attention on polyploid trees because of their importance to forestry. To evaluate the impact of intraspecies genome duplication on the transcriptome, a series of Betula platyphylla autotetraploids and diploids were generated from four full-sib families. The phenotypes and transcriptomes of these autotetraploid individuals were compared with those of diploid trees. Autotetraploids were generally superior in breast-height diameter, volume, leaf, fruit and stoma and were generally inferior in height compared to diploids. Transcriptome data revealed numerous changes in gene expression attributable to autotetraploidization, which resulted in the upregulation of 7052 unigenes and the downregulation of 3658 unigenes. Pathway analysis revealed that the biosynthesis and signal transduction of indoleacetate (IAA) and ethylene were altered after genome duplication, which may have contributed to phenotypic changes. These results shed light on variations in birch autotetraploidization and help identify important genes for the genetic engineering of birch trees.

  11. Transcriptomic Analysis of Phenotypic Changes in Birch (Betula platyphylla) Autotetraploids

    PubMed Central

    Mu, Huai-Zhi; Liu, Zi-Jia; Lin, Lin; Li, Hui-Yu; Jiang, Jing; Liu, Gui-Feng

    2012-01-01

    Plant breeders have focused much attention on polyploid trees because of their importance to forestry. To evaluate the impact of intraspecies genome duplication on the transcriptome, a series of Betula platyphylla autotetraploids and diploids were generated from four full-sib families. The phenotypes and transcriptomes of these autotetraploid individuals were compared with those of diploid trees. Autotetraploids were generally superior in breast-height diameter, volume, leaf, fruit and stoma and were generally inferior in height compared to diploids. Transcriptome data revealed numerous changes in gene expression attributable to autotetraploidization, which resulted in the upregulation of 7052 unigenes and the downregulation of 3658 unigenes. Pathway analysis revealed that the biosynthesis and signal transduction of indoleacetate (IAA) and ethylene were altered after genome duplication, which may have contributed to phenotypic changes. These results shed light on variations in birch autotetraploidization and help identify important genes for the genetic engineering of birch trees. PMID:23202935

  12. Transcriptomic analysis of incised leaf-shape determination in birch.

    PubMed

    Mu, Huaizhi; Lin, Lin; Liu, Guifeng; Jiang, Jing

    2013-12-01

    Plant researchers have focused much attention on leaf shape because of its importance in the identification. To evaluate the impact of intraspecies leaf-shape variation on the transcriptome, a series of Betula pendula 'Dalecarlica' and B. pendula saplings were generated through tissue culture. The leaf shapes and transcriptomes of B. pendula 'Dalecarlica' clones were compared with those of B. pendula clones. The leaf shape of B. pendula 'Dalecarlica' was incised and that of B. pendula was ovate. Transcriptome data revealed numerous changes in gene expression between B. pendula 'Dalecarlica' and B. pendula, including upregulation of 8767 unigenes and downregulation of 8379 unigenes in B. pendula 'Dalecarlica'. A pathway analysis revealed that the transport and signal transduction of auxin were altered in 'Dalecarlica', which may have contributed to its altered leaf shape. These results shed light on variation in birch leaf shape and help identify important genes for the genetic engineering of birch trees.

  13. De Novo Transcriptome Sequencing Analysis of cDNA Library and Large-Scale Unigene Assembly in Japanese Red Pine (Pinus densiflora).

    PubMed

    Liu, Le; Zhang, Shijie; Lian, Chunlan

    2015-12-04

    Japanese red pine (Pinus densiflora) is extensively cultivated in Japan, Korea, China, and Russia and is harvested for timber, pulpwood, garden, and paper markets. However, genetic information and molecular markers were very scarce for this species. In this study, over 51 million sequencing clean reads from P. densiflora mRNA were produced using Illumina paired-end sequencing technology. It yielded 83,913 unigenes with a mean length of 751 bp, of which 54,530 (64.98%) unigenes showed similarity to sequences in the NCBI database. Among which the best matches in the NCBI Nr database were Picea sitchensis (41.60%), Amborella trichopoda (9.83%), and Pinus taeda (4.15%). A total of 1953 putative microsatellites were identified in 1784 unigenes using MISA (MicroSAtellite) software, of which the tri-nucleotide repeats were most abundant (50.18%) and 629 EST-SSR (expressed sequence tag- simple sequence repeats) primer pairs were successfully designed. Among 20 EST-SSR primer pairs randomly chosen, 17 markers yielded amplification products of the expected size in P. densiflora. Our results will provide a valuable resource for gene-function analysis, germplasm identification, molecular marker-assisted breeding and resistance-related gene(s) mapping for pine for P. densiflora.

  14. De Novo Transcriptome Sequencing Analysis of cDNA Library and Large-Scale Unigene Assembly in Japanese Red Pine (Pinus densiflora)

    PubMed Central

    Liu, Le; Zhang, Shijie; Lian, Chunlan

    2015-01-01

    Japanese red pine (Pinus densiflora) is extensively cultivated in Japan, Korea, China, and Russia and is harvested for timber, pulpwood, garden, and paper markets. However, genetic information and molecular markers were very scarce for this species. In this study, over 51 million sequencing clean reads from P. densiflora mRNA were produced using Illumina paired-end sequencing technology. It yielded 83,913 unigenes with a mean length of 751 bp, of which 54,530 (64.98%) unigenes showed similarity to sequences in the NCBI database. Among which the best matches in the NCBI Nr database were Picea sitchensis (41.60%), Amborella trichopoda (9.83%), and Pinus taeda (4.15%). A total of 1953 putative microsatellites were identified in 1784 unigenes using MISA (MicroSAtellite) software, of which the tri-nucleotide repeats were most abundant (50.18%) and 629 EST-SSR (expressed sequence tag- simple sequence repeats) primer pairs were successfully designed. Among 20 EST-SSR primer pairs randomly chosen, 17 markers yielded amplification products of the expected size in P. densiflora. Our results will provide a valuable resource for gene-function analysis, germplasm identification, molecular marker-assisted breeding and resistance-related gene(s) mapping for pine for P. densiflora. PMID:26690126

  15. Large-scale transcriptome analysis in chickpea (Cicer arietinum L.), an orphan legume crop of the semi-arid tropics of Asia and Africa

    PubMed Central

    Hiremath, Pavana J; Farmer, Andrew; Cannon, Steven B; Woodward, Jimmy; Kudapa, Himabindu; Tuteja, Reetu; Kumar, Ashish; BhanuPrakash, Amindala; Mulaosmanovic, Benjamin; Gujaria, Neha; Krishnamurthy, Laxmanan; Gaur, Pooran M; KaviKishor, Polavarapu B; Shah, Trushar; Srinivasan, Ramamurthy; Lohse, Marc; Xiao, Yongli; Town, Christopher D; Cook, Douglas R; May, Gregory D; Varshney, Rajeev K

    2011-01-01

    Chickpea (Cicer arietinum L.) is an important legume crop in the semi-arid regions of Asia and Africa. Gains in crop productivity have been low however, particularly because of biotic and abiotic stresses. To help enhance crop productivity using molecular breeding techniques, next generation sequencing technologies such as Roche/454 and Illumina/Solexa were used to determine the sequence of most gene transcripts and to identify drought-responsive genes and gene-based molecular markers. A total of 103 215 tentative unique sequences (TUSs) have been produced from 435 018 Roche/454 reads and 21 491 Sanger expressed sequence tags (ESTs). Putative functions were determined for 49 437 (47.8%) of the TUSs, and gene ontology assignments were determined for 20 634 (41.7%) of the TUSs. Comparison of the chickpea TUSs with the Medicago truncatula genome assembly (Mt 3.5.1 build) resulted in 42 141 aligned TUSs with putative gene structures (including 39 281 predicted intron/splice junctions). Alignment of ∼37 million Illumina/Solexa tags generated from drought-challenged root tissues of two chickpea genotypes against the TUSs identified 44 639 differentially expressed TUSs. The TUSs were also used to identify a diverse set of markers, including 728 simple sequence repeats (SSRs), 495 single nucleotide polymorphisms (SNPs), 387 conserved orthologous sequence (COS) markers, and 2088 intron-spanning region (ISR) markers. This resource will be useful for basic and applied research for genome analysis and crop improvement in chickpea. PMID:21615673

  16. Large-scale transcriptome analysis in chickpea (Cicer arietinum L.), an orphan legume crop of the semi-arid tropics of Asia and Africa.

    PubMed

    Hiremath, Pavana J; Farmer, Andrew; Cannon, Steven B; Woodward, Jimmy; Kudapa, Himabindu; Tuteja, Reetu; Kumar, Ashish; Bhanuprakash, Amindala; Mulaosmanovic, Benjamin; Gujaria, Neha; Krishnamurthy, Laxmanan; Gaur, Pooran M; Kavikishor, Polavarapu B; Shah, Trushar; Srinivasan, Ramamurthy; Lohse, Marc; Xiao, Yongli; Town, Christopher D; Cook, Douglas R; May, Gregory D; Varshney, Rajeev K

    2011-10-01

    Chickpea (Cicer arietinum L.) is an important legume crop in the semi-arid regions of Asia and Africa. Gains in crop productivity have been low however, particularly because of biotic and abiotic stresses. To help enhance crop productivity using molecular breeding techniques, next generation sequencing technologies such as Roche/454 and Illumina/Solexa were used to determine the sequence of most gene transcripts and to identify drought-responsive genes and gene-based molecular markers. A total of 103,215 tentative unique sequences (TUSs) have been produced from 435,018 Roche/454 reads and 21,491 Sanger expressed sequence tags (ESTs). Putative functions were determined for 49,437 (47.8%) of the TUSs, and gene ontology assignments were determined for 20,634 (41.7%) of the TUSs. Comparison of the chickpea TUSs with the Medicago truncatula genome assembly (Mt 3.5.1 build) resulted in 42,141 aligned TUSs with putative gene structures (including 39,281 predicted intron/splice junctions). Alignment of ∼37 million Illumina/Solexa tags generated from drought-challenged root tissues of two chickpea genotypes against the TUSs identified 44,639 differentially expressed TUSs. The TUSs were also used to identify a diverse set of markers, including 728 simple sequence repeats (SSRs), 495 single nucleotide polymorphisms (SNPs), 387 conserved orthologous sequence (COS) markers, and 2088 intron-spanning region (ISR) markers. This resource will be useful for basic and applied research for genome analysis and crop improvement in chickpea. Plant Biotechnology Journal © 2011 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd. No claim to original US government works.

  17. IMPROVED PERFORMANCE OF GENE SET ANALYSIS ON GENOME-WIDE TRANSCRIPTOMICS DATA WHEN USING GENE ACTIVITY STATE ESTIMATES.

    PubMed

    Kamp, Thomas; Adams, Micah; Disselkoen, Craig; Tintle, Nathan

    2016-01-01

    Gene set analysis methods continue to be a popular and powerful method of evaluating genome-wide transcriptomics data. These approach require a priori grouping of genes into biologically meaningful sets, and then conducting downstream analyses at the set (instead of gene) level of analysis. Gene set analysis methods have been shown to yield more powerful statistical conclusions than single-gene analyses due to both reduced multiple testing penalties and potentially larger observed effects due to the aggregation of effects across multiple genes in the set. Traditionally, gene set analysis methods have been applied directly to normalized, log-transformed, transcriptomics data. Recently, efforts have been made to transform transcriptomics data to scales yielding more biologically interpretable results. For example, recently proposed models transform log-transformed transcriptomics data to a confidence metric (ranging between 0 and 100%) that a gene is active (roughly speaking, that the gene product is part of an active cellular mechanism). In this manuscript, we demonstrate, on both real and simulated transcriptomics data, that tests for differential expression between sets of genes using are typically more powerful when using gene activity state estimates as opposed to log-transformed gene expression data. Our analysis suggests further exploration of techniques to transform transcriptomics data to meaningful quantities for improved downstream inference.

  18. Transcriptomic analysis of Salmonella desiccation resistance.

    PubMed

    Li, Haiping; Bhaskara, Anuhya; Megalis, Christina; Tortorello, Mary Lou

    2012-12-01

    The survival of Salmonella in low moisture foods and processing environments remains a great challenge for the food industry and public health. To explore the mechanisms of Salmonella desiccation resistance, we studied the transcriptomic responses in Salmonella Tennessee (Tennessee), using Salmonella Typhimurium LT2 (LT2), a strain weakly resistant to desiccation, as a reference strain. In response to 2 h of air-drying at 11% equilibrated relative humidity, approximately one-fourth of the open reading frames (ORFs) in the Tennessee genome and one-fifth in LT2 were differentially expressed (>2-fold). Among all differentially expressed functional groups (>5-fold) in both strains, the expression fold change associated with fatty acid metabolism was the highest, and constituted 51% and 35% of the total expression fold change in Tennessee and LT2, respectively. Tennessee showed greater changes in expression of genes associated with stress response and envelope modification than LT2, while showing lesser changes in protein biosynthesis expression. Expression of flagella genes was significantly more inhibited in stationary phase cells of Tennessee than LT2 both before and after desiccation. The accumulation of the osmolyte trehalose was significantly induced by desiccation in Tennessee, but no increase was detectable in LT2, which is consistent with the expression patterns of the entire trehalose biosynthesis and degradation pathways in both strains. Results from this study present a global view of the dynamic desiccation responses in Salmonella, which will guide future research efforts to control Salmonella in low moisture environments.

  19. Transcriptomic analysis of flower development in tea (Camellia sinensis (L.)).

    PubMed

    Liu, Feng; Wang, Yu; Ding, Zhaotang; Zhao, Lei; Xiao, Jun; Wang, Linjun; Ding, Shibo

    2017-10-05

    Flowering is a critical and complicated process in plant development, involving interactions of numerous endogenous and environmental factors, but little is known about the complex network regulating flower development in tea plants. In this study, de novo transcriptome assembly and gene expression analysis using Illumina sequencing technology were performed. Transcriptomic analysis assembles gene-related information involved in reproductive growth of C. sinensis. 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. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that metabolic pathways, biosynthesis of secondary metabolites, and plant hormone signal transduction were enriched among the DEGs. Furthermore, 207 flowering-associated unigenes were identified from our database. Some transcription factors, such as WRKY, ERF, bHLH, MYB and MADS-box were shown to be up-regulated in floral transition, which might play the role of progression of flowering. Furthermore, 14 genes were selected for confirmation of expression levels using quantitative real-time PCR (qRT-PCR). The comprehensive transcriptomic analysis presents fundamental information on the genes and pathways which are involved in flower development in C. sinensis. Our data also provided a useful database for further research of tea and other species of plants. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Transcriptome Analysis of Cytokinin Response in Tomato Leaves

    PubMed Central

    Shi, Xiuling; Gupta, Sarika; Lindquist, Ingrid E.; Cameron, Connor T.; Mudge, Joann; Rashotte, Aaron M.

    2013-01-01

    Tomato is one of the most economically and agriculturally important Solanaceous species and vegetable crops, serving as a model for examination of fruit biology and compound leaf development. Cytokinin is a plant hormone linked to the control of leaf development and is known to regulate a wide range of genes including many transcription factors. Currently there is little known of the leaf transcriptome in tomato and how it might be regulated by cytokinin. We employ high throughput mRNA sequencing technology and bioinformatic methodologies to robustly analyze cytokinin regulated tomato leaf transcriptomes. Leaf samples of two ages, 13d and 35d were treated with cytokinin or the solvent vehicle control dimethyl sulfoxide (DMSO) for 2 h or 24 h, after which RNA was extracted for sequencing. To confirm the accuracy of RNA sequencing results, we performed qPCR analysis of select transcripts identified as cytokinin regulated by the RNA sequencing approach. The resulting data provide the first hormone transcriptome analysis of leaves in tomato. Specifically we identified several previously untested tomato orthologs of cytokinin-related genes as well as numerous novel cytokinin-regulated transcripts in tomato leaves. Principal component analysis of the data indicates that length of cytokinin treatment and plant age are the major factors responsible for changes in transcripts observed in this study. Two hour cytokinin treatment showed a more robust transcript response indicated by both greater fold change of induced transcripts and the induction of twice as many cytokinin-related genes involved in signaling, metabolism, and transport in young vs. older leaves. This difference in transcriptome response in younger vs. older leaves was also found to a lesser extent with an extended (24 h) cytokinin treatment. Overall data presented here provides a solid foundation for future study of cytokinin and cytokinin regulated genes involved in compound leaf development or other

  1. Transcriptome analysis identifies novel responses and potential regulatory genes involved in seasonal dormancy transitions of leafy spurge (Euphorbia esula L.)

    USDA-ARS?s Scientific Manuscript database

    Dormancy transitions in crown buds of leafy spurge were investigated using 23K element cDNA microarrays. These data represent the first large-scale transcriptome analysis of dormancy in underground buds of an herbaceous perennial species. Crown buds collected monthly from August through December, ov...

  2. Transcriptome analysis by strand-specific sequencing of complementary DNA

    PubMed Central

    Parkhomchuk, Dmitri; Borodina, Tatiana; Amstislavskiy, Vyacheslav; Banaru, Maria; Hallen, Linda; Krobitsch, Sylvia; Lehrach, Hans; Soldatov, Alexey

    2009-01-01

    High-throughput complementary DNA sequencing (RNA-Seq) is a powerful tool for whole-transcriptome analysis, supplying information about a transcript's expression level and structure. However, it is difficult to determine the polarity of transcripts, and therefore identify which strand is transcribed. Here, we present a simple cDNA sequencing protocol that preserves information about a transcript's direction. Using Saccharomyces cerevisiae and mouse brain transcriptomes as models, we demonstrate that knowing the transcript's orientation allows more accurate determination of the structure and expression of genes. It also helps to identify new genes and enables studying promoter-associated and antisense transcription. The transcriptional landscapes we obtained are available online. PMID:19620212

  3. Transcriptome Analysis to Understand the Toxicity of Latrodectus tredecimguttatus Eggs

    PubMed Central

    Xu, Dehong; Wang, Xianchun

    2016-01-01

    Latrodectus tredecimguttatus is a kind of highly venomous black widow spider, with toxicity coming from not only venomous glands but also other parts of its body as well as newborn spiderlings and eggs. Up to date, although L. tredecimguttatus eggs have been demonstrated to be rich in proteinaceous toxins, there is no systematic investigation on such active components at transcriptome level. In this study, we performed a high-throughput transcriptome sequencing of L. tredecimguttatus eggs with Illumina sequencing technology. As a result, 53,284 protein-coding unigenes were identified, of which 14,185 unigenes produced significant hits in the available databases, including 280 unigenes encoding proteins or peptides homologous to known proteinaceous toxins. GO term and KEGG pathway enrichment analyses of the 280 unigenes showed that 375 GO terms and 18 KEGG pathways were significantly enriched. Functional analysis indicated that these unigene-coded toxins have the bioactivities to degrade tissue proteins, inhibit ion channels, block neuromuscular transmission, provoke anaphylaxis, induce apoptosis and hyperalgesia, etc. No known typical proteinaceous toxins in L. tredecimguttatus venomous glands, such as latrotoxins, were identified, suggesting that the eggs have a different toxicity mechanism from that of the venom. Our present transcriptome analysis not only helps to reveal the gene expression profile and toxicity mechanism of the L. tredecimguttatus eggs, but also provides references for the further related researches. PMID:27999389

  4. Transcriptomic analysis of the salivary glands of an invasive whitefly.

    PubMed

    Su, Yun-Lin; Li, Jun-Min; Li, Meng; Luan, Jun-Bo; Ye, Xiao-Dong; Wang, Xiao-Wei; Liu, Shu-Sheng

    2012-01-01

    Some species of the whitefly Bemisia tabaci complex cause tremendous losses to crops worldwide through feeding directly and virus transmission indirectly. The primary salivary glands of whiteflies are critical for their feeding and virus transmission. However, partly due to their tiny size, research on whitefly salivary glands is limited and our knowledge on these glands is scarce. We sequenced the transcriptome of the primary salivary glands of the Mediterranean species of B. tabaci complex using an effective cDNA amplification method in combination with short read sequencing (Illumina). In a single run, we obtained 13,615 unigenes. The quantity of the unigenes obtained from the salivary glands of the whitefly is at least four folds of the salivary gland genes from other plant-sucking insects. To reveal the functions of the primary glands, sequence similarity search and comparisons with the whole transcriptome of the whitefly were performed. The results demonstrated that the genes related to metabolism and transport were significantly enriched in the primary salivary glands. Furthermore, we found that a number of highly expressed genes in the salivary glands might be involved in secretory protein processing, secretion and virus transmission. To identify potential proteins of whitefly saliva, the translated unigenes were put into secretory protein prediction. Finally, 295 genes were predicted to encode secretory proteins and some of them might play important roles in whitefly feeding. The combined method of cDNA amplification, Illumina sequencing and de novo assembly is suitable for transcriptomic analysis of tiny organs in insects. Through analysis of the transcriptome, genomic features of the primary salivary glands were dissected and biologically important proteins, especially secreted proteins, were predicted. Our findings provide substantial sequence information for the primary salivary glands of whiteflies and will be the basis for future studies on whitefly

  5. Transcriptomic Analysis of the Salivary Glands of an Invasive Whitefly

    PubMed Central

    Su, Yun-Lin; Li, Jun-Min; Li, Meng; Luan, Jun-Bo; Ye, Xiao-Dong; Wang, Xiao-Wei; Liu, Shu-Sheng

    2012-01-01

    Background Some species of the whitefly Bemisia tabaci complex cause tremendous losses to crops worldwide through feeding directly and virus transmission indirectly. The primary salivary glands of whiteflies are critical for their feeding and virus transmission. However, partly due to their tiny size, research on whitefly salivary glands is limited and our knowledge on these glands is scarce. Methodology/Principal Findings We sequenced the transcriptome of the primary salivary glands of the Mediterranean species of B. tabaci complex using an effective cDNA amplification method in combination with short read sequencing (Illumina). In a single run, we obtained 13,615 unigenes. The quantity of the unigenes obtained from the salivary glands of the whitefly is at least four folds of the salivary gland genes from other plant-sucking insects. To reveal the functions of the primary glands, sequence similarity search and comparisons with the whole transcriptome of the whitefly were performed. The results demonstrated that the genes related to metabolism and transport were significantly enriched in the primary salivary glands. Furthermore, we found that a number of highly expressed genes in the salivary glands might be involved in secretory protein processing, secretion and virus transmission. To identify potential proteins of whitefly saliva, the translated unigenes were put into secretory protein prediction. Finally, 295 genes were predicted to encode secretory proteins and some of them might play important roles in whitefly feeding. Conclusions/Significance: The combined method of cDNA amplification, Illumina sequencing and de novo assembly is suitable for transcriptomic analysis of tiny organs in insects. Through analysis of the transcriptome, genomic features of the primary salivary glands were dissected and biologically important proteins, especially secreted proteins, were predicted. Our findings provide substantial sequence information for the primary salivary glands

  6. Analysis of the transcriptome in molecular epidemiology studies

    PubMed Central

    McHale, Cliona M.; Zhang, Luoping; Thomas, Reuben; smith, Martyn T.

    2016-01-01

    The human transcriptome is complex, comprising multiple transcript types, mostly in the form of non-coding RNA (ncRNA). The majority of ncRNA is of the long form (lncRNA, ≥200bp), which plays an important role in gene regulation through multiple mechanisms including epigenetics, chromatin modification, control of transcription factor binding, and regulation of alternative splicing. Both mRNA and ncRNA exhibit additional variability in the form of alternative splicing and RNA editing. All aspects of the human transcriptome can potentially be dysregulated by environmental exposures. Next-generation RNA sequencing (RNA-Seq) is the best available methodology to measure this although it has limitations, including experimental bias. The third phase of the MicroArray Quality Control Consortium project (MAQC-III), also called Sequencing Quality Control (SeQC), aims to address these limitations through standardization of experimental and bioinformatic methodologies. A limited number of toxicogenomic studies have been conducted to date using RNA-Seq. This review describes the complexity of the human transcriptome, the application of transcriptomics by RNA-Seq or microarray in molecular epidemiology studies, and limitations of these approaches including the type of cell or tissue analyzed, experimental variation, and confounding. By using good study designs with precise, individual exposure measurements, sufficient power and incorporation of phenotypic anchors, studies in human populations can identify biomarkers of exposure and/or early effect and elucidate mechanisms of action underlying associated diseases, even at low doses. Analysis of datasets at the pathway level can compensate for some of the limitations of RNA-Seq and, as more datasets become available, will increasingly elucidate the exposure-disease continuum. PMID:23907930

  7. Transcriptome Analysis of the Asian Honey Bee Apis cerana cerana

    PubMed Central

    Huang, Zachary Y.; Wu, Xiao Bo; Yan, Wei Yu; Zeng, Zhi Jiang

    2012-01-01

    Background The Eastern hive honey bee, Apis cerana cerana is a native and widely bred honey bee species in China. Molecular biology research about this honey bee species is scarce, and genomic information for A. c. cerana is not currently available. Transcriptome and expression profiling data for this species are therefore important resources needed to better understand the biological mechanisms of A. c. cerana. In this study, we obtained the transcriptome information of A. c. cerana by RNA-sequencing and compared gene expression differences between queens and workers of A. c. cerana by digital gene expression (DGE) analysis. Results Using high-throughput Illumina RNA sequencing we obtained 51,581,510 clean reads corresponding to 4.64 Gb total nucleotides from a single run. These reads were assembled into 46,999 unigenes with a mean length of 676 bp. Based on a sequence similarity search against the five public databases (NR, Swissport, GO, COG, KEGG) with a cut-off E-value of 10−5 using BLASTX, a total of 24,630 unigenes were annotated with gene descriptions, gene ontology terms, or metabolic pathways. Using these transcriptome data as references we analyzed the gene expression differences between the queens and workers of A. c. cerana using a tag-based digital gene expression method. We obtained 5.96 and 5.66 million clean tags from the queen and worker samples, respectively. A total of 414 genes were differentially expressed between them, with 189 up-regulated and 225 down-regulated in queens. Conclusions Our transcriptome data provide a comprehensive sequence resource for future A. c. cerana study, establishing an important public information platform for functional genomic studies in A. c. cerana. Furthermore, the DGE data provide comprehensive gene expression information for the queens and workers, which will facilitate our understanding of the molecular mechanisms of the different physiological aspects of the two castes. PMID:23112877

  8. Transcriptome and proteomic analysis of mango (Mangifera indica Linn) fruits.

    PubMed

    Wu, Hong-xia; Jia, Hui-min; Ma, Xiao-wei; Wang, Song-biao; Yao, Quan-sheng; Xu, Wen-tian; Zhou, Yi-gang; Gao, Zhong-shan; Zhan, Ru-lin

    2014-06-13

    Here we used Illumina RNA-seq technology for transcriptome sequencing of a mixed fruit sample from 'Zill' mango (Mangifera indica Linn) fruit pericarp and pulp during the development and ripening stages. RNA-seq generated 68,419,722 sequence reads that were assembled into 54,207 transcripts with a mean length of 858bp, including 26,413 clusters and 27,794 singletons. A total of 42,515(78.43%) transcripts were annotated using public protein databases, with a cut-off E-value above 10(-5), of which 35,198 and 14,619 transcripts were assigned to gene ontology terms and clusters of orthologous groups respectively. Functional annotation against the Kyoto Encyclopedia of Genes and Genomes database identified 23,741(43.79%) transcripts which were mapped to 128 pathways. These pathways revealed many previously unknown transcripts. We also applied mass spectrometry-based transcriptome data to characterize the proteome of ripe fruit. LC-MS/MS analysis of the mango fruit proteome was using tandem mass spectrometry (MS/MS) in an LTQ Orbitrap Velos (Thermo) coupled online to the HPLC. This approach enabled the identification of 7536 peptides that matched 2754 proteins. Our study provides a comprehensive sequence for a systemic view of transcriptome during mango fruit development and the most comprehensive fruit proteome to date, which are useful for further genomics research and proteomic studies. Our study provides a comprehensive sequence for a systemic view of both the transcriptome and proteome of mango fruit, and a valuable reference for further research on gene expression and protein identification. This article is part of a Special Issue entitled: Proteomics of non-model organisms. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Transcriptome Analysis of Catharanthus roseus for Gene Discovery and Expression Profiling

    PubMed Central

    Sharma, Raghvendra; Sinha, Alok K.; Jain, Mukesh

    2014-01-01

    The medicinal plant, Catharanthus roseus, accumulates wide range of terpenoid indole alkaloids, which are well documented therapeutic agents. In this study, deep transcriptome sequencing of C. roseus was carried out to identify the pathways and enzymes (genes) involved in biosynthesis of these compounds. About 343 million reads were generated from different tissues (leaf, flower and root) of C. roseus using Illumina platform. Optimization of de novo assembly involving a two-step process resulted in a total of 59,220 unique transcripts with an average length of 1284 bp. Comprehensive functional annotation and gene ontology (GO) analysis revealed the representation of many genes involved in different biological processes and molecular functions. In total, 65% of C. roseus transcripts showed homology with sequences available in various public repositories, while remaining 35% unigenes may be considered as C. roseus specific. In silico analysis revealed presence of 11,620 genic simple sequence repeats (excluding mono-nucleotide repeats) and 1820 transcription factor encoding genes in C. roseus transcriptome. Expression analysis showed roots and leaves to be actively participating in bisindole alkaloid production with clear indication that enzymes involved in pathway of vindoline and vinblastine biosynthesis are restricted to aerial tissues. Such large-scale transcriptome study provides a rich source for understanding plant-specialized metabolism, and is expected to promote research towards production of plant-derived pharmaceuticals. PMID:25072156

  10. Transcriptome analysis of adiposity in domestic ducks by transcriptomic comparison with their wild counterparts.

    PubMed

    Chen, L; Luo, J; Li, J X; Li, J J; Wang, D Q; Tian, Y; Lu, L Z

    2015-06-01

    Excessive adiposity is a major problem in the duck industry, but its molecular mechanisms remain unknown. Genetic comparisons between domestic and wild animals have contributed to the exploration of genetic mechanisms responsible for many phenotypic traits. Significant differences in body fat mass have been detected between domestic and wild ducks. In this study, we used the Peking duck and Anas platyrhynchos as the domestic breed and wild counterpart respectively and performed a transcriptomic comparison of abdominal fat between the two breeds to comprehensively analyze the transcriptome basis of adiposity in ducks. We obtained approximately 350 million clean reads; assembled 61 250 transcripts, including 23 699 novel ones; and identified alternative 5' splice sites, alternative 3' splice sites, skipped exons and retained intron as the main alternative splicing events. A differential expression analysis between the two breeds showed that 753 genes exhibited differential expression. In Peking ducks, some lipid metabolism-related genes (IGF2, FABP5, BMP7, etc.) and oncogenes (RRM2, AURKA, CYR61, etc.) were upregulated, whereas genes related to tumor suppression and immunity (TNFRSF19, TNFAIP6, IGSF21, NCF1, etc.) were downregulated, suggesting adiposity might closely associate with tumorigenesis in ducks. Furthermore, 280 576 single-nucleotide variations were found differentiated between the two breeds, including 8641 non-synonymous ones, and some of the non-synonymous ones were found enriched in genes involved in lipid-associated and immune-associated pathways, suggesting abdominal fat of the duck undertakes both a metabolic function and immune-related function. These datasets enlarge our genetic information of ducks and provide valuable resources for analyzing mechanisms underlying adiposity in ducks. © 2015 Stichting International Foundation for Animal Genetics.

  11. 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

  12. Transcriptomic analysis of purple leaf determination in birch.

    PubMed

    Lin, Lin; Mu, Huaizhi; Jiang, Jing; Liu, Guifeng

    2013-09-10

    'Purple Rain', a purple cultivar of Betula pendula, has dark purple leaves throughout the vegetative period. In this study, B. pendula 'Purple Rain' was found to have a higher anthocyanidin level compared with B. pendula, Transcriptome analysis revealed numerous changes in gene expression that could be attributed to color change, including the upregulation of 2467 unigenes and the downregulation of 2299 unigenes in 'Purple Rain'. Furthermore, anthocyanidin synthesis and transcriptional regulation were altered in 'Purple Rain', which may have contributed to phenotypic changes. These results provide unique molecular insights into the biochemical pathways and regulatory networks that function in a purple variety of B. pendula.

  13. Transcriptome Analysis of Human Diabetic Kidney Disease

    PubMed Central

    Woroniecka, Karolina I.; Park, Ae Seo Deok; Mohtat, Davoud; Thomas, David B.; Pullman, James M.; Susztak, Katalin

    2011-01-01

    OBJECTIVE Diabetic kidney disease (DKD) is the single leading cause of kidney failure in the U.S., for which a cure has not yet been found. The aim of our study was to provide an unbiased catalog of gene-expression changes in human diabetic kidney biopsy samples. RESEARCH DESIGN AND METHODS Affymetrix expression arrays were used to identify differentially regulated transcripts in 44 microdissected human kidney samples. DKD samples were significant for their racial diversity and decreased glomerular filtration rate (~25–35 mL/min). Stringent statistical analysis, using the Benjamini-Hochberg corrected two-tailed t test, was used to identify differentially expressed transcripts in control and diseased glomeruli and tubuli. Two different web-based algorithms were used to define differentially regulated pathways. RESULTS We identified 1,700 differentially expressed probesets in DKD glomeruli and 1,831 in diabetic tubuli, and 330 probesets were commonly differentially expressed in both compartments. Pathway analysis highlighted the regulation of Ras homolog gene family member A, Cdc42, integrin, integrin-linked kinase, and vascular endothelial growth factor signaling in DKD glomeruli. The tubulointerstitial compartment showed strong enrichment for inflammation-related pathways. The canonical complement signaling pathway was determined to be statistically differentially regulated in both DKD glomeruli and tubuli and was associated with increased glomerulosclerosis even in a different set of DKD samples. CONCLUSIONS Our studies have cataloged gene-expression regulation and identified multiple novel genes and pathways that may play a role in the pathogenesis of DKD or could serve as biomarkers. PMID:21752957

  14. Transcriptomic analysis of flower development in wintersweet (Chimonanthus praecox).

    PubMed

    Liu, Daofeng; Sui, Shunzhao; Ma, Jing; Li, Zhineng; Guo, Yulong; Luo, Dengpan; Yang, Jianfeng; Li, Mingyang

    2014-01-01

    Wintersweet (Chimonanthus praecox) is familiar as a garden plant and woody ornamental flower. On account of its unique flowering time and strong fragrance, it has a high ornamental and economic value. Despite a long history of human cultivation, our understanding of wintersweet genetics and molecular biology remains scant, reflecting a lack of basic genomic and transcriptomic data. In this study, we assembled three cDNA libraries, from three successive stages in flower development, designated as the flower bud with displayed petal, open flower and senescing flower stages. Using the Illumina RNA-Seq method, we obtained 21,412,928, 26,950,404, 24,912,954 qualified Illumina reads, respectively, for the three successive stages. The pooled reads from all three libraries were then assembled into 106,995 transcripts, 51,793 of which were annotated in the NCBI non-redundant protein database. Of these annotated sequences, 32,649 and 21,893 transcripts were assigned to gene ontology categories and clusters of orthologous groups, respectively. We could map 15,587 transcripts onto 312 pathways using the Kyoto Encyclopedia of Genes and Genomes pathway database. Based on these transcriptomic data, we obtained a large number of candidate genes that were differentially expressed at the open flower and senescing flower stages. An analysis of differentially expressed genes involved in plant hormone signal transduction pathways indicated that although flower opening and senescence may be independent of the ethylene signaling pathway in wintersweet, salicylic acid may be involved in the regulation of flower senescence. We also succeeded in isolating key genes of floral scent biosynthesis and proposed a biosynthetic pathway for monoterpenes and sesquiterpenes in wintersweet flowers, based on the annotated sequences. This comprehensive transcriptomic analysis presents fundamental information on the genes and pathways which are involved in flower development in wintersweet. And our data

  15. Transcriptome and proteome analysis of Eucalyptus infected with Calonectria pseudoreteaudii.

    PubMed

    Chen, Quanzhu; Guo, Wenshuo; Feng, Lizhen; Ye, Xiaozhen; Xie, Wanfeng; Huang, Xiuping; Liu, Jinyan

    2015-02-06

    Cylindrocladium leaf blight is one of the most severe diseases in Eucalyptus plantations and nurseries. There are Eucalyptus cultivars with resistance to the disease. However, little is known about the defense mechanism of resistant cultivars. Here, we investigated the transcriptome and proteome of Eucalyptus leaves (E. urophylla×E. tereticornis M1), infected or not with Calonectria pseudoreteaudii. A total of 8585 differentially expressed genes (|log2 ratio| ≥1, FDR ≤0.001) at 12 and 24hours post-inoculation were detected using RNA-seq. Transcriptional changes for five genes were further confirmed by qRT-PCR. A total of 3680 proteins at the two time points were identified using iTRAQ technique.The combined transcriptome and proteome analysis revealed that the shikimate/phenylpropanoid pathway, terpenoid biosynthesis, signalling pathway (jasmonic acid and sugar) were activated. The data also showed that some proteins (WRKY33 and PR proteins) which have been reported to involve in plant defense response were up-regulated. However, photosynthesis, nucleic acid metabolism and protein metabolism were impaired by the infection of C. pseudoreteaudii. This work will facilitate the identification of defense related genes and provide insights into Eucalyptus defense responses to Cylindrocladium leaf blight. In this study, a total of 130 proteins and genes involved in the shikimate/phenylpropanoid pathway, terpenoid biosynthesis, signalling pathway, cell transport, carbohydrate and energy metabolism, nucleic acid metabolism and protein metabolism in Eucalyptus leaves after infected with C. pseudoreteaudii were identified. This is the first report of a comprehensive transcriptomic and proteomic analysis of Eucalyptus in response to Calonectria sp. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Transcriptomic Analysis of Flower Development in Wintersweet (Chimonanthus praecox)

    PubMed Central

    Liu, Daofeng; Sui, Shunzhao; Ma, Jing; Li, Zhineng; Guo, Yulong; Luo, Dengpan; Yang, Jianfeng; Li, Mingyang

    2014-01-01

    Wintersweet (Chimonanthus praecox) is familiar as a garden plant and woody ornamental flower. On account of its unique flowering time and strong fragrance, it has a high ornamental and economic value. Despite a long history of human cultivation, our understanding of wintersweet genetics and molecular biology remains scant, reflecting a lack of basic genomic and transcriptomic data. In this study, we assembled three cDNA libraries, from three successive stages in flower development, designated as the flower bud with displayed petal, open flower and senescing flower stages. Using the Illumina RNA-Seq method, we obtained 21,412,928, 26,950,404, 24,912,954 qualified Illumina reads, respectively, for the three successive stages. The pooled reads from all three libraries were then assembled into 106,995 transcripts, 51,793 of which were annotated in the NCBI non-redundant protein database. Of these annotated sequences, 32,649 and 21,893 transcripts were assigned to gene ontology categories and clusters of orthologous groups, respectively. We could map 15,587 transcripts onto 312 pathways using the Kyoto Encyclopedia of Genes and Genomes pathway database. Based on these transcriptomic data, we obtained a large number of candidate genes that were differentially expressed at the open flower and senescing flower stages. An analysis of differentially expressed genes involved in plant hormone signal transduction pathways indicated that although flower opening and senescence may be independent of the ethylene signaling pathway in wintersweet, salicylic acid may be involved in the regulation of flower senescence. We also succeeded in isolating key genes of floral scent biosynthesis and proposed a biosynthetic pathway for monoterpenes and sesquiterpenes in wintersweet flowers, based on the annotated sequences. This comprehensive transcriptomic analysis presents fundamental information on the genes and pathways which are involved in flower development in wintersweet. And our data

  17. Transcriptomic Analysis of Murine Embryos Lacking Endogenous Retinoic Acid Signaling

    PubMed Central

    Paschaki, Marie; Schneider, Carole; Rhinn, Muriel; Thibault-Carpentier, Christelle; Dembélé, Doulaye; Niederreither, Karen; Dollé, Pascal

    2013-01-01

    Retinoic acid (RA), an active derivative of the liposoluble vitamin A (retinol), acts as an important signaling molecule during embryonic development, regulating phenomenons as diverse as anterior-posterior axial patterning, forebrain and optic vesicle development, specification of hindbrain rhombomeres, pharyngeal arches and second heart field, somitogenesis, and differentiation of spinal cord neurons. This small molecule directly triggers gene activation by binding to nuclear receptors (RARs), switching them from potential repressors to transcriptional activators. The repertoire of RA-regulated genes in embryonic tissues is poorly characterized. We performed a comparative analysis of the transcriptomes of murine wild-type and Retinaldehyde Dehydrogenase 2 null-mutant (Raldh2−/−) embryos — unable to synthesize RA from maternally-derived retinol — using Affymetrix DNA microarrays. Transcriptomic changes were analyzed in two embryonic regions: anterior tissues including forebrain and optic vesicle, and posterior (trunk) tissues, at early stages preceding the appearance of overt phenotypic abnormalities. Several genes expected to be downregulated under RA deficiency appeared in the transcriptome data (e.g. Emx2, Foxg1 anteriorly, Cdx1, Hoxa1, Rarb posteriorly), whereas reverse-transcriptase-PCR and in situ hybridization performed for additional selected genes validated the changes identified through microarray analysis. Altogether, the affected genes belonged to numerous molecular pathways and cellular/organismal functions, demonstrating the pleiotropic nature of RA-dependent events. In both tissue samples, genes upregulated were more numerous than those downregulated, probably due to feedback regulatory loops. Bioinformatic analyses highlighted groups (clusters) of genes displaying similar behaviors in mutant tissues, and biological functions most significantly affected (e.g. mTOR, VEGF, ILK signaling in forebrain tissues; pyrimidine and purine metabolism

  18. Analysis of the brain mural cell transcriptome

    PubMed Central

    He, Liqun; Vanlandewijck, Michael; Raschperger, Elisabeth; Andaloussi Mäe, Maarja; Jung, Bongnam; Lebouvier, Thibaud; Ando, Koji; Hofmann, Jennifer; Keller, Annika; Betsholtz, Christer

    2016-01-01

    Pericytes, the mural cells of blood microvessels, regulate microvascular development and function and have been implicated in many brain diseases. However, due to a paucity of defining markers, pericyte identification and functional characterization remain ambiguous and data interpretation problematic. In mice carrying two transgenic reporters, Pdgfrb-eGFP and NG2-DsRed, we found that double-positive cells were vascular mural cells, while the single reporters marked additional, but non-overlapping, neuroglial cells. Double-positive cells were isolated by fluorescence-activated cell sorting (FACS) and analyzed by RNA sequencing. To reveal defining patterns of mural cell transcripts, we compared the RNA sequencing data with data from four previously published studies. The meta-analysis provided a conservative catalogue of 260 brain mural cell-enriched gene transcripts. We validated pericyte-specific expression of two novel markers, vitronectin (Vtn) and interferon-induced transmembrane protein 1 (Ifitm1), using fluorescent in situ hybridization and immunohistochemistry. We further analyzed signaling pathways and interaction networks of the pericyte-enriched genes in silico. This work provides novel insight into the molecular composition of brain mural cells. The reported gene catalogue facilitates identification of brain pericytes by providing numerous new candidate marker genes and is a rich source for new hypotheses for future studies of brain mural cell physiology and pathophysiology. PMID:27725773

  19. Transcriptomic Analysis of Calcium Remodeling in Colorectal Cancer.

    PubMed

    Pérez-Riesgo, Enrique; Gutiérrez, Lucía G; Ubierna, Daniel; Acedo, Alberto; Moyer, Mary P; Núñez, Lucía; Villalobos, Carlos

    2017-04-27

    Colorectal cancer (CRC) cells undergo the remodeling of intracellular Ca(2+) homeostasis, which contributes to cancer hallmarks such as enhanced proliferation, invasion and survival. Ca(2+) remodeling includes critical changes in store-operated Ca(2+) entry (SOCE) and Ca(2+) store content. Some changes have been investigated at the molecular level. However, since nearly 100 genes are involved in intracellular Ca(2+) transport, a comprehensive view of Ca(2+) remodeling in CRC is lacking. We have used Next Generation Sequencing (NGS) to investigate differences in expression of 77 selected gene transcripts involved in intracellular Ca(2+) transport in CRC. To this end, mRNA from normal human colonic NCM460 cells and human colon cancer HT29 cells was isolated and used as a template for transcriptomic sequencing and expression analysis using Ion Torrent technology. After data transformation and filtering, exploratory analysis revealed that both cell types were well segregated. In addition, differential gene expression using R and bioconductor packages show significant differences in expression of selected voltage-operated Ca(2+) channels and store-operated Ca(2+) entry players, transient receptor potential (TRP) channels, Ca(2+) release channels, Ca(2+) pumps, Na⁺/Ca(2+) exchanger isoforms and genes involved in mitochondrial Ca(2+) transport. These data provide the first comprehensive transcriptomic analysis of Ca(2+) remodeling in CRC.

  20. Transcriptomic Analysis of Calcium Remodeling in Colorectal Cancer

    PubMed Central

    Pérez-Riesgo, Enrique; Gutiérrez, Lucía G.; Ubierna, Daniel; Acedo, Alberto; Moyer, Mary P.; Núñez, Lucía; Villalobos, Carlos

    2017-01-01

    Colorectal cancer (CRC) cells undergo the remodeling of intracellular Ca2+ homeostasis, which contributes to cancer hallmarks such as enhanced proliferation, invasion and survival. Ca2+ remodeling includes critical changes in store-operated Ca2+ entry (SOCE) and Ca2+ store content. Some changes have been investigated at the molecular level. However, since nearly 100 genes are involved in intracellular Ca2+ transport, a comprehensive view of Ca2+ remodeling in CRC is lacking. We have used Next Generation Sequencing (NGS) to investigate differences in expression of 77 selected gene transcripts involved in intracellular Ca2+ transport in CRC. To this end, mRNA from normal human colonic NCM460 cells and human colon cancer HT29 cells was isolated and used as a template for transcriptomic sequencing and expression analysis using Ion Torrent technology. After data transformation and filtering, exploratory analysis revealed that both cell types were well segregated. In addition, differential gene expression using R and bioconductor packages show significant differences in expression of selected voltage-operated Ca2+ channels and store-operated Ca2+ entry players, transient receptor potential (TRP) channels, Ca2+ release channels, Ca2+ pumps, Na+/Ca2+ exchanger isoforms and genes involved in mitochondrial Ca2+ transport. These data provide the first comprehensive transcriptomic analysis of Ca2+ remodeling in CRC. PMID:28448473

  1. Leading edge analysis of transcriptomic changes during pseudorabies virus infection.

    PubMed

    Fleming, Damarius S; Miller, Laura C

    2016-12-01

    Eight RNA samples taken from the tracheobronchial lymph nodes (TBLN) of pigs that were either infected or non-infected with a feral isolate of porcine pseudorabies virus (PRV) were used to investigate changes in gene expression related to the pathogen. The RNA was processed into fastq files for each library prior to being analyzed using Illumina Digital Gene Expression Tag Profiling sequences (DGETP) which were used as the downstream measure of differential expression. Analyzed tags consisted of 21 base pair sequences taken from time points 1, 3, 6, and 14 days' post infection (dpi) that generated 1,927,547 unique tag sequences. Tag sequences were analyzed for differential transcript expression and gene set enrichment analysis (GSEA) to uncover transcriptomic changes related to PRV pathology progression. In conjunction with the DGETP and GSEA, the study also incorporated use of leading edge analysis to help link the TBLN transcriptome data to clinical progression of PRV at each of the sampled time points. The purpose of this manuscript is to provide useful background on applying the leading edge analysis to GSEA and expression data to help identify genes considered to be of high biological interest. The data in the form of fastq files has been uploaded to the NCBI Gene Expression Omnibus (GEO) (GSE74473) database.

  2. RNA-seq based transcriptomic analysis of single bacterial cells.

    PubMed

    Wang, Jiangxin; Chen, Lei; Chen, Zixi; Zhang, Weiwen

    2015-11-01

    Gene-expression heterogeneity among individual cells determines the fate of a bacterial population. Here we report the first bacterial single-cell RNA sequencing (RNA-seq), BaSiC RNA-seq, a method integrating RNA isolation, cDNA synthesis and amplification, and RNA-seq analysis of the whole transcriptome of single cyanobacterium Synechocystis sp. PCC 6803 cells which typically contain approximately 5-7 femtogram total RNA per cell. We applied the method to 3 Synechocystis single cells at 24 h and 3 single cells at 72 h after nitrogen-starvation stress treatment, as well as their bulk-cell controls under the same conditions, to determine the heterogeneity upon environmental stress. With 82-98% and 31-48% of all putative Synechocystis genes identified in single cells of 24 and 72 h, respectively, the results demonstrated that the method could achieve good identification of the transcripts in single bacterial cells. In addition, the preliminary results from nitrogen-starved cells also showed a possible increasing gene-expression heterogeneity from 24 h to 72 h after nitrogen starvation stress. Moreover, preliminary analysis of single-cell transcriptomic datasets revealed that genes from the "Mobile elements" functional category have the most significant increase of gene-expression heterogeneity upon stress, which was further confirmed by single-cell RT-qPCR analysis of gene expression in 24 randomly selected cells.

  3. Simultaneous Transcriptome Analysis of Sorghum and Bipolaris sorghicola by Using RNA-seq in Combination with De Novo Transcriptome Assembly

    PubMed Central

    Yazawa, Takayuki; Kawahigashi, Hiroyuki; Matsumoto, Takashi; Mizuno, Hiroshi

    2013-01-01

    The recent development of RNA sequencing (RNA-seq) technology has enabled us to analyze the transcriptomes of plants and their pathogens simultaneously. However, RNA-seq often relies on aligning reads to a reference genome and is thus unsuitable for analyzing most plant pathogens, as their genomes have not been fully sequenced. Here, we analyzed the transcriptomes of Sorghum bicolor (L.) Moench and its pathogen Bipolaris sorghicola simultaneously by using RNA-seq in combination with de novo transcriptome assembly. We sequenced the mixed transcriptome of the disease-resistant sorghum cultivar SIL-05 and B. sorghicola in infected leaves in the early stages of infection (12 and 24 h post-inoculation) by using Illumina mRNA-Seq technology. Sorghum gene expression was quantified by aligning reads to the sorghum reference genome. For B. sorghicola, reads that could not be aligned to the sorghum reference genome were subjected to de novo transcriptome assembly. We identified genes of B. sorghicola for growth of this fungus in sorghum, as well as genes in sorghum for the defense response. The genes of B. sorghicola included those encoding Woronin body major protein, LysM domain-containing intracellular hyphae protein, transcriptional factors CpcA and HacA, and plant cell-wall degrading enzymes. The sorghum genes included those encoding two receptors of the simple eLRR domain protein family, transcription factors that are putative orthologs of OsWRKY45 and OsWRKY28 in rice, and a class III peroxidase that is a homolog involved in disease resistance in the Poaceae. These defense-related genes were particularly strongly induced among paralogs annotated in the sorghum genome. Thus, in the absence of genome sequences for the pathogen, simultaneous transcriptome analysis of plant and pathogen by using de novo assembly was useful for identifying putative key genes in the plant–pathogen interaction. PMID:23638091

  4. Simultaneous transcriptome analysis of Sorghum and Bipolaris sorghicola by using RNA-seq in combination with de novo transcriptome assembly.

    PubMed

    Yazawa, Takayuki; Kawahigashi, Hiroyuki; Matsumoto, Takashi; Mizuno, Hiroshi

    2013-01-01

    The recent development of RNA sequencing (RNA-seq) technology has enabled us to analyze the transcriptomes of plants and their pathogens simultaneously. However, RNA-seq often relies on aligning reads to a reference genome and is thus unsuitable for analyzing most plant pathogens, as their genomes have not been fully sequenced. Here, we analyzed the transcriptomes of Sorghum bicolor (L.) Moench and its pathogen Bipolaris sorghicola simultaneously by using RNA-seq in combination with de novo transcriptome assembly. We sequenced the mixed transcriptome of the disease-resistant sorghum cultivar SIL-05 and B. sorghicola in infected leaves in the early stages of infection (12 and 24 h post-inoculation) by using Illumina mRNA-Seq technology. Sorghum gene expression was quantified by aligning reads to the sorghum reference genome. For B. sorghicola, reads that could not be aligned to the sorghum reference genome were subjected to de novo transcriptome assembly. We identified genes of B. sorghicola for growth of this fungus in sorghum, as well as genes in sorghum for the defense response. The genes of B. sorghicola included those encoding Woronin body major protein, LysM domain-containing intracellular hyphae protein, transcriptional factors CpcA and HacA, and plant cell-wall degrading enzymes. The sorghum genes included those encoding two receptors of the simple eLRR domain protein family, transcription factors that are putative orthologs of OsWRKY45 and OsWRKY28 in rice, and a class III peroxidase that is a homolog involved in disease resistance in the Poaceae. These defense-related genes were particularly strongly induced among paralogs annotated in the sorghum genome. Thus, in the absence of genome sequences for the pathogen, simultaneous transcriptome analysis of plant and pathogen by using de novo assembly was useful for identifying putative key genes in the plant-pathogen interaction.

  5. Transcriptome Analysis of the Oriental Fruit Fly (Bactrocera dorsalis)

    PubMed Central

    Jiang, Hong-Bo; Yang, Wen-Jia; Jia, Fu-Xian; Hu, Fei; Cong, Lin; Wang, Jin-Jun

    2011-01-01

    Background The oriental fruit fly, Bactrocera dorsalis (Hendel), is one of the most economically important pests in the world, causing serious damage to fruit production. However, lack of genetic information on this organism is an obstacle to understanding the mechanisms behind its development and its ability to resist insecticides. Analysis of the B. dorsalis transcriptome and its expression profile data is essential to extending the genetic information resources on this species, providing a shortcut that will support studies on B. dorsalis. Methodology/Principal Findings We performed de novo assembly of a transcriptome using short read sequencing technology (Illumina). The results generated 484,628 contigs, 70,640 scaffolds, and 49,804 unigenes. Of those unigenes, 27,455 (55.13%) matched known proteins in the NCBI database, as determined by BLAST search. Clusters of orthologous groups (COG), gene orthology (GO), and the Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations were performed to better understand the functions of these unigenes. Genes related to insecticide resistance were analyzed in additional detail. Digital gene expression (DGE) libraries showed differences in gene expression profiles at different developmental stages (eggs, third-instar larvae, pupae, and adults). To confirm the DGE results, the expression profiles of six randomly selected genes were analyzed. Conclusion/Significance This transcriptome greatly improves our genetic understanding of B. dorsalis and makes a huge number of gene sequences available for further study, including both genes of known importance and genes of unknown function. The DGE data provide comprehensive insight into gene expression profiles at different developmental stages. This facilitates the study of the role of each gene in the developmental process and in insecticide resistance. PMID:22195006

  6. Analysis of the transcriptome of the Indonesian coelacanth Latimeria menadoensis

    PubMed Central

    2013-01-01

    Background Latimeria menadoensis is a coelacanth species first identified in 1997 in Indonesia, at 10,000 Km of distance from its African congener. To date, only six specimens have been caught and just a very limited molecular data is available. In the present work we describe the de novo transcriptome assembly obtained from liver and testis samples collected from the fifth specimen ever caught of this species. Results The deep RNA sequencing performed with Illumina technologies generated 145,435,156 paired-end reads, accounting for ~14 GB of sequence data, which were de novo assembled using a Trinity/CLC combined strategy. The assembly output was processed and filtered producing a set of 66,308 contigs, whose quality was thoroughly assessed. The comparison with the recently sequenced genome of the African congener Latimeria chalumnae and with the available genomic resources of other vertebrates revealed a good reconstruction of full length transcripts and a high coverage of the predicted full coelacanth transcriptome. The RNA-seq analysis revealed remarkable differences in the expression profiles between the two tissues, allowing the identification of liver- and testis-specific transcripts which may play a fundamental role in important biological processes carried out by these two organs. Conclusion Given the high genomic affinity between the two coelacanth species, the here described de novo transcriptome assembly can be considered a valuable support tool for the improvement of gene prediction within the genome of L. chalumnae and a valuable resource for investigation of many aspects of tetrapod evolution. PMID:23927401

  7. Transcriptomic analysis of heteromorphic stamens in Cassia biscapsularis L.

    PubMed Central

    Luo, Zhonglai; Hu, Jin; Zhao, Zhongtao; Zhang, Dianxiang

    2016-01-01

    Hermaphroditic flowers have evolved primarily under the selection on male function. Evolutionary modification often leads to stamen differentiation within flowers, or “heteranthery”, a phenomenon intrigued scientists since the 18th century until recently. However, the genetic basis and molecular regulation mechanism has barely been touched. Here we conducted comparative transcriptome profiling in Cassia biscapsularis L., a heterantherous species with representative patterns of stamen differentiation. Numerous differentially expressed genes (DEGs) were detected between the staminodes (the degenerated stamens) and fertile stamens, while much fewer genes differentially expressed among the three sets of fertile stamens. GO term enrichment and KEGG pathway analysis characterized functional properties of DEGs in different stamen types. Transcripts showing close correlation between expression pattern and stamen types were identified. Transcription factors from the bHLH family were suggested to have taken crucial part in the formation of staminodes. This first global transcriptomic analysis focusing on stamen differentiation opens the door toward a more comprehensive understanding on the molecular regulation of floral organ evolution. Especially, the generated unigene resource would be valuable for developing male sterile lines in agronomy. PMID:27527392

  8. Venomic and transcriptomic analysis of centipede Scolopendra subspinipes dehaani.

    PubMed

    Liu, Zi-Chao; Zhang, Rong; Zhao, Feng; Chen, Zhong-Ming; Liu, Hao-Wen; Wang, Yan-Jie; Jiang, Ping; Zhang, Yong; Wu, Ying; Ding, Jiu-Ping; Lee, Wen-Hui; Zhang, Yun

    2012-12-07

    Centipedes have venom glands in their first pair of limbs, and their venoms contain a large number of components with different biochemical and pharmacological properties. However, information about the compositions and functions of their venoms is largely unknown. In this study, Scolopendra subspinipes dehaani venoms were systematically investigated by transcriptomic and proteomic analysis coupled with biological function assays. After random screening approximately 1500 independent clones, 1122 full length cDNA sequences, which encode 543 different proteins, were cloned from a constructed cDNA library using a pair of venom glands from a single centipede species. Neurotoxins, ion channel acting components and venom allergens were the main fractions of the crude venom as revealed by transcriptomic analysis. Meanwhile, 40 proteins/peptides were purified and characterized from crude venom of S. subspinipes dehaani. The N-terminal amino acid sequencing and mass spectrum results of 29 out of these 40 proteins or peptides matched well with their corresponding cDNAs. The purified proteins/peptides showed different pharmacological properties, including the following: (1) platelet aggregating activity; (2) anticoagulant activity; (3) phospholipase A(2) activity; (4) trypsin inhibiting activity; (5) voltage-gated potassium channel activities; (6) voltage-gated sodium channel activities; (7) voltage-gated calcium channel activities. Most of them showed no significant similarity to other protein sequences deposited in the known public database. This work provides the largest number of protein or peptide candidates with medical-pharmaceutical significance and reveals the toxin nature of centipede S. subspinipes dehaani venom.

  9. Transcriptome analysis of the Octopus vulgaris central nervous system.

    PubMed

    Zhang, Xiang; Mao, Yong; Huang, Zixia; Qu, Meng; Chen, Jun; Ding, Shaoxiong; Hong, Jingni; Sun, Tiantian

    2012-01-01

    Cephalopoda are a class of Mollusca species found in all the world's oceans. They are an important model organism in neurobiology. Unfortunately, the lack of neuronal molecular sequences, such as ESTs, transcriptomic or genomic information, has limited the development of molecular neurobiology research in this unique model organism. With high-throughput Illumina Solexa sequencing technology, we have generated 59,859 high quality sequences from 12,918,391 paired-end reads. Using BLASTx/BLASTn, 12,227 contigs have blast hits in the Swissprot, NR protein database and NT nucleotide database with E-value cutoff 1e(-5). The comparison between the Octopus vulgaris central nervous system (CNS) library and the Aplysia californica/Lymnaea stagnalis CNS ESTs library yielded 5.93%/13.45% of O. vulgaris sequences with significant matches (1e(-5)) using BLASTn/tBLASTx. Meanwhile the hit percentage of the recently published Schistocerca gregaria, Tilapia or Hirudo medicinalis CNS library to the O. vulgaris CNS library is 21.03%-46.19%. We constructed the Phylogenetic tree using two genes related to CNS function, Synaptotagmin-7 and Synaptophysin. Lastly, we demonstrated that O. vulgaris may have a vertebrate-like Blood-Brain Barrier based on bioinformatic analysis. This study provides a mass of molecular information that will contribute to further molecular biology research on O. vulgaris. In our presentation of the first CNS transcriptome analysis of O. vulgaris, we hope to accelerate the study of functional molecular neurobiology and comparative evolutionary biology.

  10. 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

  11. Transcriptome Analysis of Oryza sativa Calli Under Microgravity

    NASA Astrophysics Data System (ADS)

    Jin, Jing; Chen, Haiying; Cai, Weiming

    2015-11-01

    The transcriptome of Oryza sativacalli was analyzed on board the Chinese spaceship "Shenzhou 8" to study the effects of microgravity on plant signal transduction and secondary metabolism (as one of the experiments with SIMBOX on Shenzhou 8). Calli of Oryza sativa were pre-cultured for 4 days on ground and then loaded into the stationary platform or the rotating platform of a biological incubator, called SIMBOX, to grow in space under microgravity conditions or 1g-conditions, respectively. The calli were fixed by RNAlater after grew 324 h under microgravity. After 17 days, Shenzhou 8 returned to Earth carrying SIMBOX. Oryza sativa calli were recovered, and the RNA was extracted for transcriptome analysis. After comparing 1 gspaceflight controls-inflight controls with 1 g-ground controls, 157 probe sets with different expression levels (fold change ≥2, p<0.05) were identified. When comparing spaceflight controls to 1 g-ground controls and to 1 g-inflight controls, 678 probe sets with different expression levels (fold change ≥2, p<0.05) were identified. The fact that the same 678 probe sets were identified in these two comparisons suggests that transcription was affected under microgravity conditions. MapMan analysis was used to classify 627 microgravity responsive (MR) transcripts. The MR transcripts were mainly involved in cell wall structure, the TCA cycle, primary metabolism, transcription, protein modification and degradation, hormone metabolism, calcium regulation, receptor like kinase activity and transport.

  12. Stress-induced and epigenetic-mediated maize transcriptome regulation study by means of transcriptome reannotation and differential expression analysis

    PubMed Central

    Forestan, Cristian; Aiese Cigliano, Riccardo; Farinati, Silvia; Lunardon, Alice; Sanseverino, Walter; Varotto, Serena

    2016-01-01

    Plant’s response and adaptation to abiotic stresses involve sophisticated genetic and epigenetic regulatory systems. To obtain a global view of molecular response to osmotic stresses, including the non-coding portion of genome, we conducted a total leaf transcriptome analysis on maize plants subjected to prolonged drought and salt stresses. Stress application to both B73 wild type and the epiregulator mutant rpd1-1/rmr6 allowed dissection of the epigenetic component of stress response. Coupling total RNA-Seq and transcriptome re-assembly we annotated thousands of new maize transcripts, together with 13,387 lncRNAs that may play critical roles in regulating gene expression. Differential expression analysis revealed hundreds of genes modulated by long-term stress application, including also many lncRNAs and transposons specifically induced by stresses. The amplitude and dynamic of the stress-modulated gene sets are very different between B73 and rpd1-1/rmr6 mutant plants, as result of stress-like effect on genome regulation caused by the mutation itself, which activates many stress-related genes even in control condition. The analyzed extensive set of total RNA-Seq data, together with the improvement of the transcriptome and the identification of the non-coding portion of the transcriptome give a revealing insight into the genetic and epigenetic mechanism responsible for maize molecular response to abiotic stresses. PMID:27461139

  13. Pesticide toxicogenomics across scales: in vitro transcriptome predicts mechanisms and outcomes of exposure in vivo

    PubMed Central

    Porreca, Immacolata; D’Angelo, Fulvio; De Franceschi, Lucia; Mattè, Alessandro; Ceccarelli, Michele; Iolascon, Achille; Zamò, Alberto; Russo, Filomena; Ravo, Maria; Tarallo, Roberta; Scarfò, Marzia; Weisz, Alessandro; De Felice, Mario; Mallardo, Massimo; Ambrosino, Concetta

    2016-01-01

    In vitro Omics analysis (i.e. transcriptome) is suggested to predict in vivo toxicity and adverse effects in humans, although the causal link between high-throughput data and effects in vivo is not easily established. Indeed, the chemical-organism interaction can involve processes, such as adaptation, not established in cell cultures. Starting from this consideration we investigate the transcriptomic response of immortalized thyrocytes to ethylenthiourea and chlorpyrifos. In vitro data revealed specific and common genes/mechanisms of toxicity, controlling the proliferation/survival of the thyrocytes and unrelated hematopoietic cell lineages. These results were phenotypically confirmed in vivo by the reduction of circulating T4 hormone and the development of pancytopenia after long exposure. Our data imply that in vitro toxicogenomics is a powerful tool in predicting adverse effects in vivo, experimentally confirming the vision described as Tox21c (Toxicity Testing in the 21st century) although not fully recapitulating the biocomplexity of a living animal. PMID:27905518

  14. MCbiclust: a novel algorithm to discover large-scale functionally related gene sets from massive transcriptomics data collections.

    PubMed

    Bentham, Robert B; Bryson, Kevin; Szabadkai, Gyorgy

    2017-09-06

    The potential to understand fundamental biological processes from gene expression data has grown in parallel with the recent explosion of the size of data collections. However, to exploit this potential, novel analytical methods are required, capable of discovering large co-regulated gene networks. We found current methods limited in the size of correlated gene sets they could discover within biologically heterogeneous data collections, hampering the identification of multi-gene controlled fundamental cellular processes such as energy metabolism, organelle biogenesis and stress responses. Here we describe a novel biclustering algorithm called Massively Correlated Biclustering (MCbiclust) that selects samples and genes from large datasets with maximal correlated gene expression, allowing regulation of complex networks to be examined. The method has been evaluated using synthetic data and applied to large bacterial and cancer cell datasets. We show that the large biclusters discovered, so far elusive to identification by existing techniques, are biologically relevant and thus MCbiclust has great potential in the analysis of transcriptomics data to identify large-scale unknown effects hidden within the data. The identified massive biclusters can be used to develop improved transcriptomics based diagnosis tools for diseases caused by altered gene expression, or used for further network analysis to understand genotype-phenotype correlations. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  15. A Comprehensive Transcriptomic and Proteomic Analysis of Hydra Head Regeneration

    PubMed Central

    Petersen, Hendrik O.; Höger, Stefanie K.; Looso, Mario; Lengfeld, Tobias; Kuhn, Anne; Warnken, Uwe; Nishimiya-Fujisawa, Chiemi; Schnölzer, Martina; Krüger, Marcus; Özbek, Suat; Simakov, Oleg; Holstein, Thomas W.

    2015-01-01

    The cnidarian freshwater polyp Hydra sp. exhibits an unparalleled regeneration capacity in the animal kingdom. Using an integrative transcriptomic and stable isotope labeling by amino acids in cell culture proteomic/phosphoproteomic approach, we studied stem cell-based regeneration in Hydra polyps. As major contributors to head regeneration, we identified diverse signaling pathways adopted for the regeneration response as well as enriched novel genes. Our global analysis reveals two distinct molecular cascades: an early injury response and a subsequent, signaling driven patterning of the regenerating tissue. A key factor of the initial injury response is a general stabilization of proteins and a net upregulation of transcripts, which is followed by a subsequent activation cascade of signaling molecules including Wnts and transforming growth factor (TGF) beta-related factors. We observed moderate overlap between the factors contributing to proteomic and transcriptomic responses suggesting a decoupled regulation between the transcriptional and translational levels. Our data also indicate that interstitial stem cells and their derivatives (e.g., neurons) have no major role in Hydra head regeneration. Remarkably, we found an enrichment of evolutionarily more recent genes in the early regeneration response, whereas conserved genes are more enriched in the late phase. In addition, genes specific to the early injury response were enriched in transposon insertions. Genetic dynamicity and taxon-specific factors might therefore play a hitherto underestimated role in Hydra regeneration. PMID:25841488

  16. Genome-wide transcriptome analysis of human epidermal melanocytes

    PubMed Central

    Haltaufderhyde, Kirk D.; Oancea, Elena

    2015-01-01

    Because human epidermal melanocytes (HEMs) provide critical protection against skin cancer, sunburn, and photoaging, a genome-wide perspective of gene expression in these cells is vital to understanding human skin physiology. In this study we performed high throughput sequencing of HEMs to obtain a complete data set of transcript sizes, abundances, and splicing. As expected, we found that melanocyte specific genes that function in pigmentation were among the highest expressed genes. We analyzed receptor, ion channel and transcription factor gene families to get a better understanding of the cell signalling pathways used by melanocytes. We also performed a comparative transcriptomic analysis of lightly versus darkly pigmented HEMs and found 16 genes differentially expressed in the two pigmentation phenotypes; of those, only one putative melanosomal transporter (SLC45A2) has known function in pigmentation. In addition, we found 166 genes with splice isoforms expressed exclusively in one pigmentation phenotype, 17 of which are genes involved in signal transduction. Our melanocyte transcriptome study provides a comprehensive view and may help identify novel pigmentation genes and potential pharmacological targets. PMID:25451175

  17. Differential Transcriptome Analysis between Paulownia fortunei and Its Synthesized Autopolyploid

    PubMed Central

    Zhang, Xiaoshen; Deng, Minjie; Fan, Guoqiang

    2014-01-01

    Paulownia fortunei is an ecologically and economically important tree species that is widely used as timber and chemical pulp. Its autotetraploid, which carries a number of valuable traits, was successfully induced with colchicine. To identify differences in gene expression between P. fortunei and its synthesized autotetraploid, we performed transcriptome sequencing using an Illumina Genome Analyzer IIx (GAIIx). About 94.8 million reads were generated and assembled into 383,056 transcripts, including 18,984 transcripts with a complete open reading frame. A conducted Basic Local Alignment Search Tool (BLAST) search indicated that 16,004 complete transcripts had significant hits in the National Center for Biotechnology Information (NCBI) non-redundant database. The complete transcripts were given functional assignments using three public protein databases. One thousand one hundred fifty eight differentially expressed complete transcripts were screened through a digital abundance analysis, including transcripts involved in energy metabolism and epigenetic regulation. Finally, the expression levels of several transcripts were confirmed by quantitative real-time PCR. Our results suggested that polyploidization caused epigenetic-related changes, which subsequently resulted in gene expression variation between diploid and autotetraploid P. fortunei. This might be the main mechanism affected by the polyploidization. Our results represent an extensive survey of the P. fortunei transcriptome and will facilitate subsequent functional genomics research in P. fortunei. Moreover, the gene expression profiles of P. fortunei and its autopolyploid will provide a valuable resource for the study of polyploidization. PMID:24663058

  18. De novo assembly and analysis of crow lungs transcriptome.

    PubMed

    Vijayakumar, Periyasamy; Raut, Ashwin Ashok; Kumar, Pushpendra; Sharma, Deepak; Mishra, Anamika

    2014-09-01

    The jungle crow (Corvus macrorhynchos) belongs to the order Passeriformes of bird species and is important for avian ecological and evolutionary genetics studies. However, there is limited information on the transcriptome data of this species. In the present study, we report the characterization of the lung transcriptome of the jungle crow using GS FLX Titanium XLR70. Altogether, 1,510,303 high-quality sequence reads with 581,198,230 bases was de novo assembled into 22,169 isotigs (isotig represents an individual transcript) and 784,009 singletons. Using these isotigs and 581,681 length-filtered (greater than 300 bp) singletons, 20,010 unique protein-coding genes were identified by BLASTx comparison against a nonredundant (nr) protein sequence database. Comparative analysis revealed that 46,604 (70.29%) and 51,642 (72.48%) of the assembled transcripts have significant similarity to zebra finch and chicken RefSeq proteins, respectively. As determined by GO annotation and KEGG pathway mapping, functional annotation of the unigenes recovered diverse biological functions and processes. Transcripts putatively involved in the immune response were identified. Furthermore, 20,599 single nucleotide polymorphisms (SNPs) and 7525 simple sequence repeats (SSRs) were retrieved from the assembled transcript database. This resource should lay an important base for future ecological, evolutionary, and conservation genetic studies on this species and in other related species.

  19. Comparative transcriptome analysis of the metal hyperaccumulator Noccaea caerulescens

    PubMed Central

    Halimaa, Pauliina; Blande, Daniel; Aarts, Mark G. M.; Tuomainen, Marjo; Tervahauta, Arja; Kärenlampi, Sirpa

    2014-01-01

    The metal hyperaccumulator Noccaea caerulescens is an established model to study the adaptation of plants to metalliferous soils. Various comparators have been used in these studies. The choice of suitable comparators is important and depends on the hypothesis to be tested and methods to be used. In high-throughput analyses such as microarray, N. caerulescens has been compared to non-tolerant, non-accumulator plants like Arabidopsis thaliana or Thlaspi arvense rather than to the related hypertolerant or hyperaccumulator plants. An underutilized source is N. caerulescens populations with considerable variation in their capacity to accumulate and tolerate metals. Whole transcriptome sequencing (RNA-Seq) is revealing interesting variation in their gene expression profiles. Combining physiological characteristics of N. caerulescens accessions with their RNA-Seq has a great potential to provide detailed insight into the underlying molecular mechanisms, including entirely new gene products. In this review we will critically consider comparative transcriptome analyses carried out to explore metal hyperaccumulation and hypertolerance of N. caerulescens, and demonstrate the potential of RNA-Seq analysis as a tool in evolutionary genomics. PMID:24904610

  20. Transcriptome analysis of Haloquadratum walsbyi: vanity is but the surface.

    PubMed

    Bolhuis, Henk; Martín-Cuadrado, Ana Belén; Rosselli, Riccardo; Pašić, Lejla; Rodriguez-Valera, Francisco

    2017-07-03

    Haloquadratum walsbyi dominates saturated thalassic lakes worldwide where they can constitute up to 80-90% of the total prokaryotic community. Despite the abundance of the enigmatic square-flattened cells, only 7 isolates are currently known with 2 genomes fully sequenced and annotated due to difficulties to grow them under laboratory conditions. We have performed a transcriptomic analysis of one of these isolates, the Spanish strain HBSQ001 in order to investigate gene transcription under light and dark conditions. Despite a potential advantage for light as additional source of energy, no significant differences were found between light and dark expressed genes. Constitutive high gene expression was observed in genes encoding surface glycoproteins, light mediated proton pumping by bacteriorhodopsin, several nutrient uptake systems, buoyancy and storage of excess carbon. Two low expressed regions of the genome were characterized by a lower codon adaptation index, low GC content and high incidence of hypothetical genes. Under the extant cultivation conditions, the square hyperhalophile devoted most of its transcriptome towards processes maintaining cell integrity and exploiting solar energy. Surface glycoproteins are essential for maintaining the large surface to volume ratio that facilitates light and organic nutrient harvesting whereas constitutive expression of bacteriorhodopsin warrants an immediate source of energy when light becomes available.

  1. A Comprehensive Transcriptomic and Proteomic Analysis of Hydra Head Regeneration.

    PubMed

    Petersen, Hendrik O; Höger, Stefanie K; Looso, Mario; Lengfeld, Tobias; Kuhn, Anne; Warnken, Uwe; Nishimiya-Fujisawa, Chiemi; Schnölzer, Martina; Krüger, Marcus; Özbek, Suat; Simakov, Oleg; Holstein, Thomas W

    2015-08-01

    The cnidarian freshwater polyp Hydra sp. exhibits an unparalleled regeneration capacity in the animal kingdom. Using an integrative transcriptomic and stable isotope labeling by amino acids in cell culture proteomic/phosphoproteomic approach, we studied stem cell-based regeneration in Hydra polyps. As major contributors to head regeneration, we identified diverse signaling pathways adopted for the regeneration response as well as enriched novel genes. Our global analysis reveals two distinct molecular cascades: an early injury response and a subsequent, signaling driven patterning of the regenerating tissue. A key factor of the initial injury response is a general stabilization of proteins and a net upregulation of transcripts, which is followed by a subsequent activation cascade of signaling molecules including Wnts and transforming growth factor (TGF) beta-related factors. We observed moderate overlap between the factors contributing to proteomic and transcriptomic responses suggesting a decoupled regulation between the transcriptional and translational levels. Our data also indicate that interstitial stem cells and their derivatives (e.g., neurons) have no major role in Hydra head regeneration. Remarkably, we found an enrichment of evolutionarily more recent genes in the early regeneration response, whereas conserved genes are more enriched in the late phase. In addition, genes specific to the early injury response were enriched in transposon insertions. Genetic dynamicity and taxon-specific factors might therefore play a hitherto underestimated role in Hydra regeneration. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  2. A Comprehensive Analysis of Metabolomics and Transcriptomics in Cervical Cancer

    PubMed Central

    Yang, Kai; Xia, Bairong; Wang, Wenjie; Cheng, Jinlong; Yin, Mingzhu; Xie, Hongyu; Li, Junnan; Ma, Libing; Yang, Chunyan; Li, Ang; Fan, Xin; Dhillon, Harman S.; Hou, Yan; Lou, Ge; Li, Kang

    2017-01-01

    Cervical cancer (CC) still remains a common and deadly malignancy among females in developing countries. More accurate and reliable diagnostic methods/biomarkers should be discovered. In this study, we performed a comprehensive analysis of metabolomics (285 samples) and transcriptomics (52 samples) on the potential diagnostic implication and metabolic characteristic description in cervical cancer. Sixty-two metabolites were different between CC and normal controls (NOR), in which 5 metabolites (bilirubin, LysoPC(17:0), n-oleoyl threonine, 12-hydroxydodecanoic acid and tetracosahexaenoic acid) were selected as candidate biomarkers for CC. The AUC value, sensitivity (SE), and specificity (SP) of these 5 biomarkers were 0.99, 0.98 and 0.99, respectively. We further analysed the genes in 7 significantly enriched pathways, of which 117 genes, that were expressed differentially, were mainly involved in catalytic activity. Finally, a fully connected network of metabolites and genes in these pathways was built, which can increase the credibility of our selected metabolites. In conclusion, our biomarkers from metabolomics could set a path for CC diagnosis and screening. Our results also showed that variables of both transcriptomics and metabolomics were associated with CC. PMID:28225065

  3. Transcriptome Comparison Analysis of Ostrinia furnacalis in Four Developmental Stages

    PubMed Central

    Zhang, Tiantao; He, Kanglai; Wang, Zhenying

    2016-01-01

    The Asian corn borer, Ostrinia furnacalis, is one of the most destructive pests of maize and causes huge losses in maize yield each year. In order to characterize the different developmental stages, a high-throughput sequencing platform was employed to perform de novo transcriptome assembly and gene expression analysis for the egg, larva, pupa and adult stages. Approximately 185 million reads were obtained, trimmed, and assembled into 42,638 unigenes with an average length of 801.94 bp and an N50 length of 1,152 bp. These unigene sequences were annotated and classified by performing Gene Ontology (GO), Cluster of Orthologous Groups (KOG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional classifications. Comparison of the gene expression profiles of the two transitional stages revealed dramatic differences. Some differentially expressed genes are associated with digestion, cuticularization olfactory recognition and wing formation as well as growth and development. In total, 12 putative insect development-related genes were identified. Real-time quantitative PCR (RT-qPCR) results and sequencing based on relative expression levels of randomly selected genes confirmed these expression patterns. These data represent the most comprehensive transcriptomic resource currently available for O. furnacalis and will facilitate the study of developmental pathways, cuticularization, wing formation and olfactory recognition. PMID:27713521

  4. An evaluation of transcriptome-based exon capture for frog phylogenomics across multiple scales of divergence (Class: Amphibia, Order: Anura).

    PubMed

    Portik, Daniel M; Smith, Lydia L; Bi, Ke

    2016-09-01

    Custom sequence capture experiments are becoming an efficient approach for gathering large sets of orthologous markers in nonmodel organisms. Transcriptome-based exon capture utilizes transcript sequences to design capture probes, typically using a reference genome to identify intron-exon boundaries to exclude shorter exons (<200 bp). Here, we test directly using transcript sequences for probe design, which are often composed of multiple exons of varying lengths. Using 1260 orthologous transcripts, we conducted sequence captures across multiple phylogenetic scales for frogs, including outgroups ~100 Myr divergent from the ingroup. We recovered a large phylogenomic data set consisting of sequence alignments for 1047 of the 1260 transcriptome-based loci (~561 000 bp) and a large quantity of highly variable regions flanking the exons in transcripts (~70 000 bp), the latter improving substantially by only including ingroup species (~797 000 bp). We recovered both shorter (<100 bp) and longer exons (>200 bp), with no major reduction in coverage towards the ends of exons. We observed significant differences in the performance of blocking oligos for target enrichment and nontarget depletion during captures, and differences in PCR duplication rates resulting from the number of individuals pooled for capture reactions. We explicitly tested the effects of phylogenetic distance on capture sensitivity, specificity, and missing data, and provide a baseline estimate of expectations for these metrics based on a priori knowledge of nuclear pairwise differences among samples. We provide recommendations for transcriptome-based exon capture design based on our results, cost estimates and offer multiple pipelines for data assembly and analysis.

  5. Integrated Analysis of Transcriptomic and Proteomic Datasets Reveals Information on Protein Expressivity and Factors Affecting Translational Efficiency.

    PubMed

    Wang, Jiangxin; Wu, Gang; Chen, Lei; Zhang, Weiwen

    2016-01-01

    Integrated analysis of large-scale transcriptomic and proteomic data can provide important insights into the metabolic mechanisms underlying complex biological systems. In this chapter, we present methods to address two aspects of issues related to integrated transcriptomic and proteomic analysis. First, due to the fact that proteomic datasets are often incomplete, and integrated analysis of partial proteomic data may introduce significant bias. To address these issues, we describe a zero-inflated Poisson (ZIP)-based model to uncover the complicated relationships between protein abundances and mRNA expression levels, and then apply them to predict protein abundance for the proteins not experimentally detected. The ZIP model takes into consideration the undetected proteins by assuming that there is a probability mass at zero representing expressed proteins that were undetected owing to technical limitations. The model validity is demonstrated using biological information of operons, regulons, and pathways. Second, weak correlation between transcriptomic and proteomic datasets is often due to biological factors affecting translational processes. To quantify the effects of these factors, we describe a multiple regression-based statistical framework to quantitatively examine the effects of various translational efficiency-related sequence features on mRNA-protein correlation. Using the datasets from sulfate-reducing bacteria Desulfovibrio vulgaris, the analysis shows that translation-related sequence features can contribute up to 15.2-26.2% of the total variation of the correlation between transcriptomic and proteomic datasets, and also reveals the relative importance of various features in translation process.

  6. Single-cell transcriptome analysis of endometrial tissue.

    PubMed

    Krjutškov, K; Katayama, S; Saare, M; Vera-Rodriguez, M; Lubenets, D; Samuel, K; Laisk-Podar, T; Teder, H; Einarsdottir, E; Salumets, A; Kere, J

    2016-04-01

    How can we study the full transcriptome of endometrial stromal and epithelial cells at the single-cell level? By compiling and developing novel analytical tools for biopsy, tissue cryopreservation and disaggregation, single-cell sorting, library preparation, RNA sequencing (RNA-seq) and statistical data analysis. Although single-cell transcriptome analyses from various biopsied tissues have been published recently, corresponding protocols for human endometrium have not been described. The frozen-thawed endometrial biopsies were fluorescence-activated cell sorted (FACS) to distinguish CD13-positive stromal and CD9-positive epithelial cells and single-cell transcriptome analysis performed from biopsied tissues without culturing the cells. We studied gene transcription, applying a modern and efficient RNA-seq protocol. In parallel, endometrial stromal cells were cultured and global expression profiles were compared with uncultured cells. For method validation, we used two endometrial biopsies, one from mid-secretory phase (Day 21, LH+8) and another from late-secretory phase (Day 25). The samples underwent single-cell FACS sorting, single-cell RNA-seq library preparation and Illumina sequencing. Here we present a complete pipeline for single-cell gene-expression studies, from clinical sampling to statistical data analysis. Tissue manipulation, starting from disaggregation and cell-type-specific labelling and ending with single-cell automated sorting, is managed within 90 min at low temperature to minimize changes in the gene expression profile. The single living stromal and epithelial cells were sorted using CD13- and CD9-specific antibodies, respectively. Of the 8622 detected genes, 2661 were more active in cultured stromal cells than in biopsy cells. In the comparison of biopsy versus cultured cells, 5603 commonly expressed genes were detected, with 241 significantly differentially expressed genes. Of these, 231 genes were up- and 10 down-regulated in cultured cells

  7. Single-cell transcriptome analysis of endometrial tissue

    PubMed Central

    Krjutškov, K.; Katayama, S.; Saare, M.; Vera-Rodriguez, M.; Lubenets, D.; Samuel, K.; Laisk-Podar, T.; Teder, H.; Einarsdottir, E.; Salumets, A.; Kere, J.

    2016-01-01

    STUDY QUESTION How can we study the full transcriptome of endometrial stromal and epithelial cells at the single-cell level? SUMMARY ANSWER By compiling and developing novel analytical tools for biopsy, tissue cryopreservation and disaggregation, single-cell sorting, library preparation, RNA sequencing (RNA-seq) and statistical data analysis. WHAT IS KNOWN ALREADY Although single-cell transcriptome analyses from various biopsied tissues have been published recently, corresponding protocols for human endometrium have not been described. STUDY DESIGN, SIZE, DURATION The frozen-thawed endometrial biopsies were fluorescence-activated cell sorted (FACS) to distinguish CD13-positive stromal and CD9-positive epithelial cells and single-cell transcriptome analysis performed from biopsied tissues without culturing the cells. We studied gene transcription, applying a modern and efficient RNA-seq protocol. In parallel, endometrial stromal cells were cultured and global expression profiles were compared with uncultured cells. PARTICIPANTS/MATERIALS, SETTING, METHODS For method validation, we used two endometrial biopsies, one from mid-secretory phase (Day 21, LH+8) and another from late-secretory phase (Day 25). The samples underwent single-cell FACS sorting, single-cell RNA-seq library preparation and Illumina sequencing. MAIN RESULTS AND THE ROLE OF CHANCE Here we present a complete pipeline for single-cell gene-expression studies, from clinical sampling to statistical data analysis. Tissue manipulation, starting from disaggregation and cell-type-specific labelling and ending with single-cell automated sorting, is managed within 90 min at low temperature to minimize changes in the gene expression profile. The single living stromal and epithelial cells were sorted using CD13- and CD9-specific antibodies, respectively. Of the 8622 detected genes, 2661 were more active in cultured stromal cells than in biopsy cells. In the comparison of biopsy versus cultured cells, 5603

  8. Large-scale transcriptome sequencing and gene analyses in the crab-eating macaque (Macaca fascicularis) for biomedical research

    PubMed Central

    2012-01-01

    Background As a human replacement, the crab-eating macaque (Macaca fascicularis) is an invaluable non-human primate model for biomedical research, but the lack of genetic information on this primate has represented a significant obstacle for its broader use. Results Here, we sequenced the transcriptome of 16 tissues originated from two individuals of crab-eating macaque (male and female), and identified genes to resolve the main obstacles for understanding the biological response of the crab-eating macaque. From 4 million reads with 1.4 billion base sequences, 31,786 isotigs containing genes similar to those of humans, 12,672 novel isotigs, and 348,160 singletons were identified using the GS FLX sequencing method. Approximately 86% of human genes were represented among the genes sequenced in this study. Additionally, 175 tissue-specific transcripts were identified, 81 of which were experimentally validated. In total, 4,314 alternative splicing (AS) events were identified and analyzed. Intriguingly, 10.4% of AS events were associated with transposable element (TE) insertions. Finally, investigation of TE exonization events and evolutionary analysis were conducted, revealing interesting phenomena of human-specific amplified trends in TE exonization events. Conclusions This report represents the first large-scale transcriptome sequencing and genetic analyses of M. fascicularis and could contribute to its utility for biomedical research and basic biology. PMID:22554259

  9. Analysis of Whole Transcriptome Sequencing Data: Workflow and Software

    PubMed Central

    Yang, In Seok

    2015-01-01

    RNA is a polymeric molecule implicated in various biological processes, such as the coding, decoding, regulation, and expression of genes. Numerous studies have examined RNA features using whole transcriptome sequencing (RNA-seq) approaches. RNA-seq is a powerful technique for characterizing and quantifying the transcriptome and accelerates the development of bioinformatics software. In this review, we introduce routine RNA-seq workflow together with related software, focusing particularly on transcriptome reconstruction and expression quantification. PMID:26865842

  10. 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

  11. Transcriptome Analysis and Comparison of Marmota monax and Marmota himalayana.

    PubMed

    Liu, Yanan; Wang, Baoju; Wang, Lu; Vikash, Vikash; Wang, Qin; Roggendorf, Michael; Lu, Mengji; Yang, Dongliang; Liu, Jia

    2016-01-01

    The Eastern woodchuck (Marmota monax) is a classical animal model for studying hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) in humans. Recently, we found that Marmota himalayana, an Asian animal species closely related to Marmota monax, is susceptible to woodchuck hepatitis virus (WHV) infection and can be used as a new mammalian model for HBV infection. However, the lack of genomic sequence information of both Marmota models strongly limited their application breadth and depth. To address this major obstacle of the Marmota models, we utilized Illumina RNA-Seq technology to sequence the cDNA libraries of liver and spleen samples of two Marmota monax and four Marmota himalayana. In total, over 13 billion nucleotide bases were sequenced and approximately 1.5 billion clean reads were obtained. Following assembly, 106,496 consensus sequences of Marmota monax and 78,483 consensus sequences of Marmota himalayana were detected. For functional annotation, in total 73,603 Unigenes of Marmota monax and 78,483 Unigenes of Marmota himalayana were identified using different databases (NR, NT, Swiss-Prot, KEGG, COG, GO). The Unigenes were aligned by blastx to protein databases to decide the coding DNA sequences (CDS) and in total 41,247 CDS of Marmota monax and 34,033 CDS of Marmota himalayana were predicted. The single nucleotide polymorphisms (SNPs) and the simple sequence repeats (SSRs) were also analyzed for all Unigenes obtained. Moreover, a large-scale transcriptome comparison was performed and revealed a high similarity in transcriptome sequences between the two marmota species. Our study provides an extensive amount of novel sequence information for Marmota monax and Marmota himalayana. This information may serve as a valuable genomics resource for further molecular, developmental and comparative evolutionary studies, as well as for the identification and characterization of functional genes that are involved in WHV infection and HCC development

  12. Transcriptome Analysis and Comparison of Marmota monax and Marmota himalayana

    PubMed Central

    Wang, Lu; Vikash, Vikash; Wang, Qin; Roggendorf, Michael; Lu, Mengji; Yang, Dongliang; Liu, Jia

    2016-01-01

    The Eastern woodchuck (Marmota monax) is a classical animal model for studying hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC) in humans. Recently, we found that Marmota himalayana, an Asian animal species closely related to Marmota monax, is susceptible to woodchuck hepatitis virus (WHV) infection and can be used as a new mammalian model for HBV infection. However, the lack of genomic sequence information of both Marmota models strongly limited their application breadth and depth. To address this major obstacle of the Marmota models, we utilized Illumina RNA-Seq technology to sequence the cDNA libraries of liver and spleen samples of two Marmota monax and four Marmota himalayana. In total, over 13 billion nucleotide bases were sequenced and approximately 1.5 billion clean reads were obtained. Following assembly, 106,496 consensus sequences of Marmota monax and 78,483 consensus sequences of Marmota himalayana were detected. For functional annotation, in total 73,603 Unigenes of Marmota monax and 78,483 Unigenes of Marmota himalayana were identified using different databases (NR, NT, Swiss-Prot, KEGG, COG, GO). The Unigenes were aligned by blastx to protein databases to decide the coding DNA sequences (CDS) and in total 41,247 CDS of Marmota monax and 34,033 CDS of Marmota himalayana were predicted. The single nucleotide polymorphisms (SNPs) and the simple sequence repeats (SSRs) were also analyzed for all Unigenes obtained. Moreover, a large-scale transcriptome comparison was performed and revealed a high similarity in transcriptome sequences between the two marmota species. Our study provides an extensive amount of novel sequence information for Marmota monax and Marmota himalayana. This information may serve as a valuable genomics resource for further molecular, developmental and comparative evolutionary studies, as well as for the identification and characterization of functional genes that are involved in WHV infection and HCC development

  13. Characterization of citrate utilization in Corynebacterium glutamicum by transcriptome and proteome analysis.

    PubMed

    Polen, Tino; Schluesener, Daniela; Poetsch, Ansgar; Bott, Michael; Wendisch, Volker F

    2007-08-01

    Corynebacterium glutamicum grows aerobically on a variety of carbohydrates and organic acids as single or combined sources of carbon and energy. To characterize the citrate utilization in C. glutamicum on a genomewide scale, a comparative analysis was carried out by combining transcriptome and proteome analysis. In cells grown on citrate, transcriptome analysis revealed highest expression changes for two different citrate-uptake systems encoded by citM and tctCBA, whereas genes encoding uptake systems for the glucose- (ptsG), sucrose- (ptsS) and fructose- (ptsF) specific PTS components and permeases for gluconate (gntP) and glutamate (gluC) displayed decreased mRNA levels in citrate-grown cells. This pattern was also observed when cells grown in Luria-Bertani (LB) medium plus citrate were compared with cells grown in LB medium, indicating some kind of catabolite repression. Genes encoding enzymes of the tricarboxylic acid cycle (aconitase, succinyl-CoA synthetase, succinate dehydrogenase and fumarase), malic enzyme, PEP carboxykinase, gluconeogenic glyceraldehyde-3-phosphate dehydrogenase and ATP synthase displayed increased expression in cells grown on citrate. Accordingly, proteome analysis revealed elevated protein levels of these enzymes and showed a good correlation with the mRNA levels. In conclusion, this study revealed the citrate stimulon in C. glutamicum and the regulated central metabolic genes when grown on citrate.

  14. Comprehensive analysis of circadian periodic pattern in plant transcriptome

    PubMed Central

    Ptitsyn, Andrey

    2008-01-01

    Background Circadian rhythm is a crucial factor in orchestration of plant physiology, keeping it in synchrony with the daylight cycle. Previous studies have reported that up to 16% of plant transcriptome are circadially expressed. Results Our studies of mammalian gene expression revealed circadian baseline oscillation in nearly 100% of genes. Here we present a comprehensive analysis of periodicity in two independent data sets. Application of the advanced algorithms and analytic approached already tested on animal data reveals oscillation in almost every gene of Arabidopsis thaliana. Conclusion This study indicates an even more pervasive role of oscillation in molecular physiology of plants than previously believed. Earlier studies have dramatically underestimated the prevalence of circadian oscillation in plant gene expression. PMID:18793463

  15. 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

  16. Transcriptome analysis of the uniparous and multiparous goats ovaries.

    PubMed

    Wang, L J; Sun, X W; Guo, F Y; Zhao, Y J; Zhang, J H; Zhao, Z Q

    2016-12-01

    Transcriptome analysis of Inner Mongolia Cashmere goat and Dazu black goat generated 38,772,947 and 38,771,668 clean pair end reads, respectively, which were assembled into 72,422 and 80,069 unigenes by Trinity, respectively. For Inner Mongolia Cashmere goat, 26,051 and 10,100 unigenes were assigned to gene ontology (GO) categories and clusters of orthologous groups, respectively. A total of 32,772 unigenes can comment to SWISS-Prot database, and the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG) mapped 24,420 unigenes. While annotating the unigenes about Dazu black goats, we found 29,444(45.42%), 11,271 (38.28%), 36,910(56.94%) and 27,766 (42.83%) unigenes were assigned to GO database, COG database, SWISS-Prot database and KEGG database, respectively. In addition, we performed the bioinformatics analysis of gene expression profiling aimed at the ovarian transcriptome difference between Inner Mongolia Cashmere goat and Dazu black goat. We obtained a sequencing depth of over 5.5 million and 5.8 million tags. There were 1,133 DEGs between two species, of which 632 genes upregulated in the Dazu black goat and 501 genes downregulated compared with which in Inner Mongolia Cashmere goat. By annotating the 1,133 DEGs into KEGG database, we found 525 DEGs. And there were 68 DEGs annotated in metabolic pathways, 31 DEGs annotated in ribosome, 28 DEGs annotated in focal adhesion, 27 DEGs annotated in phagosome, 26 DEGs annotated in pathways in cancer, 25 DEGs annotated in ECM-receptor interaction, 23 DEGs annotated in protein digestion and absorption, 20 DEGs annotated in oxidative phosphorylation, 17 DEGs annotated in lysosome, and 16 DEGs annotated in cell adhesion molecules. © 2016 Blackwell Verlag GmbH.

  17. A comprehensive transcriptomic analysis of infant and adult mouse ovary.

    PubMed

    Pan, Linlin; Gong, Wei; Zhou, Yuanyuan; Li, Xiaonuan; Yu, Jun; Hu, Songnian

    2014-10-01

    Ovary development is a complex process involving numerous genes. A well-developed ovary is essential for females to keep fertility and reproduce offspring. In order to gain a better insight into the molecular mechanisms related to the process of mammalian ovary development, we performed a comparative transcriptomic analysis on ovaries isolated from infant and adult mice by using next-generation sequencing technology (SOLiD). We identified 15,454 and 16,646 transcriptionally active genes at the infant and adult stage, respectively. Among these genes, we also identified 7021 differentially expressed genes. Our analysis suggests that, in general, the adult ovary has a higher level of transcriptomic activity. However, it appears that genes related to primordial follicle development, such as those encoding Figla and Nobox, are more active in the infant ovary, whereas expression of genes vital for follicle development, such as Gdf9, Bmp4 and Bmp15, is upregulated in the adult. These data suggest a dynamic shift in gene expression during ovary development and it is apparent that these changes function to facilitate follicle maturation, when additional functional gene studies are considered. Furthermore, our investigation has also revealed several important functional pathways, such as apoptosis, MAPK and steroid biosynthesis, that appear to be much more active in the adult ovary compared to those of the infant. These findings will provide a solid foundation for future studies on ovary development in mice and other mammals and help to expand our understanding of the complex molecular and cellular events that occur during postnatal ovary development.

  18. Transcriptome Analysis of the Octopus vulgaris Central Nervous System

    PubMed Central

    Zhang, Xiang; Mao, Yong; Huang, Zixia; Qu, Meng; Chen, Jun; Ding, Shaoxiong; Hong, Jingni; Sun, Tiantian

    2012-01-01

    Background Cephalopoda are a class of Mollusca species found in all the world's oceans. They are an important model organism in neurobiology. Unfortunately, the lack of neuronal molecular sequences, such as ESTs, transcriptomic or genomic information, has limited the development of molecular neurobiology research in this unique model organism. Results With high-throughput Illumina Solexa sequencing technology, we have generated 59,859 high quality sequences from 12,918,391 paired-end reads. Using BLASTx/BLASTn, 12,227 contigs have blast hits in the Swissprot, NR protein database and NT nucleotide database with E-value cutoff 1e−5. The comparison between the Octopus vulgaris central nervous system (CNS) library and the Aplysia californica/Lymnaea stagnalis CNS ESTs library yielded 5.93%/13.45% of O. vulgaris sequences with significant matches (1e−5) using BLASTn/tBLASTx. Meanwhile the hit percentage of the recently published Schistocerca gregaria, Tilapia or Hirudo medicinalis CNS library to the O. vulgaris CNS library is 21.03%–46.19%. We constructed the Phylogenetic tree using two genes related to CNS function, Synaptotagmin-7 and Synaptophysin. Lastly, we demonstrated that O. vulgaris may have a vertebrate-like Blood-Brain Barrier based on bioinformatic analysis. Conclusion This study provides a mass of molecular information that will contribute to further molecular biology research on O. vulgaris. In our presentation of the first CNS transcriptome analysis of O. vulgaris, we hope to accelerate the study of functional molecular neurobiology and comparative evolutionary biology. PMID:22768275

  19. 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

  20. Transcriptome analysis of a microbial coculture in which the cell populations are separated by a membrane.

    PubMed

    Hosoda, Kazufumi; Ono, Naoaki; Suzuki, Shingo; Yomo, Tetsuya

    2014-01-01

    The microbial coculture of multiple cell populations is used to study community evolution and for bioengineering applications. The cells in coculture undergo dynamic changes because of cell-cell and cell-environment interactions. Transcriptome analysis allows us to study the molecular basis of these changes in cell physiology. For transcriptome analysis, it is essential that the cell populations in the coculture are harvested separately. Here, we describe a method for transcriptome analysis of a microbial coculture in which two different cell populations are separated by a porous membrane.

  1. Analysis of Transcriptomic Dose Response Data in the ...

    EPA Pesticide Factsheets

    Slide presentation at the HESI-HEALTH Canada-McGill Workshop on Transcriptomic Dose Response Data in the Context of Chemical Risk Assessment Slide presentation at the HESI-HEALTH Canada-McGill Workshop on Transcriptomic Dose Response Data in the Context of Chemical Risk Assessment

  2. Transcriptome analysis of genes involved in defence response in Polyporus umbellatus with Armillaria mellea infection

    PubMed Central

    Liu, Meng-Meng; Xing, Yong-Mei; Zhang, Da-Wei; Guo, Shun-Xing

    2015-01-01

    Polyporus umbellatus, a species symbiotic with Armillaria mellea and it also exhibits substantial defence response to Armillaria mellea infection. There are no genomics resources databases for understanding the molecular mechanism underlying the infection stress of P. umbellatus. Therefore, we performed a large-scale transcriptome sequencing of this fungus with A. mellea infection using Illumina sequencing technology. The assembly of the clean reads resulted in 120,576 transcripts, including 38,444 unigenes. Additionally, we performed a gene expression profiling analysis upon infection treatment. The results indicated significant differences in the gene expression profiles between the control and the infection group. In total, 10933 genes were identified between the two groups. Based on the differentially expressed genes, a Gene Ontology annotation analysis showed many defence-relevant categories. Meanwhile, the Kyoto Encyclopedia of Genes and Genomes pathway analysis uncovered some important pathways. Furthermore, the expression patterns of 13 putative genes that are involved in defence response resulting from quantitative real-time PCR were consistent with their transcript abundance changes as identified by RNA-seq. The sequenced genes covered a considerable proportion of the P. umbellatus transcriptome, and the expression results may be useful to strengthen the knowledge on the defence response of this fungus defend against Armillaria mellea invasion. PMID:26526032

  3. Combined snake venomics and venom gland transcriptomic analysis of the ocellated carpet viper, Echis ocellatus.

    PubMed

    Wagstaff, Simon C; Sanz, Libia; Juárez, Paula; Harrison, Robert A; Calvete, Juan J

    2009-01-30

    Snakebite in Africa causes thousands of deaths annually and considerable permanent physical disability. The saw-scaled viper, Echis ocellatus, represents the single most medically important snake species in West Africa. To provide a detailed compositional analysis of the venom of E. ocellatus for designing novel toxin-specific immunotherapy and to delineate sequence structure-function relationships of individual toxins, we characterised the venom proteome and the venom gland transcriptome. Whole E. ocellatus venom was fractionated by reverse-phase HPLC, followed by analysis of each chromatographic fraction using a combination of SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and CID-MS/MS of tryptic peptides. This analysis identified around 35 distinct proteins of molecular masses in the range of 5.5-110 kDa belonging to 8 different toxin families (disintegrin, DC-fragment, phospholipase A(2), cysteine-rich secretory protein, serine proteinase, C-type lectin, l-amino acid oxidase, and Zn(2+)-dependent metalloprotease). Comparison of the toxin composition of E. ocellatus venom determined using a proteomic approach, with the predicted proteome derived from assembly of 1000 EST sequences from a E. ocellatus venom gland cDNA library, shows some differences. Most notably, peptides derived from 26% of the venom proteins could not be ascribed an exact match in the transcriptome. Similarly, 64 (67%) out of the 95 putative toxin clusters reported in the transcriptome did not match to peptides detected in the venom proteome. These data suggest that the final composition of venom is influenced by transcriptional and post-translational mechanisms that may be more complex than previously appreciated. This, in turn, emphasises the value of combining proteomic and transcriptomic approaches to acquire a more complete understanding of the precise composition of snake venom, than would be gleaned from using one analysis alone. From a clinical perspective, the large

  4. De novo transcriptome assembly analysis of weed Apera spica-venti from seven tissues and growth stages.

    PubMed

    Babineau, Marielle; Mahmood, Khalid; Mathiassen, Solvejg K; Kudsk, Per; Kristensen, Michael

    2017-02-06

    Loose silky bentgrass (Apera spica-venti) is an important weed in Europe with a recent increase in herbicide resistance cases. The lack of genetic information about this noxious weed limits its biological understanding such as growth, reproduction, genetic variation, molecular ecology and metabolic herbicide resistance. This study produced a reference transcriptome for A. spica-venti from different tissues (leaf, root, stem) and various growth stages (seed at phenological stages 05, 07, 08, 09). The de novo assembly was performed on individual and combined dataset followed by functional annotations. Individual transcripts and gene families involved in metabolic based herbicide resistance were identified. Eight separate transcriptome assemblies were performed and compared. The combined transcriptome assembly consists of 83,349 contigs with an N50 and average contig length of 762 and 658 bp, respectively. This dataset contains 74,724 transcripts consisting of total 54,846,111 bp. Among them 94% had a homologue to UniProtKB, 73% retrieved a GO mapping, and 50% were functionally annotated. Compared with other grass species, A. spica-venti has 26% proteins in common to Brachypodium distachyon, and 41% to Lolium spp. Glycosyltransferases had the highest number of transcripts in each tissue followed by the cytochrome P450s. The GSTF1 and CYP89A2 transcripts were recovered from the majority of tissues and aligned at a maximum of 66 and 30% to proven herbicide resistant allele from Alopecurus myosuroides and Lolium rigidum, respectively. De novo transcriptome assembly enabled the generation of the first reference transcriptome of A. spica-venti. This can serve as stepping stone for understanding the metabolic herbicide resistance as well as the general biology of this problematic weed. Furthermore, this large-scale sequence data is a valuable scientific resource for comparative transcriptome analysis for Poaceae grasses.

  5. Transcriptome analysis of zebrafish embryos exposed to deltamethrin.

    PubMed

    Chueh, Tsung-Cheng; Hsu, Li-Sung; Kao, Chin-Ming; Hsu, Tung-Wei; Liao, Hung-Yu; Wang, Kuan-Yi; Chen, Ssu Ching

    2016-10-27

    Deltamethrin (DTM), a type II pyrethroid, is one of the most commonly used insecticides. The increased use of pyrethroid leads to potential adverse effects, particularly in sensitive populations such as children and pregnant women. None of the related studies was focused on the transcriptome responses in zebrafish embryos after treatment with DTM; therefore, RNA-seq, a high-throughput method, was performed to analyze the global expression of differential expressed genes (DEGs) in zebrafish embryos treated with DTM (40 and 80 μg/L) from fertilization to 48 h postfertilization (hpf) as compared with that in the control group (without DTM treatment). Two cDNA libraries were generated from treated embryos and one cDNA library from nontreated embryos, respectively. Over 92% of reads mapped to the reference in these three libraries. It was observed that many differential genes were expressed in comparison with embryos before and after DTM. The 20 most differentially expressed upregulated or downregulated genes were majorly involved in the signaling transduction. Validation of selected nine genes expression using qRT-PCR confirmed RNA-seq results. The transcriptome sequences were further subjected to gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, showing G-protein-coupled receptor signaling pathway and neuroactive ligand-receptor interaction, respectively, were most enriched. The data from this study contributed to a better understanding of the potential consequences of fish exposed to DTM, to an evaluation of the potential threat of DTM to fish populations in aquatic environments. © 2016 Wiley Periodicals, Inc. Environ Toxicol, 2016.

  6. Exploiting Gene Families for Phylogenomic Analysis of Myzostomid Transcriptome Data

    PubMed Central

    Hartmann, Stefanie; Helm, Conrad; Nickel, Birgit; Meyer, Matthias; Struck, Torsten H.; Tiedemann, Ralph; Selbig, Joachim; Bleidorn, Christoph

    2012-01-01

    Background In trying to understand the evolutionary relationships of organisms, the current flood of sequence data offers great opportunities, but also reveals new challenges with regard to data quality, the selection of data for subsequent analysis, and the automation of steps that were once done manually for single-gene analyses. Even though genome or transcriptome data is available for representatives of most bilaterian phyla, some enigmatic taxa still have an uncertain position in the animal tree of life. This is especially true for myzostomids, a group of symbiotic (or parasitic) protostomes that are either placed with annelids or flatworms. Methodology Based on similarity criteria, Illumina-based transcriptome sequences of one myzostomid were compared to protein sequences of one additional myzostomid and 29 reference metazoa and clustered into gene families. These families were then used to investigate the phylogenetic position of Myzostomida using different approaches: Alignments of 989 sequence families were concatenated, and the resulting superalignment was analyzed under a Maximum Likelihood criterion. We also used all 1,878 gene trees with at least one myzostomid sequence for a supertree approach: the individual gene trees were computed and then reconciled into a species tree using gene tree parsimony. Conclusions Superalignments require strictly orthologous genes, and both the gene selection and the widely varying amount of data available for different taxa in our dataset may cause anomalous placements and low bootstrap support. In contrast, gene tree parsimony is designed to accommodate multilocus gene families and therefore allows a much more comprehensive data set to be analyzed. Results of this supertree approach showed a well-resolved phylogeny, in which myzostomids were part of the annelid radiation, and major bilaterian taxa were found to be monophyletic. PMID:22276131

  7. De novo sequencing, assembly and analysis of eight different transcriptomes from the Malayan pangolin

    PubMed Central

    Mohamed Yusoff, Aini; Tan, Tze King; Hari, Ranjeev; Koepfli, Klaus-Peter; Wee, Wei Yee; Antunes, Agostinho; Sitam, Frankie Thomas; Rovie-Ryan, Jeffrine Japning; Karuppannan, Kayal Vizi; Wong, Guat Jah; Lipovich, Leonard; Warren, Wesley C.; O’Brien, Stephen J.; Choo, Siew Woh

    2016-01-01

    Pangolins are scale-covered mammals, containing eight endangered species. Maintaining pangolins in captivity is a significant challenge, in part because little is known about their genetics. Here we provide the first large-scale sequencing of the critically endangered Manis javanica transcriptomes from eight different organs using Illumina HiSeq technology, yielding ~75 Giga bases and 89,754 unigenes. We found some unigenes involved in the insect hormone biosynthesis pathway and also 747 lipids metabolism-related unigenes that may be insightful to understand the lipid metabolism system in pangolins. Comparative analysis between M. javanica and other mammals revealed many pangolin-specific genes significantly over-represented in stress-related processes, cell proliferation and external stimulus, probably reflecting the traits and adaptations of the analyzed pregnant female M. javanica. Our study provides an invaluable resource for future functional works that may be highly relevant for the conservation of pangolins. PMID:27618997

  8. De novo sequencing, assembly and analysis of eight different transcriptomes from the Malayan pangolin.

    PubMed

    Mohamed Yusoff, Aini; Tan, Tze King; Hari, Ranjeev; Koepfli, Klaus-Peter; Wee, Wei Yee; Antunes, Agostinho; Sitam, Frankie Thomas; Rovie-Ryan, Jeffrine Japning; Karuppannan, Kayal Vizi; Wong, Guat Jah; Lipovich, Leonard; Warren, Wesley C; O'Brien, Stephen J; Choo, Siew Woh

    2016-09-13

    Pangolins are scale-covered mammals, containing eight endangered species. Maintaining pangolins in captivity is a significant challenge, in part because little is known about their genetics. Here we provide the first large-scale sequencing of the critically endangered Manis javanica transcriptomes from eight different organs using Illumina HiSeq technology, yielding ~75 Giga bases and 89,754 unigenes. We found some unigenes involved in the insect hormone biosynthesis pathway and also 747 lipids metabolism-related unigenes that may be insightful to understand the lipid metabolism system in pangolins. Comparative analysis between M. javanica and other mammals revealed many pangolin-specific genes significantly over-represented in stress-related processes, cell proliferation and external stimulus, probably reflecting the traits and adaptations of the analyzed pregnant female M. javanica. Our study provides an invaluable resource for future functional works that may be highly relevant for the conservation of pangolins.

  9. Sugarcane Giant Borer Transcriptome Analysis and Identification of Genes Related to Digestion

    PubMed Central

    de Assis Fonseca, Fernando Campos; Firmino, Alexandre Augusto Pereira; de Macedo, Leonardo Lima Pepino; Coelho, Roberta Ramos; de Sousa Júnior, José Dijair Antonino; Silva-Junior, Orzenil Bonfim; Togawa, Roberto Coiti; Pappas, Georgios Joannis; de Góis, Luiz Avelar Brandão; da Silva, Maria Cristina Mattar; Grossi-de-Sá, Maria Fátima

    2015-01-01

    Sugarcane is a widely cultivated plant that serves primarily as a source of sugar and ethanol. Its annual yield can be significantly reduced by the action of several insect pests including the sugarcane giant borer (Telchin licus licus), a lepidopteran that presents a long life cycle and which efforts to control it using pesticides have been inefficient. Although its economical relevance, only a few DNA sequences are available for this species in the GenBank. Pyrosequencing technology was used to investigate the transcriptome of several developmental stages of the insect. To maximize transcript diversity, a pool of total RNA was extracted from whole body insects and used to construct a normalized cDNA database. Sequencing produced over 650,000 reads, which were de novo assembled to generate a reference library of 23,824 contigs. After quality score and annotation, 43% of the contigs had at least one BLAST hit against the NCBI non-redundant database, and 40% showed similarities with the lepidopteran Bombyx mori. In a further analysis, we conducted a comparison with Manduca sexta midgut sequences to identify transcripts of genes involved in digestion. Of these transcripts, many presented an expansion or depletion in gene number, compared to B. mori genome. From the sugarcane giant borer (SGB) transcriptome, a number of aminopeptidase N (APN) cDNAs were characterized based on homology to those reported as Cry toxin receptors. This is the first report that provides a large-scale EST database for the species. Transcriptome analysis will certainly be useful to identify novel developmental genes, to better understand the insect’s biology and to guide the development of new strategies for insect-pest control. PMID:25706301

  10. Transcriptome Analysis of a Petal Anthocyanin Polymorphism in the Arctic Mustard, Parrya nudicaulis

    PubMed Central

    Butler, Timothy; Dick, Cynthia; Carlson, Matthew L.; Whittall, Justen B.

    2014-01-01

    Angiosperms are renown for their diversity of flower colors. Often considered adaptations to pollinators, the most common underlying pigments, anthocyanins, are also involved in plants’ stress response. Although the anthocyanin biosynthetic pathway is well characterized across many angiosperms and is composed of a few candidate genes, the consequences of blocking this pathway and producing white flowers has not been investigated at the transcriptome scale. We take a transcriptome-wide approach to compare expression differences between purple and white petal buds in the arctic mustard, Parrya nudicaulis, to determine which genes’ expression are consistently correlated with flower color. Using mRNA-Seq and de novo transcriptome assembly, we assembled an average of 722 bp per gene (49.81% coding sequence based on the A. thaliana homolog) for 12,795 genes from the petal buds of a pair of purple and white samples. Our results correlate strongly with qRT-PCR analysis of nine candidate genes in the anthocyanin biosynthetic pathway where chalcone synthase has the greatest difference in expression between color morphs (P/W = ∼7×). Among the most consistently differentially expressed genes between purple and white samples, we found 3× more genes with higher expression in white petals than in purple petals. These include four unknown genes, two drought-response genes (CDSP32, ERD5), a cold-response gene (GR-RBP2), and a pathogen defense gene (DND1). Gene ontology analysis of the top 2% of genes with greater expression in white relative to purple petals revealed enrichment in genes associated with stress responses including cold, drought and pathogen defense. Unlike the uniform downregulation of chalcone synthase that may be directly involved in the loss of petal anthocyanins, the variable expression of several genes with greater expression in white petals suggest that the physiological and ecological consequences of having white petals may be microenvironment

  11. A high quality Arabidopsis transcriptome for accurate transcript-level analysis of alternative splicing

    PubMed Central

    Zhang, Runxuan; Calixto, Cristiane P. G.; Marquez, Yamile; Venhuizen, Peter; Tzioutziou, Nikoleta A.; Guo, Wenbin; Spensley, Mark; Entizne, Juan Carlos; Lewandowska, Dominika; ten Have, Sara; Frei dit Frey, Nicolas; James, Allan B.; Nimmo, Hugh G.; Barta, Andrea

    2017-01-01

    Abstract Alternative splicing generates multiple transcript and protein isoforms from the same gene and thus is important in gene expression regulation. To date, RNA-sequencing (RNA-seq) is the standard method for quantifying changes in alternative splicing on a genome-wide scale. Understanding the current limitations of RNA-seq is crucial for reliable analysis and the lack of high quality, comprehensive transcriptomes for most species, including model organisms such as Arabidopsis, is a major constraint in accurate quantification of transcript isoforms. To address this, we designed a novel pipeline with stringent filters and assembled a comprehensive Reference Transcript Dataset for Arabidopsis (AtRTD2) containing 82,190 non-redundant transcripts from 34 212 genes. Extensive experimental validation showed that AtRTD2 and its modified version, AtRTD2-QUASI, for use in Quantification of Alternatively Spliced Isoforms, outperform other available transcriptomes in RNA-seq analysis. This strategy can be implemented in other species to build a pipeline for transcript-level expression and alternative splicing analyses. PMID:28402429

  12. Scaling in sensitivity analysis

    USGS Publications Warehouse

    Link, W.A.; Doherty, P.F.

    2002-01-01

    Population matrix models allow sets of demographic parameters to be summarized by a single value 8, the finite rate of population increase. The consequences of change in individual demographic parameters are naturally measured by the corresponding changes in 8; sensitivity analyses compare demographic parameters on the basis of these changes. These comparisons are complicated by issues of scale. Elasticity analysis attempts to deal with issues of scale by comparing the effects of proportional changes in demographic parameters, but leads to inconsistencies in evaluating demographic rates. We discuss this and other problems of scaling in sensitivity analysis, and suggest a simple criterion for choosing appropriate scales. We apply our suggestions to data for the killer whale, Orcinus orca.

  13. Sequencing and Analysis of the Mediterranean Amphioxus (Branchiostoma lanceolatum) Transcriptome

    PubMed Central

    Oulion, Silvan; Bertrand, Stephanie; Belgacem, Mohamed R.; Le Petillon, Yann; Escriva, Hector

    2012-01-01

    Background The basally divergent phylogenetic position of amphioxus (Cephalochordata), as well as its conserved morphology, development and genetics, make it the best proxy for the chordate ancestor. Particularly, studies using the amphioxus model help our understanding of vertebrate evolution and development. Thus, interest for the amphioxus model led to the characterization of both the transcriptome and complete genome sequence of the American species, Branchiostoma floridae. However, recent technical improvements allowing induction of spawning in the laboratory during the breeding season on a daily basis with the Mediterranean species Branchiostoma lanceolatum have encouraged European Evo-Devo researchers to adopt this species as a model even though no genomic or transcriptomic data have been available. To fill this need we used the pyrosequencing method to characterize the B. lanceolatum transcriptome and then compared our results with the published transcriptome of B. floridae. Results Starting with total RNA from nine different developmental stages of B. lanceolatum, a normalized cDNA library was constructed and sequenced on Roche GS FLX (Titanium mode). Around 1.4 million of reads were produced and assembled into 70,530 contigs (average length of 490 bp). Overall 37% of the assembled sequences were annotated by BlastX and their Gene Ontology terms were determined. These results were then compared to genomic and transcriptomic data of B. floridae to assess similarities and specificities of each species. Conclusion We obtained a high-quality amphioxus (B. lanceolatum) reference transcriptome using a high throughput sequencing approach. We found that 83% of the predicted genes in the B. floridae complete genome sequence are also found in the B. lanceolatum transcriptome, while only 41% were found in the B. floridae transcriptome obtained with traditional Sanger based sequencing. Therefore, given the high degree of sequence conservation between different

  14. Applying thiouracil (TU)-tagging for mouse transcriptome analysis

    PubMed Central

    Gay, Leslie; Karfilis, Kate V.; Miller, Michael R.; Doe, Chris Q.; Stankunas, Kryn

    2014-01-01

    Transcriptional profiling is a powerful approach to study mouse development, physiology, and disease models. Here, we describe a protocol for mouse thiouracil-tagging (TU-tagging), a transcriptome analysis technology that includes in vivo covalent labeling, purification, and analysis of cell type-specific RNA. TU-tagging enables 1) the isolation of RNA from a given cell population of a complex tissue, avoiding transcriptional changes induced by cell isolation trauma, and 2) the identification of actively transcribed RNAs and not pre-existing transcripts. Therefore, in contrast to other cell-specific transcriptional profiling methods based on purification of tagged ribosomes or nuclei, TU-tagging provides a direct examination of transcriptional regulation. We describe how to: 1) deliver 4-thiouracil to transgenic mice to thio-label cell lineage-specific transcripts, 2) purify TU-tagged RNA and prepare libraries for Illumina sequencing, and 3) follow a straight-forward bioinformatics workflow to identify cell type-enriched or differentially expressed genes. Tissue containing TU-tagged RNA can be obtained in one day, RNA-Seq libraries generated within two days, and, following sequencing, an initial bioinformatics analysis completed in one additional day. PMID:24457332

  15. Transcriptome profiling analysis of cultivar-specific apple fruit ripening and texture attributes

    USDA-ARS?s Scientific Manuscript database

    Molecular events regulating cultivar-specific 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, transcriptome profile analysis, scanning electron microscopic examination an...

  16. Comparative transcriptome analysis during early fruit development between three seedy citrus genotypes and their seedless mutants

    USDA-ARS?s Scientific Manuscript database

    Identification of genes with differential transcript abundance (GDTA) in seedless mutants may enhance understanding of seedless citrus development. Transcriptome analysis was conducted at three time points during early fruit development (Phase 1) of three seedy citrus genotypes: Fallglo [Bower citru...

  17. De Novo Transcriptome Analysis of Cucumis melo L. var. makuwa

    PubMed Central

    Kim, Hyun A; Shin, Ah-Young; Lee, Min-Seon; Lee, Hee-Jeong; Lee, Heung-Ryul; Ahn, Jongmoon; Nahm, Seokhyeon; Jo, Sung-Hwan; Park, Jeong Mee; Kwon, Suk-Yoon

    2016-01-01

    Oriental melon (Cucumis melo L. var. makuwa) is one of six subspecies of melon and is cultivated widely in East Asia, including China, Japan, and Korea. Although oriental melon is economically valuable in Asia and is genetically distinct from other subspecies, few reports of genome-scale research on oriental melon have been published. We generated 30.5 and 36.8 Gb of raw RNA sequence data from the female and male flowers, leaves, roots, and fruit of two oriental melon varieties, Korean landrace (KM) and Breeding line of NongWoo Bio Co. (NW), respectively. From the raw reads, 64,998 transcripts from KM and 100,234 transcripts from NW were de novo assembled. The assembled transcripts were used to identify molecular markers (e.g., single-nucleotide polymorphisms and simple sequence repeats), detect tissue-specific expressed genes, and construct a genetic linkage map. In total, 234 single-nucleotide polymorphisms and 25 simple sequence repeats were screened from 7,871 and 8,052 candidates, respectively, between the KM and NW varieties and used for construction of a genetic map with 94 F2 population specimens. The genetic linkage map consisted of 12 linkage groups, and 248 markers were assigned. These transcriptome and molecular marker data provide information useful for molecular breeding of oriental melon and further comparative studies of the Cucurbitaceae family. PMID:26743902

  18. De Novo Transcriptome Analysis of Cucumis melo L. var. makuwa.

    PubMed

    Kim, Hyun A; Shin, Ah-Young; Lee, Min-Seon; Lee, Hee-Jeong; Lee, Heung-Ryul; Ahn, Jongmoon; Nahm, Seokhyeon; Jo, Sung-Hwan; Park, Jeong Mee; Kwon, Suk-Yoon

    2016-02-01

    Oriental melon (Cucumis melo L. var. makuwa) is one of six subspecies of melon and is cultivated widely in East Asia, including China, Japan, and Korea. Although oriental melon is economically valuable in Asia and is genetically distinct from other subspecies, few reports of genome-scale research on oriental melon have been published. We generated 30.5 and 36.8 Gb of raw RNA sequence data from the female and male flowers, leaves, roots, and fruit of two oriental melon varieties, Korean landrace (KM) and Breeding line of NongWoo Bio Co. (NW), respectively. From the raw reads, 64,998 transcripts from KM and 100,234 transcripts from NW were de novo assembled. The assembled transcripts were used to identify molecular markers (e.g., single-nucleotide polymorphisms and simple sequence repeats), detect tissue-specific expressed genes, and construct a genetic linkage map. In total, 234 single-nucleotide polymorphisms and 25 simple sequence repeats were screened from 7,871 and 8,052 candidates, respectively, between the KM and NW varieties and used for construction of a genetic map with 94 F2 population specimens. The genetic linkage map consisted of 12 linkage groups, and 248 markers were assigned. These transcriptome and molecular marker data provide information useful for molecular breeding of oriental melon and further comparative studies of the Cucurbitaceae family.

  19. Whole transcriptome analysis of Penicillium digitatum strains treatmented with prochloraz reveals their drug-resistant mechanisms.

    PubMed

    Liu, Jing; Wang, Shengqiang; Qin, Tingting; Li, Na; Niu, Yuhui; Li, Dandan; Yuan, Yongze; Geng, Hui; Xiong, Li; Liu, Deli

    2015-10-24

    Penicillium digitatum is one of the most destructive postharvest pathogen of citrus fruits, causing fruit decay and economic loss. The emergence of fungicide-resistant strains made the control of P. digitatum more difficult. While the genome of P. digitatum is available, there has been few reports about its resistant mechanism from the transcriptome perspective and there has been no large-scale functional annotation of the genome using expressed genes derived from transcriptomes. Total RNA of P. digitatum strain HS-F6 (prochloraz-resistant strain) and HS-E3 (prochloraz-susceptible strain) before and after prochloraz-treatment were extracted and sequenced on an Illumina Hiseq 2000 platform. The transcriptome data of four samples were compared and analyzed using differential expression analysis, novel transcripts prediction and alternative splicing analysis, SNP analysis and quantitative real-time PCR. We present a large scale analysis about the transcriptome data of P. digitatum. The whole RNA was extracted from a prochloraz-resistant strain (HS-F6) and a prochloraz-susceptible strain (HS-E3) before and after prochloraz-treatment and sequenced by Illumina technology. A total of more than 100 million reads were generated and de novo assembled into 9760 transcripts that contained annotated genes after quality control and sequence assembling. 6625 single nucleotide variations (SNVs) were identified from the sequences aligned against the reference genome. Gene expression profiling analysis was performed upon prochloraz treatment in HS-F6 and HS-E3, and differential expression analysis was used to identify genes related to prochloraz-response and drug-resistance: there are 224 differentially expressed genes in HS-E3 and 1100 differentially expressed genes in HS-F6 after prochloraz-treatment. Moreover, gene expression profile in prochloraz-resistant strain HS-F6 is quite different from that in HS-E3 before prochloraz-treatment, 1520 differential expression genes were

  20. Transcriptome Analysis of Stem and Globally Comparison with Other Tissues in Brassica napus

    PubMed Central

    Miao, Liyun; Zhang, Libin; Raboanatahiry, Nadia; Lu, Guangyuan; Zhang, Xuekun; Xiang, Jun; Gan, Jianping; Fu, Chunhua; Li, Maoteng

    2016-01-01

    Brassica napus is one of the most important oilseed crops in the world. However, there is currently no enough stem transcriptome information and comparative transcriptome analysis of different tissues, which impedes further functional genomics research on B. napus. In this study, the stem transcriptome of B. napus was characterized by RNA-seq technology. Approximately 13.4 Gb high-quality clean reads with an average length of 100 bp were generated and used for comparative transcriptome analysis with the existing transcriptome sequencing data of roots, leaves, flower buds, and immature embryos of B. napus. All the transcripts were annotated against GO and KEGG databases. The common genes in five tissues, differentially expressed genes (DEGs) of the common genes between stems and other tissues, and tissue-specific genes were detected, and the main biochemical activities and pathways implying the common genes, DEGs and tissue-specific genes were investigated. Accordingly, the common transcription factors (TFs) in the five tissues and tissue-specific TFs were identified, and a TFs-based regulation network between TFs and the target genes involved in ‘Phenylpropanoid biosynthesis’ pathway were constructed to show several important TFs and key nodes in the regulation process. Collectively, this study not only provided an available stem transcriptome resource in B. napus, but also revealed valuable comparative transcriptome information of five tissues of B. napus for future investigation on specific processes, functions and pathways. PMID:27708656

  1. Full Transcriptome Analysis of Early Dorsoventral Patterning in Zebrafish

    PubMed Central

    Horváth, Balázs; Molnár, János; Nagy, István; Tóth, Gábor; Wilson, Stephen W.; Varga, Máté

    2013-01-01

    Understanding the molecular interactions that lead to the establishment of the major body axes during embryogenesis is one of the main goals of developmental biology. Although the past two decades have revolutionized our knowledge about the genetic basis of these patterning processes, the list of genes involved in axis formation is unlikely to be complete. In order to identify new genes involved in the establishment of the dorsoventral (DV) axis during early stages of zebrafish embryonic development, we employed next generation sequencing for full transcriptome analysis of normal embryos and embryos lacking overt DV pattern. A combination of different statistical approaches yielded 41 differentially expressed candidate genes and we confirmed by in situ hybridization the early dorsal expression of 32 genes that are transcribed shortly after the onset of zygotic transcription. Although promoter analysis of the validated genes suggests no general enrichment for the binding sites of early acting transcription factors, most of these genes carry “bivalent” epigenetic histone modifications at the time when zygotic transcription is initiated, suggesting a “poised” transcriptional status. Our results reveal some new candidates of the dorsal gene regulatory network and suggest that a plurality of the earliest upregulated genes on the dorsal side have a role in the modulation of the canonical Wnt pathway. PMID:23922899

  2. Analysis of SOX2-Regulated Transcriptome in Glioma Stem Cells

    PubMed Central

    Acanda de la Rocha, Arlet M.; López-Bertoni, Hernando; Guruceaga, Elizabeth; González-Huarriz, Marisol; Martínez-Vélez, Naiara; Xipell, Enric; Fueyo, Juan; Gomez-Manzano, Candelaria

    2016-01-01

    Introduction Glioblastoma is the most malignant brain tumor in adults and is associated with poor survival despite multimodal treatments. Glioma stem-like cells (GSCs) are cells functionally defined by their self-renewal potential and the ability to reconstitute the original tumor upon orthotopic implantation. They have been postulated to be the culprit of glioma chemo- and radio-resistance ultimately leading to relapse. Understanding the molecular circuits governing the GSC compartment is essential. SOX2, a critical transcription regulator of embryonic and neural stem cell function, is deregulated in GSCs however; the precise molecular pathways regulated by this gene in GSCs remain poorly understood. Results We performed a genome-wide analysis of SOX2-regulated transcripts in GSCs, using a microarray. We identified a total of 2048 differentially expressed coding transcripts and 261 non-coding transcripts. Cell adhesion and cell-cell signaling are among the most enriched terms using Gene Ontology (GO) classification. The pathways altered after SOX2 down-modulation includes multiple cellular processes such as amino-acid metabolism and intercellular signaling cascades. We also defined and classified the set of non-coding transcripts differentially expressed regulated by SOX2 in GSCs, and validated two of them. Conclusions We present a comprehensive analysis of the transcriptome controlled by SOX2 in GSCs, gaining insights in the understanding of the potential roles of SOX2 in glioblastoma. PMID:27669421

  3. Transcriptomic Analysis of the Porcine Endometrium during Embryo Implantation

    PubMed Central

    Lin, Haichao; Wang, Huaizhong; Wang, Yanping; Liu, Chang; Wang, Cheng; Guo, Jianfeng

    2015-01-01

    In pigs, successful embryo implantation is an important guarantee for producing litter size, and early embryonic loss occurring on day 12–30 of gestation critically affects the potential litter size. The implantation process is regulated by the expression of numerous genes, so comprehensive analysis of the endometrium is necessary. In this study, RNA sequencing (RNA-Seq) technology is used to analyze endometrial tissues during early pregnancy. We investigated the changes of gene expression between three stages (day 12, 18, and 25) by multiple comparisons. There were 1557, 8951, and 2345 differentially expressed genes (DEGs) revealed between the different periods of implantation. We selected several genes for validation by the use of quantitative real-time RT-PCR. Bioinformatic analysis of differentially expressed genes in the endometrium revealed a number of biological processes and pathways potentially involved in embryo implantation in the pig, most noticeably cell proliferation, regulation of immune response, interaction of cytokine-cytokine receptors, and cell adhesion. These results showed that specific gene expression patterns reflect the different functions of the endometrium in three stages (maternal recognition, conceptus attachment, and embryo implantation). This study identified comprehensive transcriptomic profile in the porcine endometrium and thus could be a foundation for targeted studies of genes and pathways potentially involved in abnormal endometrial receptivity and embryo loss in early pregnancy. PMID:26703736

  4. Function-informed transcriptome analysis of Drosophila renal tubule

    PubMed Central

    Wang, Jing; Kean, Laura; Yang, Jingli; Allan, Adrian K; Davies, Shireen A; Herzyk, Pawel; Dow, Julian AT

    2004-01-01

    Background Comprehensive, tissue-specific, microarray analysis is a potent tool for the identification of tightly defined expression patterns that might be missed in whole-organism scans. We applied such an analysis to Drosophila melanogaster Malpighian (renal) tubule, a defined differentiated tissue. Results The transcriptome of the D. melanogaster Malpighian tubule is highly reproducible and significantly different from that obtained from whole-organism arrays. More than 200 genes are more than 10-fold enriched and over 1,000 are significantly enriched. Of the top 200 genes, only 18 have previously been named, and only 45% have even estimates of function. In addition, 30 transcription factors, not previously implicated in tubule development, are shown to be enriched in adult tubule, and their expression patterns respect precisely the domains and cell types previously identified by enhancer trapping. Of Drosophila genes with close human disease homologs, 50 are enriched threefold or more, and eight enriched 10-fold or more, in tubule. Intriguingly, several of these diseases have human renal phenotypes, implying close conservation of renal function across 400 million years of divergent evolution. Conclusions From those genes that are identifiable, a radically new view of the function of the tubule, emphasizing solute transport rather than fluid secretion, can be obtained. The results illustrate the phenotype gap: historically, the effort expended on a model organism has tended to concentrate on a relatively small set of processes, rather than on the spread of genes in the genome. PMID:15345053

  5. Transcriptome Sequencing and Positive Selected Genes Analysis of Bombyx mandarina

    PubMed Central

    Wu, Yuqian; Long, Renwen; Liu, Chun; Xia, Qingyou

    2015-01-01

    The wild silkworm Bombyx mandarina is widely believed to be an ancestor of the domesticated silkworm, Bombyx mori. Silkworms are often used as a model for studying the mechanism of species domestication. Here, we performed transcriptome sequencing of the wild silkworm using an Illumina HiSeq2000 platform. We produced 100,004,078 high-quality reads and assembled them into 50,773 contigs with an N50 length of 1764 bp and a mean length of 941.62 bp. A total of 33,759 unigenes were identified, with 12,805 annotated in the Nr database, 8273 in the Pfam database, and 9093 in the Swiss-Prot database. Expression profile analysis found significant differential expression of 1308 unigenes between the middle silk gland (MSG) and posterior silk gland (PSG). Three sericin genes (sericin 1, sericin 2, and sericin 3) were expressed specifically in the MSG and three fibroin genes (fibroin-H, fibroin-L, and fibroin/P25) were expressed specifically in the PSG. In addition, 32,297 Single-nucleotide polymorphisms (SNPs) and 361 insertion-deletions (INDELs) were detected. Comparison with the domesticated silkworm p50/Dazao identified 5,295 orthologous genes, among which 400 might have experienced or to be experiencing positive selection by Ka/Ks analysis. These data and analyses presented here provide insights into silkworm domestication and an invaluable resource for wild silkworm genomics research. PMID:25806526

  6. 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

  7. 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.

  8. Identification and analysis of common bean (Phaseolus vulgaris L.) transcriptomes by massively parallel pyrosequencing

    PubMed Central

    2011-01-01

    Background Common bean (Phaseolus vulgaris) is the most important food legume in the world. Although this crop is very important to both the developed and developing world as a means of dietary protein supply, resources available in common bean are limited. Global transcriptome analysis is important to better understand gene expression, genetic variation, and gene structure annotation in addition to other important features. However, the number and description of common bean sequences are very limited, which greatly inhibits genome and transcriptome research. Here we used 454 pyrosequencing to obtain a substantial transcriptome dataset for common bean. Results We obtained 1,692,972 reads with an average read length of 207 nucleotides (nt). These reads were assembled into 59,295 unigenes including 39,572 contigs and 19,723 singletons, in addition to 35,328 singletons less than 100 bp. Comparing the unigenes to common bean ESTs deposited in GenBank, we found that 53.40% or 31,664 of these unigenes had no matches to this dataset and can be considered as new common bean transcripts. Functional annotation of the unigenes carried out by Gene Ontology assignments from hits to Arabidopsis and soybean indicated coverage of a broad range of GO categories. The common bean unigenes were also compared to the bean bacterial artificial chromosome (BAC) end sequences, and a total of 21% of the unigenes (12,724) including 9,199 contigs and 3,256 singletons match to the 8,823 BAC-end sequences. In addition, a large number of simple sequence repeats (SSRs) and transcription factors were also identified in this study. Conclusions This work provides the first large scale identification of the common bean transcriptome derived by 454 pyrosequencing. This research has resulted in a 150% increase in the number of Phaseolus vulgaris ESTs. The dataset obtained through this analysis will provide a platform for functional genomics in common bean and related legumes and will aid in the

  9. Comprehensive analysis of Sichuan white geese (Anser cygnoides) transcriptome.

    PubMed

    Ding, Ning; Han, Qing; Li, Qin; Zhao, Xianzhi; Li, Jing; Su, Jian; Wang, Qigui

    2014-06-01

    High-throughput RNA sequencing was performed for comprehensively analyzing the transcriptome of geese. A total of 28,803,759 bp of raw sequence data was generated by 454 GS Flx+. After removal of adaptor sequences, 28,730,361 bp remained and 117,279 reads were obtained, with an average length of 244 bases. Simultaneously, complementary DNA samples from two different reproductive stages of goose ovarian, hypothalamus and pituitary tissue were sequenced separately using Illumina MiSeq platform. A total of 12 688 673 148 bp of raw sequence data were generated by Illumina MiSeq. After removal of adaptor sequences, 8 198 126 562 bp remained and 60 382 786 clean reads were obtained, with an average length of 135 bases. Assembly of all the reads from both 454 Flx+ and Illumina platforms formed 56,839 contigs. The sequence size ranges from 38 to 28,206 bp in size, with an average size of 2584 bp and an N50 of 4624. The assembly produced a substantial number of large contigs: 35,545 (62.5%) were longer than 1 kb, of which 8850 (15.6%) were longer than 5 kb. The sequencing depth was 85 X on average. We performed comprehensive function annotations on unigenes including protein sequence similarity, gene ontology (GO) term classification, and Kyoto Encylcopedia of Genes and Genomes (KEGG) pathway enrichment. GO analysis showed that approximately 63% of the contigs had annotation information, among the 35,953 annotated isotigs in Nr database, 24,783 (68.9%) sequences were assigned with one or more GO terms. There were 14,634 (40.7%) isotigs for biological processes, 10,557(29.3%) isotigs for cellular component, 22,607 (62.9%) isotigs for molecular function. The result of KEGG pathway mapping 8926 sequences had the pathway annotation, and took part in 477 pathways. Additionally, 10,685 simple sequence repeat (SSR) markers were identified from the assembled sequences. The most frequent repeat motifs were trinucleotides, which accounted for 53.03% of all SSRs

  10. Comprehensive Transcriptome Analysis of the Periodontopathogenic Bacterium Porphyromonas gingivalis W83

    PubMed Central

    Høvik, Hedda; Yu, Wen-Han; Olsen, Ingar

    2012-01-01

    High-density tiling microarray and RNA sequencing technologies were used to analyze the transcriptome of the periodontopathogenic bacterium Porphyromonas gingivalis. The compiled P. gingivalis transcriptome profiles were based on total RNA samples isolated from three different laboratory culturing conditions, and the strand-specific transcription profiles generated covered the entire genome, including both protein coding and noncoding regions. The transcription profiles revealed various operon structures, 5′- and 3′-end untranslated regions (UTRs), differential expression patterns, and many novel, not-yet-annotated transcripts within intergenic and antisense regions. Further transcriptome analysis identified the majority of the genes as being expressed within operons and most 5′ and 3′ ends to be protruding UTRs, of which several 3′ UTRs were extended to overlap genes carried on the opposite/antisense strand. Extensive antisense RNAs were detected opposite most insertion sequence (IS) elements. Pairwise comparative analyses were also performed among transcriptome profiles of the three culture conditions, and differentially expressed genes and metabolic pathways were identified. With the growing realization that noncoding RNAs play important biological functions, the discovery of novel RNAs and the comprehensive transcriptome profiles compiled in this study may provide a foundation to further understand the gene regulation and virulence mechanisms in P. gingivalis. The transcriptome profiles can be viewed at and downloaded from the Microbial Transcriptome Database website, http://bioinformatics.forsyth.org/mtd. PMID:22037400

  11. Sex Change in Clownfish: Molecular Insights from Transcriptome Analysis

    PubMed Central

    Casas, Laura; Saborido-Rey, Fran; Ryu, Taewoo; Michell, Craig; Ravasi, Timothy; Irigoien, Xabier

    2016-01-01

    Sequential hermaphroditism is a unique reproductive strategy among teleosts that is displayed mainly in fish species living in the coral reef environment. The reproductive biology of hermaphrodites has long been intriguing; however, very little is known about the molecular pathways underlying their sex change. Here, we provide the first de novo transcriptome analyses of a hermaphrodite teleost´s undergoing sex change in its natural environment. Our study has examined relative gene expression across multiple groups—rather than just two contrasting conditions— and has allowed us to explore the differential expression patterns throughout the whole process. Our analysis has highlighted the rapid and complex genomic response of the brain associated with sex change, which is subsequently transmitted to the gonads, identifying a large number of candidate genes, some well-known and some novel, involved in the process. The present study provides strong evidence of the importance of the sex steroidogenic machinery during sex change in clownfish, with the aromatase gene playing a central role, both in the brain and the gonad. This work constitutes the first genome-wide study in a social sex-changing species and provides insights into the genetic mechanism governing social sex change and gonadal restructuring in protandrous hermaphrodites. PMID:27748421

  12. Integrated Analysis of Transcriptome in Cancer Patient-Derived Xenografts

    PubMed Central

    Li, Hong; Zhu, Yinjie; Tang, Xiaoyan; Li, Junyi; Li, Yuanyuan; Zhong, Zhaomin; Ding, Guohui; Li, Yixue

    2015-01-01

    Patient-derived xenograft (PDX) tumor model is a powerful technology in evaluating anti-cancer drugs and facilitating personalized medicines. Multiple research centers and commercial companies have put huge efforts into building PDX mouse models. However, PDX models have not been widely available and their molecular features have not been systematically characterized. In this study, we provided a comprehensive survey of PDX transcriptome by integrating analysis of 58 patients involving 8 different tumors. The median correlation coefficient between patients and xenografts is 0.94, which is higher than that between patients and cell line panel or between patients with the same tumor. Major differential gene expressions in PDX occur in the engraftment of human tumor tissue into mice, while gene expressions are relatively stable over passages. 48 genes are frequently differentially expressed in PDX mice of multiple cancers. They are enriched in extracellular matrix and immune response, and some are reported as targets for anticancer drugs. A simulation study showed that expression change between PDX and patient tumor (6%) would result in acceptable change in drug sensitivity (3%). Our findings demonstrate that PDX mice represent the gene-expression and drug-response features of primary tumors effectively, and it is recommended to monitoring the overall expression profiles and drug target genes in clinical application. PMID:25951608

  13. Comparative transcriptome analysis of grapevine in response to copper stress

    PubMed Central

    Leng, Xiangpeng; Jia, Haifeng; Sun, Xin; Shangguan, Lingfei; Mu, Qian; Wang, Baoju; Fang, Jinggui

    2015-01-01

    Grapevine is one of the most economically important and widely cultivated fruit crop worldwide. With the industrialization and the popular application of cupric fungicides in grape industry, copper stress and copper pollution are also the factors affecting grape production and berry and wine quality. Here, 3,843 transcripts were significantly differently expressed genes in response to Cu stress by RNA-seq, which included 1,892 up-regulated and 1,951 down-regulated transcripts. During this study we found many known and novel Cu-induced and -repressed genes. Biological analysis of grape samples were indicated that exogenous Cu can influence chlorophylls metabolism and photosynthetic activities of grapevine. Most ROS detoxification systems, including antioxidant enzyme, stress-related proteins and secondary metabolites were strongly induced. Concomitantly, abscisic acid functioned as a negative regulator in Cu stress, in opposite action to ethylene, auxin, jasmonic acid, and brassinolide. This study also identified a set of Cu stress specifically activated genes coding copper transporter, P1B-type ATPase, multidrug transporters. Overall, this work was carried out to gain insights into the copper-regulated and stress-responsive mechanisms in grapevine at transcriptome level. This research can also provide some genetic information that can help us in better vinery management and breeding Cu-resistant grape cultivars. PMID:26673527

  14. Transcriptomic analysis of basidiocarp development in Ustilago maydis (DC) Cda.

    PubMed

    León-Ramírez, C G; Cabrera-Ponce, J L; Martínez-Soto, D; Sánchez-Arreguin, A; Aréchiga-Carvajal, E T; Ruiz-Herrera, J

    2017-03-07

    Previously, we demonstrated that when Ustilago maydis (DC) Cda., a phytopathogenic basidiomycete and the causal agent of corn smut, is grown in the vicinity of maize embryogenic calli in a medium supplemented with the herbicide Dicamba, it developed gastroid-like basidiocarps. To elucidate the molecular mechanisms involved in the basidiocarp development by the fungus, we proceeded to analyze the transcriptome of the process, identifying a total of 2002 and 1064 differentially expressed genes at two developmental stages, young and mature basidiocarps, respectively. Function of these genes was analyzed with the use of different databases. MIPS analysis revealed that in the stage of young basidiocarp, among the ca. two thousand differentially expressed genes, there were some previously described for basidiocarp development in other fungal species. Additional elements that operated at this stage included, among others, genes encoding the transcription factors FOXO3, MIG3, PRO1, TEC1, copper and MFS transporters, and cytochromes P450. During mature basidiocarp development, important up-regulated genes included those encoding hydrophobins, laccases, and ferric reductase (FRE/NOX). The demonstration that a mapkk mutant was unable to form basidiocarps, indicated the importance of the MAPK signaling pathway in this developmental process.

  15. Comparative transcriptome analysis of grapevine in response to copper stress.

    PubMed

    Leng, Xiangpeng; Jia, Haifeng; Sun, Xin; Shangguan, Lingfei; Mu, Qian; Wang, Baoju; Fang, Jinggui

    2015-12-17

    Grapevine is one of the most economically important and widely cultivated fruit crop worldwide. With the industrialization and the popular application of cupric fungicides in grape industry, copper stress and copper pollution are also the factors affecting grape production and berry and wine quality. Here, 3,843 transcripts were significantly differently expressed genes in response to Cu stress by RNA-seq, which included 1,892 up-regulated and 1,951 down-regulated transcripts. During this study we found many known and novel Cu-induced and -repressed genes. Biological analysis of grape samples were indicated that exogenous Cu can influence chlorophylls metabolism and photosynthetic activities of grapevine. Most ROS detoxification systems, including antioxidant enzyme, stress-related proteins and secondary metabolites were strongly induced. Concomitantly, abscisic acid functioned as a negative regulator in Cu stress, in opposite action to ethylene, auxin, jasmonic acid, and brassinolide. This study also identified a set of Cu stress specifically activated genes coding copper transporter, P1B-type ATPase, multidrug transporters. Overall, this work was carried out to gain insights into the copper-regulated and stress-responsive mechanisms in grapevine at transcriptome level. This research can also provide some genetic information that can help us in better vinery management and breeding Cu-resistant grape cultivars.

  16. Transcriptome analysis of the salivary glands of Nephotettix cincticeps (Uhler).

    PubMed

    Matsumoto, Yukiko; Suetsugu, Yoshitaka; Nakamura, Masatoshi; Hattori, Makoto

    2014-12-01

    The green rice leafhopper (GRH), Nephotettix cincticeps, is one of the most important pests of rice in temperate Asian countries. GRH, a vascular feeder, secretes watery and gelling saliva in the process of feeding on phloem and xylem sap. It is known that GRH saliva contains several bioactive proteins, including enzymes such as laccase and beta-glucosidase. In this study, we performed transcriptome analysis of salivary glands of GRH using Illumina paired-end sequencing. Of 51,788 assembled contigs, 16,017 (30.9%) showed significant similarity to known proteins in the NCBI nr database, while 34,978 (67.5%) could not be annotated by similarity search, Pfam, or gene ontology (GO). Contigs (905) with predicted signal peptides and no putative transmembrane domains are suggested to represent secreted protein coding genes. Among the 76 most highly expressed putative secretory protein contigs, 68 transcripts were found to be salivary gland-specific or at least -dominant, but not expressed in stomach or Malpighian tubules. However, 45 of the 68 transcripts were unknown proteins. These findings suggest that most of the GRH transcripts encoding secreted proteins expressed in salivary glands are species and/or tissue specific. Our results provide a fundamental list of genes involved in GRH-Poaceae host plant interactions including successful feeding and plant pathogen transmission.

  17. Transcriptome characterization and SSR discovery in large-scale loach Paramisgurnus dabryanus (Cobitidae, Cypriniformes).

    PubMed

    Li, Caijuan; Ling, Qufei; Ge, Chen; Ye, Zhuqing; Han, Xiaofei

    2015-02-25

    The large-scale loach (Paramisgurnus dabryanus, Cypriniformes) is a bottom-dwelling freshwater species of fish found mainly in eastern Asia. The natural germplasm resources of this important aquaculture species has been recently threatened due to overfishing and artificial propagation. The objective of this study is to obtain the first functional genomic resource and candidate molecular markers for future conservation and breeding research. Illumina paired-end sequencing generated over one hundred million reads that resulted in 71,887 assembled transcripts, with an average length of 1465bp. 42,093 (58.56%) protein-coding sequences were predicted; and 43,837 transcripts had significant matches to NCBI nonredundant protein (Nr) database. 29,389 and 14,419 transcripts were assigned into gene ontology (GO) categories and Eukaryotic Orthologous Groups (KOG), respectively. 22,102 (31.14%) transcripts were mapped to 302 KEGG pathways. In addition, 15,106 candidate SSR markers were identified, with 11,037 pairs of PCR primers designed. 400 primers pairs of SSR selected randomly were validated, of which 364 (91%) pairs of primers were able to produce PCR products. Further test with 41 loci and 20 large-scale loach specimens collected from the four largest lakes in China showed that 36 (87.8%) loci were polymorphic. The transcriptomic profile and SSR repertoire obtained in this study will facilitate population genetic studies and selective breeding of large-scale loach in the future. Copyright © 2015. Published by Elsevier B.V.

  18. Comprehensive comparison of Pacific Biosciences and Oxford Nanopore Technologies and their applications to transcriptome analysis.

    PubMed

    Weirather, Jason L; de Cesare, Mariateresa; Wang, Yunhao; Piazza, Paolo; Sebastiano, Vittorio; Wang, Xiu-Jie; Buck, David; Au, Kin Fai

    2017-01-01

    Background: Given the demonstrated utility of Third Generation Sequencing [Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT)] long reads in many studies, a comprehensive analysis and comparison of their data quality and applications is in high demand. Methods: Based on the transcriptome sequencing data from human embryonic stem cells, we analyzed multiple data features of PacBio and ONT, including error pattern, length, mappability and technical improvements over previous platforms. We also evaluated their application to transcriptome analyses, such as isoform identification and quantification and characterization of transcriptome complexity, by comparing the performance of size-selected PacBio, non-size-selected ONT and their corresponding Hybrid-Seq strategies (PacBio+Illumina and ONT+Illumina). Results: PacBio shows overall better data quality, while ONT provides a higher yield. As with data quality, PacBio performs marginally better than ONT in most aspects for both long reads only and Hybrid-Seq strategies in transcriptome analysis. In addition, Hybrid-Seq shows superior performance over long reads only in most transcriptome analyses. Conclusions: Both PacBio and ONT sequencing are suitable for full-length single-molecule transcriptome analysis. As this first use of ONT reads in a Hybrid-Seq analysis has shown, both PacBio and ONT can benefit from a combined Illumina strategy. The tools and analytical methods developed here provide a resource for future applications and evaluations of these rapidly-changing technologies.

  19. De novo transcriptome analysis of mulberry (Morus L.) under drought stress using RNA-seq technology.

    PubMed

    Wang, Heng; Tong, Wei; Feng, Li; Jiao, Qian; Long, Li; Fang, Rongjun; Zhao, Weiguo

    2014-01-01

    A large-scale RNA sequencing (RNA-seq) of mulberry (Morus L.) was carried out between two samples in regular and drought stress condition. In this research, de novo assembly was performed, and totally 54736 contigs were obtained from the reads, including the scaffolded regions. 1051 genes were identified that were significantly differently expressed between the two samples. As determined by Gene Ontology (GO) annotation and the Kyoto Encyclopedia of Genes and Genomes pathway mapping, 10110 GO terms and 247 pathways were assigned and then analyzed. Thousands of SSR markers produced in this study will enable genetic linkage mapping construction and gene-based association studies. Seven unique genes showing different expression level in control and drought stress groups were subsequently analyzed and identified by real-time PCR. For lack of mulberry whole genome information, transcriptome and de novo analysis from the two samples will provide important and useful information for later research and help genetic breeding of mulberry.

  20. Comparative transcriptomic analysis reveals phenol tolerance mechanism of evolved Chlorella strain.

    PubMed

    Zhou, Lin; Cheng, Dujia; Wang, Liang; Gao, Juan; Zhao, Quanyu; Wei, Wei; Sun, Yuhan

    2017-03-01

    The growth of microalgae is inhibited by high concentration phenol due to reactive oxygen species. An evolved strain tolerated to 500mg/L phenol, Chlorella sp. L5, was obtained in previous study. In this study, comparative transcriptomic analysis was performed for Chlorella sp. L5 and its original strain (Chlorella sp. L3). The tolerance mechanism of Chlorella sp. L5 for high concentration phenol was explored on genome scale. It was identified that the up-regulations of the related genes according to antioxidant enzymes (SOD, APX, CAT and GR) and carotenoids (astaxanthin, lutein and lycopene) biosynthesis had critical roles to tolerate high concentration phenol. In addition, most of genes of PS I, PS II, photosynthetic electron transport chain and starch biosynthesis were also up-regulated. It was consistent to the experimental results of total carbohydrate contents of Chlorella sp. L3 and Chlorella sp. L5 under 0mg/L and 500mg/L phenol.

  1. A genome survey and postharvest transcriptome analysis in Lentinula edodes.

    PubMed

    Sakamoto, Yuichi; Nakade, Keiko; Sato, Shiho; Yoshida, Kentaro; Miyazaki, Kazuhiro; Natsume, Satoshi; Konno, Naotake

    2017-03-17

    Lentinula edodes is a popular cultivated edible and medicinal mushroom. Lentinula edodes is susceptible to postharvest problems such as gill browning, fruiting body softening, and lentinan degradation. We constructed a de novo assembly draft genome sequence and performed gene prediction of Lentinula edodesDe novo assembly was carried out using short reads from paired-end and mate-paired libraries and long reads by PacBio, resulting in a contig number of 1951 and an N50 of 1 Mb. Further, we predicted genes by Augustus using RNA-seq data from the whole life cycle of Lentinula edodes, resulting in 12,959 predicted genes. This analysis revealed that Lentinula edodes lacks lignin peroxidase. To reveal genes involved in Lentinula edodes postharvest fruiting body quality loss, transcriptome analysis was carried out using Super-SAGE. This analysis revealed that many cell wall-related enzymes are upregulated after harvest, such as β-1,3-1,6-glucan-degrading enzymes in glycoside hydrolase (GH) families 5, 16, 30, 55, 128, and thaumatin-like proteins. In addition, we found several chitin-related genes are upregulated, such as putative chitinases in GH family18, exo-chitinases in GH 20, and a putative chitosanase in GH 75. The results suggest that cell wall-degrading enzymes synergistically cooperate for rapid fruiting body autolysis. Many putative transcription factor genes were upregulated postharvest, such as genes containing high mobility group (HMG) domains and zinc finger domains. Several cell death-related proteins were also upregulated postharvest.Importance Our data collectively suggest that there is a rapid fruiting body autolysis system in Lentinula edodes The genes for postharvest quality loss newly found in this research will be targets for future breeding of strains that can keep freshness longer than present strains. De novo Lentinula edodes genome assembly data will be used for construction of the complete Lentinula edodes chromosome map for the future

  2. RNA-seq analysis of Rubus idaeus cv. Nova: transcriptome sequencing and de novo assembly for subsequent functional genomics approaches.

    PubMed

    Hyun, Tae Kyung; Lee, Sarah; Kumar, Dhinesh; Rim, Yeonggil; Kumar, Ritesh; Lee, Sang Yeol; Lee, Choong Hwan; Kim, Jae-Yean

    2014-10-01

    Using Illumina sequencing technology, we have generated the large-scale transcriptome sequencing data containing abundant information on genes involved in the metabolic pathways in R. idaeus cv. Nova fruits. Rubus idaeus (Red raspberry) is one of the important economical crops that possess numerous nutrients, micronutrients and phytochemicals with essential health benefits to human. The molecular mechanism underlying the ripening process and phytochemical biosynthesis in red raspberry is attributed to the changes in gene expression, but very limited transcriptomic and genomic information in public databases is available. To address this issue, we generated more than 51 million sequencing reads from R. idaeus cv. Nova fruit using Illumina RNA-Seq technology. After de novo assembly, we obtained 42,604 unigenes with an average length of 812 bp. At the protein level, Nova fruit transcriptome showed 77 and 68 % sequence similarities with Rubus coreanus and Fragaria versa, respectively, indicating the evolutionary relationship between them. In addition, 69 % of assembled unigenes were annotated using public databases including NCBI non-redundant, Cluster of Orthologous Groups and Gene ontology database, suggesting that our transcriptome dataset provides a valuable resource for investigating metabolic processes in red raspberry. To analyze the relationship between several novel transcripts and the amounts of metabolites such as γ-aminobutyric acid and anthocyanins, real-time PCR and target metabolite analysis were performed on two different ripening stages of Nova. This is the first attempt using Illumina sequencing platform for RNA sequencing and de novo assembly of Nova fruit without reference genome. Our data provide the most comprehensive transcriptome resource available for Rubus fruits, and will be useful for understanding the ripening process and for breeding R. idaeus cultivars with improved fruit quality.

  3. Transcriptome analysis of resistant soybean roots infected by Meloidogyne javanica

    PubMed Central

    de Sá, Maria Eugênia Lisei; Conceição Lopes, Marcus José; de Araújo Campos, Magnólia; Paiva, Luciano Vilela; dos Santos, Regina Maria Amorim; Beneventi, Magda Aparecida; Firmino, Alexandre Augusto Pereira; de Sá, Maria Fátima Grossi

    2012-01-01

    Soybean is an important crop for Brazilian agribusiness. However, many factors can limit its production, especially root-knot nematode infection. Studies on the mechanisms employed by the resistant soybean genotypes to prevent infection by these nematodes are of great interest for breeders. For these reasons, the aim of this work is to characterize the transcriptome of soybean line PI 595099-Meloidogyne javanica interaction through expression analysis. Two cDNA libraries were obtained using a pool of RNA from PI 595099 uninfected and M. javanica (J2) infected roots, collected at 6, 12, 24, 48, 96, 144 and 192 h after inoculation. Around 800 ESTs (Expressed Sequence Tags) were sequenced and clustered into 195 clusters. In silico subtraction analysis identified eleven differentially expressed genes encoding putative proteins sharing amino acid sequence similarities by using BlastX: metallothionein, SLAH4 (SLAC1 Homologue 4), SLAH1 (SLAC1 Homologue 1), zinc-finger proteins, AN1-type proteins, auxin-repressed proteins, thioredoxin and nuclear transport factor 2 (NTF-2). Other genes were also found exclusively in nematode stressed soybean roots, such as NAC domain-containing proteins, MADS-box proteins, SOC1 (suppressor of overexpression of constans 1) proteins, thioredoxin-like protein 4-Coumarate-CoA ligase and the transcription factor (TF) MYBZ2. Among the genes identified in non-stressed roots only were Ser/Thr protein kinases, wound-induced basic protein, ethylene-responsive family protein, metallothionein-like protein cysteine proteinase inhibitor (cystatin) and Putative Kunitz trypsin protease inhibitor. An understanding of the roles of these differentially expressed genes will provide insights into the resistance mechanisms and candidate genes involved in soybean-M. javanica interaction and contribute to more effective control of this pathogen. PMID:22802712

  4. Transcriptome analysis of resistant soybean roots infected by Meloidogyne javanica.

    PubMed

    de Sá, Maria Eugênia Lisei; Conceição Lopes, Marcus José; de Araújo Campos, Magnólia; Paiva, Luciano Vilela; Dos Santos, Regina Maria Amorim; Beneventi, Magda Aparecida; Firmino, Alexandre Augusto Pereira; de Sá, Maria Fátima Grossi

    2012-06-01

    Soybean is an important crop for Brazilian agribusiness. However, many factors can limit its production, especially root-knot nematode infection. Studies on the mechanisms employed by the resistant soybean genotypes to prevent infection by these nematodes are of great interest for breeders. For these reasons, the aim of this work is to characterize the transcriptome of soybean line PI 595099-Meloidogyne javanica interaction through expression analysis. Two cDNA libraries were obtained using a pool of RNA from PI 595099 uninfected and M. javanica (J(2)) infected roots, collected at 6, 12, 24, 48, 96, 144 and 192 h after inoculation. Around 800 ESTs (Expressed Sequence Tags) were sequenced and clustered into 195 clusters. In silico subtraction analysis identified eleven differentially expressed genes encoding putative proteins sharing amino acid sequence similarities by using BlastX: metallothionein, SLAH4 (SLAC1 Homologue 4), SLAH1 (SLAC1 Homologue 1), zinc-finger proteins, AN1-type proteins, auxin-repressed proteins, thioredoxin and nuclear transport factor 2 (NTF-2). Other genes were also found exclusively in nematode stressed soybean roots, such as NAC domain-containing proteins, MADS-box proteins, SOC1 (suppressor of overexpression of constans 1) proteins, thioredoxin-like protein 4-Coumarate-CoA ligase and the transcription factor (TF) MYBZ2. Among the genes identified in non-stressed roots only were Ser/Thr protein kinases, wound-induced basic protein, ethylene-responsive family protein, metallothionein-like protein cysteine proteinase inhibitor (cystatin) and Putative Kunitz trypsin protease inhibitor. An understanding of the roles of these differentially expressed genes will provide insights into the resistance mechanisms and candidate genes involved in soybean-M. javanica interaction and contribute to more effective control of this pathogen.

  5. Transcriptomic analysis of cyclic AMP response in bovine cumulus cells.

    PubMed

    Khan, D R; Guillemette, C; Sirard, M A; Richard, F J

    2015-09-01

    Acquisition of oocyte developmental competence needs to be understood to improve clinical outcomes of assisted reproduction. The stimulation of cumulus cell concentration of cyclic adenosine 3'5'-monophosphate (cAMP) by pharmacological agents during in vitro maturation (IVM) participates in improvement of oocyte quality. However, precise coordination and downstream targets of cAMP signaling in cumulus cells are largely unknown. We have previously demonstrated better embryo development after cAMP stimulation for first 6 h during IVM. Using this model, we investigated cAMP signaling in cumulus cells through in vitro culture of cumulus-oocyte complexes (COCs) in the presence of cAMP raising agents: forskolin, IBMX, and dipyridamole (here called FID treatment). Transcriptomic analysis of cumulus cells indicated that FID-induced differentially expressed transcripts were implicated in cumulus expansion, steroidogenesis, cell metabolism, and oocyte competence. Functional genomic analysis revealed that protein kinase-A (PKA), extracellular signal regulated kinases (ERK1/2), and calcium (Ca(2+)) pathways as key regulators of FID signaling. Inhibition of PKA (H89) in FID-supplemented COCs or substitution of FID with calcium ionophore (A23187) demonstrated that FID activated primarily the PKA pathway which inhibited ERK1/2 phosphorylation and was upstream of calcium signaling. Furthermore, inhibition of ERK1/2 phosphorylation by FID supported a regulation by dual specific phosphatase (DUSP1) via PKA. Our findings imply that cAMP (FID) regulates cell metabolism, steroidogenesis, intracellular signaling and cumulus expansion through PKA which modulates these functions through optimization of ERK1/2 phosphorylation and coordination of calcium signaling. These findings have implications for development of new strategies for improving oocyte in vitro maturation leading to better developmental competence.

  6. Spider Transcriptomes Identify Ancient Large-Scale Gene Duplication Event Potentially Important in Silk Gland Evolution

    PubMed Central

    Clarke, Thomas H.; Garb, Jessica E.; Hayashi, Cheryl Y.; Arensburger, Peter; Ayoub, Nadia A.

    2015-01-01

    The evolution of specialized tissues with novel functions, such as the silk synthesizing glands in spiders, is likely an influential driver of adaptive success. Large-scale gene duplication events and subsequent paralog divergence are thought to be required for generating evolutionary novelty. Such an event has been proposed for spiders, but not tested. We de novo assembled transcriptomes from three cobweb weaving spider species. Based on phylogenetic analyses of gene families with representatives from each of the three species, we found numerous duplication events indicative of a whole genome or segmental duplication. We estimated the age of the gene duplications relative to several speciation events within spiders and arachnids and found that the duplications likely occurred after the divergence of scorpions (order Scorpionida) and spiders (order Araneae), but before the divergence of the spider suborders Mygalomorphae and Araneomorphae, near the evolutionary origin of spider silk glands. Transcripts that are expressed exclusively or primarily within black widow silk glands are more likely to have a paralog descended from the ancient duplication event and have elevated amino acid replacement rates compared with other transcripts. Thus, an ancient large-scale gene duplication event within the spider lineage was likely an important source of molecular novelty during the evolution of silk gland-specific expression. This duplication event may have provided genetic material for subsequent silk gland diversification in the true spiders (Araneomorphae). PMID:26058392

  7. Spider Transcriptomes Identify Ancient Large-Scale Gene Duplication Event Potentially Important in Silk Gland Evolution.

    PubMed

    Clarke, Thomas H; Garb, Jessica E; Hayashi, Cheryl Y; Arensburger, Peter; Ayoub, Nadia A

    2015-06-08

    The evolution of specialized tissues with novel functions, such as the silk synthesizing glands in spiders, is likely an influential driver of adaptive success. Large-scale gene duplication events and subsequent paralog divergence are thought to be required for generating evolutionary novelty. Such an event has been proposed for spiders, but not tested. We de novo assembled transcriptomes from three cobweb weaving spider species. Based on phylogenetic analyses of gene families with representatives from each of the three species, we found numerous duplication events indicative of a whole genome or segmental duplication. We estimated the age of the gene duplications relative to several speciation events within spiders and arachnids and found that the duplications likely occurred after the divergence of scorpions (order Scorpionida) and spiders (order Araneae), but before the divergence of the spider suborders Mygalomorphae and Araneomorphae, near the evolutionary origin of spider silk glands. Transcripts that are expressed exclusively or primarily within black widow silk glands are more likely to have a paralog descended from the ancient duplication event and have elevated amino acid replacement rates compared with other transcripts. Thus, an ancient large-scale gene duplication event within the spider lineage was likely an important source of molecular novelty during the evolution of silk gland-specific expression. This duplication event may have provided genetic material for subsequent silk gland diversification in the true spiders (Araneomorphae). © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  8. Transcriptomic analysis of porcine PBMCs in response to FMDV infection.

    PubMed

    Zhao, Fu-Rong; Xie, Yin-Li; Liu, Ze-Zhong; Shao, Jun-Jun; Li, Shi-Fang; Zhang, Yong-Guang; Chang, Hui-Yun

    2017-09-01

    one or more of the three categories: biological process, cellular component, and molecular function. According to KEGG analysis,the main pathway was represented including protein processing in endoplasmic reticulum, phagosome, cell cycle and cytokine-cytokine receptor interaction. Some key DE genes related to immune process and signaling pathways were analyzed and quantified by RT-PCR. This is the first systematical transcriptome analysis of pig PBMCs infected by FMDV. These findings will help us better understand the host Cell-FMDV interaction and its relationship to pathogenesis, as well as contribute to the prevention and control of FMDV. Copyright © 2017. Published by Elsevier B.V.

  9. Transcriptome analysis of the silkworm (Bombyx mori) by high-throughput RNA sequencing.

    PubMed

    Li, Yinü; Wang, Guozeng; Tian, Jian; Liu, Huifen; Yang, Huipeng; Yi, Yongzhu; Wang, Jinhui; Shi, Xiaofeng; Jiang, Feng; Yao, Bin; Zhang, Zhifang

    2012-01-01

    The domestic silkworm, Bombyx mori, is a model insect with important economic value for silk production that also acts as a bioreactor for biomaterial production. The functional complexity of the silkworm transcriptome has not yet been fully elucidated, although genomic sequencing and other tools have been widely used in its study. We explored the transcriptome of silkworm at different developmental stages using high-throughput paired-end RNA sequencing. A total of about 3.3 gigabases (Gb) of sequence was obtained, representing about a 7-fold coverage of the B. mori genome. From the reads that were mapped to the genome sequence; 23,461 transcripts were obtained, 5,428 of them were novel. Of the 14,623 predicted protein-coding genes in the silkworm genome database, 11,884 of them were found to be expressed in the silkworm transcriptome, giving a coverage of 81.3%. A total of 13,195 new exons were detected, of which, 5,911 were found in the annotated genes in the Silkworm Genome Database (SilkDB). An analysis of alternative splicing in the transcriptome revealed that 3,247 genes had undergone alternative splicing. To help with the data analysis, a transcriptome database that integrates our transcriptome data with the silkworm genome data was constructed and is publicly available at http://124.17.27.136/gbrowse2/. To our knowledge, this is the first study to elucidate the silkworm transcriptome using high-throughput RNA sequencing technology. Our data indicate that the transcriptome of silkworm is much more complex than previously anticipated. This work provides tools and resources for the identification of new functional elements and paves the way for future functional genomics studies.

  10. Analysis of Saprolegnia parasitica Transcriptome following Treatment with Copper Sulfate.

    PubMed

    Hu, Kun; Ma, Rong-Rong; Cheng, Jun-Ming; Ye, Xin; Sun, Qi; Yuan, Hai-Lan; Liang, Nan; Fang, Wen-Hong; Li, Hao-Ran; Yang, Xian-Le

    2016-01-01

    Massive infection caused by oomycete fungus Saprolegnia parasitica is detrimental to freshwater fish. Recently, we showed that copper sulfate demonstrated good efficacy for controlling S. parasitica infection in grass carp. In this study, we investigated the mechanism of inhibition of S. parasitica growth by copper sulfate by analyzing the transcriptome of copper sulfate-treated S. parasitica. To examine the mechanism of copper sulfate inhibiting S. parasitica, we utilized RNA-seq technology to compare differential gene expression in S. parasitica treated with or without copper sulfate. The total mapped rates of the reads with the reference genome were 90.50% in the control group and 73.50% in the experimental group. In the control group, annotated splice junctions, partial novel splice junctions and complete novel splice junctions were about 83%, 3% and 14%, respectively. In the treatment group, the corresponding values were about 75%, 6% and 19%. Following copper sulfate treatment, a total 310 genes were markedly upregulated and 556 genes were markedly downregulated in S. parasitica. Material metabolism related GO terms including cofactor binding (33 genes), 1,3-beta-D-glucan synthase complex (4 genes), carboxylic acid metabolic process (40 genes) were the most significantly enriched. KEGG pathway analysis also determined that the metabolism-related biological pathways were significantly enriched, including the metabolic pathways (98 genes), biosynthesis of secondary metabolites pathways (42 genes), fatty acid metabolism (13 genes), phenylalanine metabolism (7 genes), starch and sucrose metabolism pathway (12 genes). The qRT-PCR results were largely consistent with the RNA-Seq results. Our results indicate that copper sulfate inhibits S. parasitica growth by affecting multiple biological functions, including protein synthesis, energy biogenesis, and metabolism.

  11. Neuropeptidomics in Triatoma infestans. Comparative transcriptomic analysis among triatomines.

    PubMed

    Traverso, Lucila; Sierra, Ivana; Sterkel, Marcos; Francini, Flavio; Ons, Sheila

    2016-12-18

    Chagas' disease, affecting up to 6-7 million people worldwide, is transmitted to humans through the feces of triatomine kissing bugs. From these, Rhodnius prolixus, Triatoma dimidiata, Triatoma infestans and Triatoma pallidipennis are important vectors distributed throughout the Latin American subcontinent. Resistance to pyrethroids has been developed by some triatomine populations, especially T. infestans, obstructing their control. Given their role in the regulation of physiological processes, neuroendocrine-derived factors have been proposed as a source of molecular targets for new-generation insecticides. However, the involvement of neuropeptides in insecticide metabolism and resistance in insects has been poorly studied. In the present work, the sequences of 20 neuropeptide precursor genes in T. infestans, 16 in T. dimidiata, and 13 in T. pallidipennis detected in transcriptomic databases are reported, and a comparative analysis in triatomines is presented. A total of 59 neuropeptides were validated by liquid chromatography-tandem mass spectrometry in brain and nervous ganglia from T. infestans, revealing the existence of differential post-translational modifications, extended and truncated forms. The results suggest a high sequence conservation in some neuropeptide systems in triatomines, whereas remarkable differences occur in several others within the core domains. Comparisons of the basal expression levels for several neuropeptide precursor genes between pyrethroid sensitive and resistant population of T. infestans are also presented here, in order to introduce a proof of concept to test the involvement of neuropeptides in insecticide resistance. From the precursors tested, NVP and ITG peptides are significantly higher expressed in the resistant population. To our knowledge, this is the first report to associate differential neuropeptide expression with insecticide resistance. The information provided here contributes to creating conditions to widely

  12. Analysis of Saprolegnia parasitica Transcriptome following Treatment with Copper Sulfate

    PubMed Central

    Ye, Xin; Sun, Qi; Yuan, Hai-Lan; Liang, Nan; Fang, Wen-Hong; Li, Hao-Ran; Yang, Xian-Le

    2016-01-01

    Background Massive infection caused by oomycete fungus Saprolegnia parasitica is detrimental to freshwater fish. Recently, we showed that copper sulfate demonstrated good efficacy for controlling S. parasitica infection in grass carp. In this study, we investigated the mechanism of inhibition of S. parasitica growth by copper sulfate by analyzing the transcriptome of copper sulfate—treated S. parasitica. To examine the mechanism of copper sulfate inhibiting S. parasitica, we utilized RNA-seq technology to compare differential gene expression in S. parasitica treated with or without copper sulfate. Results The total mapped rates of the reads with the reference genome were 90.50% in the control group and 73.50% in the experimental group. In the control group, annotated splice junctions, partial novel splice junctions and complete novel splice junctions were about 83%, 3% and 14%, respectively. In the treatment group, the corresponding values were about 75%, 6% and 19%. Following copper sulfate treatment, a total 310 genes were markedly upregulated and 556 genes were markedly downregulated in S. parasitica. Material metabolism related GO terms including cofactor binding (33 genes), 1,3-beta-D-glucan synthase complex (4 genes), carboxylic acid metabolic process (40 genes) were the most significantly enriched. KEGG pathway analysis also determined that the metabolism-related biological pathways were significantly enriched, including the metabolic pathways (98 genes), biosynthesis of secondary metabolites pathways (42 genes), fatty acid metabolism (13 genes), phenylalanine metabolism (7 genes), starch and sucrose metabolism pathway (12 genes). The qRT-PCR results were largely consistent with the RNA-Seq results. Conclusion Our results indicate that copper sulfate inhibits S. parasitica growth by affecting multiple biological functions, including protein synthesis, energy biogenesis, and metabolism. PMID:26895329

  13. RNA-Seq transcriptome analysis of Spirodela dormancy without reproduction

    PubMed Central

    2014-01-01

    Background Higher plants exhibit a remarkable phenotypic plasticity to adapt to adverse environmental changes. The Greater Duckweed Spirodela, as an aquatic plant, presents exceptional tolerance to cold winters through its dormant structure of turions in place of seeds. Abundant starch in turions permits them to sink and escape the freezing surface of waters. Due to their clonal propagation, they are the fastest growing biomass on earth, providing yet an untapped source for industrial applications. Results We used next generation sequencing technology to examine the transcriptome of turion development triggered by exogenous ABA. A total of 208 genes showed more than a 4-fold increase compared with 154 down-regulated genes in developing turions. The analysis of up-regulated differential expressed genes in response to dormancy exposed an enriched interplay among various pathways: signal transduction, seed dehydration, carbohydrate and secondary metabolism, and senescence. On the other side, the genes responsible for rapid growth and biomass accumulation through DNA assembly, protein synthesis and carbon fixation are repressed. Noticeably, three members of late embryogenesis abundant protein family are exclusively expressed during turion formation. High expression level of key genes in starch synthesis are APS1, APL3 and GBSSI, which could artificially be reduced for re-directing carbon flow from photosynthesis to create a higher energy biomass. Conclusions The identification and functional annotation of differentially expressed genes open a major step towards understanding the molecular network underlying vegetative frond dormancy. Moreover, genes have been identified that could be engineered in duckweeds for practical applications easing agricultural production of food crops. PMID:24456086

  14. Comparative transcriptome analysis between planarian Dugesia japonica and other platyhelminth species.

    PubMed

    Nishimura, Osamu; Hirao, Yukako; Tarui, Hiroshi; Agata, Kiyokazu

    2012-06-29

    Planarians are considered to be among the extant animals close to one of the earliest groups of organisms that acquired a central nervous system (CNS) during evolution. Planarians have a bilobed brain with nine lateral branches from which a variety of external signals are projected into different portions of the main lobes. Various interneurons process different signals to regulate behavior and learning/memory. Furthermore, planarians have robust regenerative ability and are attracting attention as a new model organism for the study of regeneration. Here we conducted large-scale EST analysis of the head region of the planarian Dugesia japonica to construct a database of the head-region transcriptome, and then performed comparative analyses among related species. A total of 54,752 high-quality EST reads were obtained from a head library of the planarian Dugesia japonica, and 13,167 unigene sequences were produced by de novo assembly. A new method devised here revealed that proteins related to metabolism and defense mechanisms have high flexibility of amino-acid substitutions within the planarian family. Eight-two CNS-development genes were found in the planarian (cf. C. elegans 3; chicken 129). Comparative analysis revealed that 91% of the planarian CNS-development genes could be mapped onto the schistosome genome, but one-third of these shared genes were not expressed in the schistosome. We constructed a database that is a useful resource for comparative planarian transcriptome studies. Analysis comparing homologous genes between two planarian species showed that the potential of genes is important for accumulation of amino-acid substitutions. The presence of many CNS-development genes in our database supports the notion that the planarian has a fundamental brain with regard to evolution and development at not only the morphological/functional, but also the genomic, level. In addition, our results indicate that the planarian CNS-development genes already existed

  15. A comprehensive analysis of the human placenta transcriptome

    USDA-ARS?s Scientific Manuscript database

    As the conduit for nutrients and growth signals, the placenta is critical to establishing an environment sufficient for fetal growth and development. To better understand the mechanisms regulating placental development and gene expression, we characterized the transcriptome of term placenta from 20 ...

  16. Characterization and Transcriptome Analysis of Mycobacterium tuberculosis Persisters

    PubMed Central

    Keren, Iris; Minami, Shoko; Rubin, Eric; Lewis, Kim

    2011-01-01

    ABSTRACT Tuberculosis continues to be a major public health problem in many parts of the world. Significant obstacles in controlling the epidemic are the length of treatment and the large reservoir of latently infected people. Bacteria form dormant, drug-tolerant persister cells, which may be responsible for the difficulty in treating both acute and latent infections. We find that in Mycobacterium  tuberculosis, low numbers of drug-tolerant persisters are present in lag and early exponential phases, increasing sharply at late exponential and stationary phases to make up ~1% of the population. This suggests that persister formation is governed by both stochastic and deterministic mechanisms. In order to isolate persisters, an exponentially growing population was treated with d-cycloserine, and cells surviving lysis were collected by centrifugation. A transcriptome of persisters was obtained by using hybridization to an Affymetrix array. The transcriptome shows downregulation of metabolic and biosynthetic pathways, consistent with a certain degree of dormancy. A set of genes was upregulated in persisters, and these are likely involved in persister formation and maintenance. A comparison of the persister transcriptome with transcriptomes obtained for several in vitro dormancy models identified a small number of genes upregulated in all cases, which may represent a core dormancy response. PMID:21673191

  17. Analysis, annotation, and profiling of the oat seed transcriptome

    USDA-ARS?s Scientific Manuscript database

    Novel high-throughput next generation sequencing (NGS) technologies are providing opportunities to explore genomes and transcriptomes in a cost-effective manner. To construct a gene expression atlas of developing oat (Avena sativa) seeds, two software packages specifically designed for RNA-seq (Trin...

  18. Comparative Transcriptome Analysis Reveals Substantial Tissue Specificity in Human Aortic Valve

    PubMed Central

    Wang, Jun; Wang, Ying; Gu, Weidong; Ni, Buqing; Sun, Haoliang; Yu, Tong; Gu, Wanjun; Chen, Liang; Shao, Yongfeng

    2016-01-01

    RNA sequencing (RNA-seq) has revolutionary roles in transcriptome identification and quantification of different types of tissues and cells in many organisms. Although numerous RNA-seq data derived from many types of human tissues and cell lines, little is known on the transcriptome repertoire of human aortic valve. In this study, we sequenced the total RNA prepared from two calcified human aortic valves and reported the whole transcriptome of human aortic valve. Integrating RNA-seq data of 13 human tissues from Human Body Map 2 Project, we constructed a transcriptome repertoire of human tissues, including 19,505 protein-coding genes and 4,948 long intergenic noncoding RNAs (lincRNAs). Among them, 263 lincRNAs were identified as novel noncoding transcripts in our data. By comparing transcriptome data among different human tissues, we observed substantial tissue specificity of RNA transcripts, both protein-coding genes and lincRNAs, in human aortic valve. Further analysis revealed that aortic valve-specific lincRNAs were more likely to be recently derived from repetitive elements in the primate lineage, but were less likely to be conserved at the nucleotide level. Expression profiling analysis showed significant lower expression levels of aortic valve-specific protein-coding genes and lincRNA genes, when compared with genes that were universally expressed in various tissues. Isoform-level expression analysis also showed that a majority of mRNA genes had a major isoform expressed in the human aortic valve. To our knowledge, this is the first comparative transcriptome analysis between human aortic valve and other human tissues. Our results are helpful to understand the transcriptome diversity of human tissues and the underlying mechanisms that drive tissue specificity of protein-coding genes and lincRNAs in human aortic valve. PMID:27493474

  19. Integrative approaches for large-scale transcriptome-wide association studies

    PubMed Central

    Gusev, Alexander; Ko, Arthur; Shi, Huwenbo; Bhatia, Gaurav; Chung, Wonil; Penninx, Brenda W J H; Jansen, Rick; de Geus, Eco JC; Boomsma, Dorret I; Wright, Fred A; Sullivan, Patrick F; Nikkola, Elina; Alvarez, Marcus; Civelek, Mete; Lusis, Aldons J.; Lehtimäki, Terho; Raitoharju, Emma; Kähönen, Mika; Seppälä, Ilkka; Raitakari, Olli T.; Kuusisto, Johanna; Laakso, Markku; Price, Alkes L.; Pajukanta, Päivi; Pasaniuc, Bogdan

    2016-01-01

    Many genetic variants influence complex traits by modulating gene expression, thus altering the abundance levels of one or multiple proteins. Here, we introduce a powerful strategy that integrates gene expression measurements with summary association statistics from large-scale genome-wide association studies (GWAS) to identify genes whose cis-regulated expression is associated to complex traits. We leverage expression imputation to perform a transcriptome wide association scan (TWAS) to identify significant expression-trait associations. We applied our approaches to expression data from blood and adipose tissue measured in ~3,000 individuals overall. We imputed gene expression into GWAS data from over 900,000 phenotype measurements to identify 69 novel genes significantly associated to obesity-related traits (BMI, lipids, and height). Many of the novel genes are associated with relevant phenotypes in the Hybrid Mouse Diversity Panel. Our results showcase the power of integrating genotype, gene expression and phenotype to gain insights into the genetic basis of complex traits. PMID:26854917

  20. Integrative approaches for large-scale transcriptome-wide association studies.

    PubMed

    Gusev, Alexander; Ko, Arthur; Shi, Huwenbo; Bhatia, Gaurav; Chung, Wonil; Penninx, Brenda W J H; Jansen, Rick; de Geus, Eco J C; Boomsma, Dorret I; Wright, Fred A; Sullivan, Patrick F; Nikkola, Elina; Alvarez, Marcus; Civelek, Mete; Lusis, Aldons J; Lehtimäki, Terho; Raitoharju, Emma; Kähönen, Mika; Seppälä, Ilkka; Raitakari, Olli T; Kuusisto, Johanna; Laakso, Markku; Price, Alkes L; Pajukanta, Päivi; Pasaniuc, Bogdan

    2016-03-01

    Many genetic variants influence complex traits by modulating gene expression, thus altering the abundance of one or multiple proteins. Here we introduce a powerful strategy that integrates gene expression measurements with summary association statistics from large-scale genome-wide association studies (GWAS) to identify genes whose cis-regulated expression is associated with complex traits. We leverage expression imputation from genetic data to perform a transcriptome-wide association study (TWAS) to identify significant expression-trait associations. We applied our approaches to expression data from blood and adipose tissue measured in ∼ 3,000 individuals overall. We imputed gene expression into GWAS data from over 900,000 phenotype measurements to identify 69 new genes significantly associated with obesity-related traits (BMI, lipids and height). Many of these genes are associated with relevant phenotypes in the Hybrid Mouse Diversity Panel. Our results showcase the power of integrating genotype, gene expression and phenotype to gain insights into the genetic basis of complex traits.

  1. Comparative Analysis of the Testis and Ovary Transcriptomes in Zebrafish by Combining Experimental and Computational Tools

    PubMed Central

    Li, Yang; Chia, Jer Ming; Bartfai, Richard; Christoffels, Alan; Yue, Gen Hua; Ding, Ke; Ho, Mei Yin; Hill, James A.

    2004-01-01

    Studies on the zebrafish model have contributed to our understanding of several important developmental processes, especially those that can be easily studied in the embryo. However, our knowledge on late events such as gonad differentiation in the zebrafish is still limited. Here we provide an analysis on the gene sets expressed in the adult zebrafish testis and ovary in an attempt to identify genes with potential role in (zebra)fish gonad development and function. We produced 10 533 expressed sequence tags (ESTs) from zebrafish testis or ovary and downloaded an additional 23 642 gonad-derived sequences from the zebrafish EST database. We clustered these sequences together with over 13 000 kidney-derived zebrafish ESTs to study partial transcriptomes for these three organs. We searched for genes with gonad-specific expression by screening macroarrays containing at least 2600 unique cDNA inserts with testis-, ovary- and kidney-derived cDNA probes. Clones hybridizing to only one of the two gonad probes were selected, and subsequently screened with computational tools to identify 72 genes with potentially testis-specific and 97 genes with potentially ovary-specific expression, respectively. PCR-amplification confirmed gonad-specificity for 21 of the 45 clones tested (all without known function). Our study, which involves over 47 000 EST sequences and specialized cDNA arrays, is the first analysis of adult organ transcriptomes of zebrafish at such a scale. The study of genes expressed in adult zebrafish testis and ovary will provide useful information on regulation of gene expression in teleost gonads and might also contribute to our understanding of the development and differentiation of reproductive organs in vertebrates. PMID:18629171

  2. Principles of transcriptome analysis and gene expression quantification: an RNA-seq tutorial.

    PubMed

    Wolf, Jochen B W

    2013-07-01

    Genome-wide analyses and high-throughput screening was long reserved for biomedical applications and genetic model organisms. With the rapid development of massively parallel sequencing nanotechnology (or next-generation sequencing) and simultaneous maturation of bioinformatic tools, this situation has dramatically changed. Genome-wide thinking is forging its way into disciplines like evolutionary biology or molecular ecology that were historically confined to small-scale genetic approaches. Accessibility to genome-scale information is transforming these fields, as it allows us to answer long-standing questions like the genetic basis of local adaptation and speciation or the evolution of gene expression profiles that until recently were out of reach. Many in the eco-evolutionary sciences will be working with large-scale genomic data sets, and a basic understanding of the concepts and underlying methods is necessary to judge the work of others. Here, I briefly introduce next-generation sequencing and then focus on transcriptome shotgun sequencing (RNA-seq). This article gives a broad overview and provides practical guidance for the many steps involved in a typical RNA-seq work flow from sampling, to RNA extraction, library preparation and data analysis. I focus on principles, present useful tools where appropriate and point out where caution is needed or progress to be expected. This tutorial is mostly targeted at beginners, but also contains potentially useful reflections for the more experienced.

  3. Cancer Transcriptome Dataset Analysis: Comparing Methods of Pathway and Gene Regulatory Network-Based Cluster Identification.

    PubMed

    Nam, Seungyoon

    2017-04-01

    Cancer transcriptome analysis is one of the leading areas of Big Data science, biomarker, and pharmaceutical discovery, not to forget personalized medicine. Yet, cancer transcriptomics and postgenomic medicine require innovation in bioinformatics as well as comparison of the performance of available algorithms. In this data analytics context, the value of network generation and algorithms has been widely underscored for addressing the salient questions in cancer pathogenesis. Analysis of cancer trancriptome often results in complicated networks where identification of network modularity remains critical, for example, in delineating the "druggable" molecular targets. Network clustering is useful, but depends on the network topology in and of itself. Notably, the performance of different network-generating tools for network cluster (NC) identification has been little investigated to date. Hence, using gastric cancer (GC) transcriptomic datasets, we compared two algorithms for generating pathway versus gene regulatory network-based NCs, showing that the pathway-based approach better agrees with a reference set of cancer-functional contexts. Finally, by applying pathway-based NC identification to GC transcriptome datasets, we describe cancer NCs that associate with candidate therapeutic targets and biomarkers in GC. These observations collectively inform future research on cancer transcriptomics, drug discovery, and rational development of new analysis tools for optimal harnessing of omics data.

  4. Transcriptomic and proteomic analyses on the supercooling ability and mining of antifreeze proteins of the Chinese white wax scale insect.

    PubMed

    Yu, Shu-Hui; Yang, Pu; Sun, Tao; Qi, Qian; Wang, Xue-Qing; Chen, Xiao-Ming; Feng, Ying; Liu, Bo-Wen

    2016-06-01

    The Chinese white wax scale insect, Ericerus pela, can survive at extremely low temperatures, and some overwintering individuals exhibit supercooling at temperatures below -30°C. To investigate the deep supercooling ability of E. pela, transcriptomic and proteomic analyses were performed to delineate the major gene and protein families responsible for the deep supercooling ability of overwintering females. Gene Ontology (GO) classification and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that genes involved in the mitogen-activated protein kinase, calcium, and PI3K-Akt signaling pathways and pathways associated with the biosynthesis of soluble sugars, sugar alcohols and free amino acids were dominant. Proteins responsible for low-temperature stress, such as cold acclimation proteins, glycerol biosynthesis-related enzymes and heat shock proteins (HSPs) were identified. However, no antifreeze proteins (AFPs) were identified through sequence similarity search methods. A random forest approach identified 388 putative AFPs in the proteome. The AFP gene ep-afp was expressed in Escherichia coli, and the expressed protein exhibited a thermal hysteresis activity of 0.97°C, suggesting its potential role in the deep supercooling ability of E. pela.

  5. Detection of driver protein complexes in breast cancer metastasis by large-scale transcriptome-interactome integration.

    PubMed

    Garcia, Maxime; Finetti, Pascal; Bertucci, Francois; Birnbaum, Daniel; Bidaut, Ghislain

    2014-01-01

    With the development of high-throughput gene expression profiling technologies came the opportunity to define genomic signatures predicting clinical condition or cancer patient outcome. However, such signatures show dependency on training set, lack of generalization, and instability, partly due to microarray data topology. Additional issues for analyzing tumor gene expression are that subtle molecular perturbations in driver genes leading to cancer and metastasis (masked in typical differential expression analysis) may provoke expression changes of greater amplitude in downstream genes (easily detected). In this chapter, we are describing an interactome-based algorithm, Interactome-Transcriptome Integration (ITI) that is used to find a generalizable signature for prediction of breast cancer relapse by superimposition of a large-scale protein-protein interaction data (human interactome) over several gene expression datasets. ITI extracts regions in the interactome whose expression is discriminating for predicting relapse-free survival in cancer and allow detection of subnetworks that constitutes a generalizable and stable genomic signature. In this chapter, we describe the practical aspects of running the full ITI pipeline (subnetwork detection and classification) on six microarray datasets.

  6. Transcriptome analysis of the response of Burmese python to digestion

    PubMed Central

    Sanggaard, Kristian Wejse; Schauser, Leif; Lauridsen, Sanne Enok; Enghild, Jan J.

    2017-01-01

    Abstract Exceptional and extreme feeding behaviour makes the Burmese python (Python bivittatus) an interesting model to study physiological remodelling and metabolic adaptation in response to refeeding after prolonged starvation. In this study, we used transcriptome sequencing of 5 visceral organs during fasting as well as 24 hours and 48 hours after ingestion of a large meal to unravel the postprandial changes in Burmese pythons. We first used the pooled data to perform a de novo assembly of the transcriptome and supplemented this with a proteomic survey of enzymes in the plasma and gastric fluid. We constructed a high-quality transcriptome with 34 423 transcripts, of which 19 713 (57%) were annotated. Among highly expressed genes (fragments per kilo base per million sequenced reads > 100 in 1 tissue), we found that the transition from fasting to digestion was associated with differential expression of 43 genes in the heart, 206 genes in the liver, 114 genes in the stomach, 89 genes in the pancreas, and 158 genes in the intestine. We interrogated the function of these genes to test previous hypotheses on the response to feeding. We also used the transcriptome to identify 314 secreted proteins in the gastric fluid of the python. Digestion was associated with an upregulation of genes related to metabolic processes, and translational changes therefore appear to support the postprandial rise in metabolism. We identify stomach-related proteins from a digesting individual and demonstrate that the sensitivity of modern liquid chromatography/tandem mass spectrometry equipment allows the identification of gastric juice proteins that are present during digestion. PMID:28873961

  7. Transcriptomic Dose-Response Analysis for Mode of Action ...

    EPA Pesticide Factsheets

    Microarray and RNA-seq technologies can play an important role in assessing the health risks associated with environmental exposures. The utility of gene expression data to predict hazard has been well documented. Early toxicogenomics studies used relatively high, single doses with minimal replication. Thus, they were not useful in understanding health risks at environmentally-relevant doses. Until the past decade, application of toxicogenomics in dose response assessment and determination of chemical mode of action has been limited. New transcriptomic biomarkers have evolved to detect chemical hazards in multiple tissues together with pathway methods to study biological effects across the full dose response range and critical time course. Comprehensive low dose datasets are now available and with the use of transcriptomic benchmark dose estimation techniques within a mode of action framework, the ability to incorporate informative genomic data into human health risk assessment has substantially improved. The key advantage to applying transcriptomic technology to risk assessment is both the sensitivity and comprehensive examination of direct and indirect molecular changes that lead to adverse outcomes. Book Chapter with topic on future application of toxicogenomics technologies for MoA and risk assessment

  8. Transcriptome Analysis of Spartina pectinata in Response to Freezing Stress.

    PubMed

    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.

  9. Analysis of the transcriptome of the Indonesian coelacanth Latimeria menadoensis.

    PubMed

    Pallavicini, Alberto; Canapa, Adriana; Barucca, Marco; Alfőldi, Jessica; Biscotti, Maria Assunta; Buonocore, Francesco; De Moro, Gianluca; Di Palma, Federica; Fausto, Anna Maria; Forconi, Mariko; Gerdol, Marco; Makapedua, Daisy Monica; Turner-Meier, Jason; Olmo, Ettore; Scapigliati, Giuseppe

    2013-08-08

    Latimeria menadoensis is a coelacanth species first identified in 1997 in Indonesia, at 10,000 Km of distance from its African congener. To date, only six specimens have been caught and just a very limited molecular data is available. In the present work we describe the de novo transcriptome assembly obtained from liver and testis samples collected from the fifth specimen ever caught of this species. The deep RNA sequencing performed with Illumina technologies generated 145,435,156 paired-end reads, accounting for ~14 GB of sequence data, which were de novo assembled using a Trinity/CLC combined strategy. The assembly output was processed and filtered producing a set of 66,308 contigs, whose quality was thoroughly assessed. The comparison with the recently sequenced genome of the African congener Latimeria chalumnae and with the available genomic resources of other vertebrates revealed a good reconstruction of full length transcripts and a high coverage of the predicted full coelacanth transcriptome. Given the high genomic affinity between the two coelacanth species, the here described de novo transcriptome assembly can be considered a valuable support tool for the improvement of gene prediction within the genome of L. chalumnae and a valuable resource for investigation of many aspects of tetrapod evolution.

  10. 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

  11. Transcriptome analysis of Enterococcus faecalis in response to alkaline stress

    PubMed Central

    Ran, Shujun; Liu, Bin; Jiang, Wei; Sun, Zhe; Liang, Jingping

    2015-01-01

    Enterococcus faecalis is the most commonly isolated species from endodontic failure root canals; its persistence in treated root canals has been attributed to its ability to resist high pH stress. The goal of this study was to characterize the E. faecalis transcriptome and to identify candidate genes for response and resistance to alkaline stress using Illumina HiSeq 2000 sequencing. We found that E. faecalis could survive and form biofilms in a pH 10 environment and that alkaline stress had a great impact on the transcription of many genes in the E. faecalis genome. The transcriptome sequencing results revealed that 613 genes were differentially expressed (DEGs) for E. faecalis grown in pH 10 medium; 211 genes were found to be differentially up-regulated and 402 genes differentially down-regulated. Many of the down-regulated genes found are involved in cell energy production and metabolism and carbohydrate and amino acid metabolism, and the up-regulated genes are mostly related to nucleotide transport and metabolism. The results presented here reveal that cultivation of E. faecalis in alkaline stress has a profound impact on its transcriptome. The observed regulation of genes and pathways revealed that E. faecalis reduced its carbohydrate and amino acid metabolism and increased nucleotide synthesis to adapt and grow in alkaline stress. A number of the regulated genes may be useful candidates for the development of new therapeutic approaches for the treatment of E. faecalis infections. PMID:26300863

  12. 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.

  13. Machine learning-based differential network analysis: a study of stress-responsive transcriptomes in Arabidopsis.

    PubMed

    Ma, Chuang; Xin, Mingming; Feldmann, Kenneth A; Wang, Xiangfeng

    2014-02-01

    Machine learning (ML) is an intelligent data mining technique that builds a prediction model based on the learning of prior knowledge to recognize patterns in large-scale data sets. We present an ML-based methodology for transcriptome analysis via comparison of gene coexpression networks, implemented as an R package called machine learning-based differential network analysis (mlDNA) and apply this method to reanalyze a set of abiotic stress expression data in Arabidopsis thaliana. The mlDNA first used a ML-based filtering process to remove nonexpressed, constitutively expressed, or non-stress-responsive "noninformative" genes prior to network construction, through learning the patterns of 32 expression characteristics of known stress-related genes. The retained "informative" genes were subsequently analyzed by ML-based network comparison to predict candidate stress-related genes showing expression and network differences between control and stress networks, based on 33 network topological characteristics. Comparative evaluation of the network-centric and gene-centric analytic methods showed that mlDNA substantially outperformed traditional statistical testing-based differential expression analysis at identifying stress-related genes, with markedly improved prediction accuracy. To experimentally validate the mlDNA predictions, we selected 89 candidates out of the 1784 predicted salt stress-related genes with available SALK T-DNA mutagenesis lines for phenotypic screening and identified two previously unreported genes, mutants of which showed salt-sensitive phenotypes.

  14. Transcriptome Sequencing (RNA-seq) Analysis of the Effects of Metal Nanoparticle Exposure on the Transcriptome of Chlamydomonas reinhardtii

    PubMed Central

    Simon, Dana F.; Domingos, Rute F.; Hauser, Charles; Hutchins, Colin M.

    2013-01-01

    The widespread use of nanoparticles (NPs) raises concern over their potential toxicological effects in humans and ecosystems. Here we used transcriptome sequencing (RNA-seq) to evaluate the effects of exposure to four different metal-based NPs, nano-Ag (nAg), nano-TiO2 (nTiO2), nano-ZnO (nZnO), and CdTe/CdS quantum dots (QDs), in the eukaryotic green alga Chlamydomonas reinhardtii. The transcriptome was characterized before and after exposure to each NP type. Specific toxicological effects were inferred from the functions of genes whose transcripts either increased or decreased. Data analysis resulted in important differences and also similarities among the NPs. Elevated levels of transcripts of several marker genes for stress were observed, suggesting that only nZnO caused nonspecific global stress to the cells under environmentally relevant conditions. Genes with photosynthesis-related functions were decreased drastically during exposure to nTiO2 and slightly during exposures to the other NP types. This pattern suggests either toxicological effects in the chloroplast or effects that mimic a transition from low to high light. nAg exposure dramatically elevated the levels of transcripts encoding known or predicted components of the cell wall and the flagella, suggesting that it damages structures exposed to the external milieu. Exposures to nTiO2, nZnO, and QDs elevated the levels of transcripts encoding subunits of the proteasome, suggesting proteasome inhibition, a phenomenon believed to underlie the development and progression of several major diseases, including Alzheimer's disease, and used in chemotherapy against multiple myeloma. PMID:23728819

  15. Gene structure in the sea urchin Strongylocentrotus purpuratus based on transcriptome analysis.

    PubMed

    Tu, Qiang; Cameron, R Andrew; Worley, Kim C; Gibbs, Richard A; Davidson, Eric H

    2012-10-01

    A comprehensive transcriptome analysis has been performed on protein-coding RNAs of Strongylocentrotus purpuratus, including 10 different embryonic stages, six feeding larval and metamorphosed juvenile stages, and six adult tissues. In this study, we pooled the transcriptomes from all of these sources and focused on the insights they provide for gene structure in the genome of this recently sequenced model system. The genome had initially been annotated by use of computational gene model prediction algorithms. A large fraction of these predicted genes were recovered in the transcriptome when the reads were mapped to the genome and appropriately filtered and analyzed. However, in a manually curated subset, we discovered that more than half the computational gene model predictions were imperfect, containing errors such as missing exons, prediction of nonexistent exons, erroneous intron/exon boundaries, fusion of adjacent genes, and prediction of multiple genes from single genes. The transcriptome data have been used to provide a systematic upgrade of the gene model predictions throughout the genome, very greatly improving the research usability of the genomic sequence. We have constructed new public databases that incorporate information from the transcriptome analyses. The transcript-based gene model data were used to define average structural parameters for S. purpuratus protein-coding genes. In addition, we constructed a custom sea urchin gene ontology, and assigned about 7000 different annotated transcripts to 24 functional classes. Strong correlations became evident between given functional ontology classes and structural properties, including gene size, exon number, and exon and intron size.

  16. Annotation of the zebrafish genome through an integrated transcriptomic and proteomic analysis.

    PubMed

    Kelkar, Dhanashree S; Provost, Elayne; Chaerkady, Raghothama; Muthusamy, Babylakshmi; Manda, Srikanth S; Subbannayya, Tejaswini; Selvan, Lakshmi Dhevi N; Wang, Chieh-Huei; Datta, Keshava K; Woo, Sunghee; Dwivedi, Sutopa B; Renuse, Santosh; Getnet, Derese; Huang, Tai-Chung; Kim, Min-Sik; Pinto, Sneha M; Mitchell, Christopher J; Madugundu, Anil K; Kumar, Praveen; Sharma, Jyoti; Advani, Jayshree; Dey, Gourav; Balakrishnan, Lavanya; Syed, Nazia; Nanjappa, Vishalakshi; Subbannayya, Yashwanth; Goel, Renu; Prasad, T S Keshava; Bafna, Vineet; Sirdeshmukh, Ravi; Gowda, Harsha; Wang, Charles; Leach, Steven D; Pandey, Akhilesh

    2014-11-01

    Accurate annotation of protein-coding genes is one of the primary tasks upon the completion of whole genome sequencing of any organism. In this study, we used an integrated transcriptomic and proteomic strategy to validate and improve the existing zebrafish genome annotation. We undertook high-resolution mass-spectrometry-based proteomic profiling of 10 adult organs, whole adult fish body, and two developmental stages of zebrafish (SAT line), in addition to transcriptomic profiling of six organs. More than 7,000 proteins were identified from proteomic analyses, and ∼ 69,000 high-confidence transcripts were assembled from the RNA sequencing data. Approximately 15% of the transcripts mapped to intergenic regions, the majority of which are likely long non-coding RNAs. These high-quality transcriptomic and proteomic data were used to manually reannotate the zebrafish genome. We report the identification of 157 novel protein-coding genes. In addition, our data led to modification of existing gene structures including novel exons, changes in exon coordinates, changes in frame of translation, translation in annotated UTRs, and joining of genes. Finally, we discovered four instances of genome assembly errors that were supported by both proteomic and transcriptomic data. Our study shows how an integrative analysis of the transcriptome and the proteome can extend our understanding of even well-annotated genomes.

  17. Analysis of Pigeon (Columba) Ovary Transcriptomes to Identify Genes Involved in Blue Light Regulation.

    PubMed

    Wang, Ying; Ding, Jia-Tong; Yang, Hai-Ming; Yan, Zheng-Jie; Cao, Wei; Li, Yang-Bai

    2015-01-01

    Monochromatic light is widely applied to promote poultry reproductive performance, yet little is currently known regarding the mechanism by which light wavelengths affect pigeon reproduction. Recently, high-throughput sequencing technologies have been used to provide genomic information for solving this problem. In this study, we employed Illumina Hiseq 2000 to identify differentially expressed genes in ovary tissue from pigeons under blue and white light conditions and de novo transcriptome assembly to construct a comprehensive sequence database containing information on the mechanisms of follicle development. A total of 157,774 unigenes (mean length: 790 bp) were obtained by the Trinity program, and 35.83% of these unigenes were matched to genes in a non-redundant protein database. Gene description, gene ontology, and the clustering of orthologous group terms were performed to annotate the transcriptome assembly. Differentially expressed genes between blue and white light conditions included those related to oocyte maturation, hormone biosynthesis, and circadian rhythm. Furthermore, 17,574 SSRs and 533,887 potential SNPs were identified in this transcriptome assembly. This work is the first transcriptome analysis of the Columba ovary using Illumina technology, and the resulting transcriptome and differentially expressed gene data can facilitate further investigations into the molecular mechanism of the effect of blue light on follicle development and reproduction in pigeons and other bird species.

  18. Comparative transcriptome analysis of microsclerotia development in Nomuraea rileyi.

    PubMed

    Song, Zhangyong; Yin, Youping; Jiang, Shasha; Liu, Juanjuan; Chen, Huan; Wang, Zhongkang

    2013-06-19

    Nomuraea rileyi is used as an environmental-friendly biopesticide. However, mass production and commercialization of this organism are limited due to its fastidious growth and sporulation requirements. When cultured in amended medium, we found that N. rileyi could produce microsclerotia bodies, replacing conidiophores as the infectious agent. However, little is known about the genes involved in microsclerotia development. In the present study, the transcriptomes were analyzed using next-generation sequencing technology to find the genes involved in microsclerotia development. A total of 4.69 Gb of clean nucleotides comprising 32,061 sequences was obtained, and 20,919 sequences were annotated (about 65%). Among the annotated sequences, only 5928 were annotated with 34 gene ontology (GO) functional categories, and 12,778 sequences were mapped to 165 pathways by searching against the Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) database. Furthermore, we assessed the transcriptomic differences between cultures grown in minimal and amended medium. In total, 4808 sequences were found to be differentially expressed; 719 differentially expressed unigenes were assigned to 25 GO classes and 1888 differentially expressed unigenes were assigned to 161 KEGG pathways, including 25 enrichment pathways. Subsequently, we examined the up-regulation or uniquely expressed genes following amended medium treatment, which were also expressed on the enrichment pathway, and found that most of them participated in mediating oxidative stress homeostasis. To elucidate the role of oxidative stress in microsclerotia development, we analyzed the diversification of unigenes using quantitative reverse transcription-PCR (RT-qPCR). Our findings suggest that oxidative stress occurs during microsclerotia development, along with a broad metabolic activity change. Our data provide the most comprehensive sequence resource available for the study of N. rileyi. We believe that the transcriptome

  19. Comparative transcriptome analysis of microsclerotia development in Nomuraea rileyi

    PubMed Central

    2013-01-01

    Background Nomuraea rileyi is used as an environmental-friendly biopesticide. However, mass production and commercialization of this organism are limited due to its fastidious growth and sporulation requirements. When cultured in amended medium, we found that N. rileyi could produce microsclerotia bodies, replacing conidiophores as the infectious agent. However, little is known about the genes involved in microsclerotia development. In the present study, the transcriptomes were analyzed using next-generation sequencing technology to find the genes involved in microsclerotia development. Results A total of 4.69 Gb of clean nucleotides comprising 32,061 sequences was obtained, and 20,919 sequences were annotated (about 65%). Among the annotated sequences, only 5928 were annotated with 34 gene ontology (GO) functional categories, and 12,778 sequences were mapped to 165 pathways by searching against the Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) database. Furthermore, we assessed the transcriptomic differences between cultures grown in minimal and amended medium. In total, 4808 sequences were found to be differentially expressed; 719 differentially expressed unigenes were assigned to 25 GO classes and 1888 differentially expressed unigenes were assigned to 161 KEGG pathways, including 25 enrichment pathways. Subsequently, we examined the up-regulation or uniquely expressed genes following amended medium treatment, which were also expressed on the enrichment pathway, and found that most of them participated in mediating oxidative stress homeostasis. To elucidate the role of oxidative stress in microsclerotia development, we analyzed the diversification of unigenes using quantitative reverse transcription-PCR (RT-qPCR). Conclusion Our findings suggest that oxidative stress occurs during microsclerotia development, along with a broad metabolic activity change. Our data provide the most comprehensive sequence resource available for the study of N. rileyi. We

  20. De novo Assembly and Analysis of the Chilean Pencil Catfish Trichomycterus areolatus Transcriptome

    PubMed Central

    Schulze, Thomas T.; Ali, Jonathan M.; Bartlett, Maggie L.; McFarland, Madalyn M.; Clement, Emalie J.; Won, Harim I.; Sanford, Austin G.; Monzingo, Elyssa B.; Martens, Matthew C.; Hemsley, Ryan M.; Kumar, Sidharta; Gouin, Nicolas; Kolok, Alan S.; Davis, Paul H.

    2016-01-01

    Trichomycterus areolatus is an endemic species of pencil catfish that inhabits the riffles and rapids of many freshwater ecosystems of Chile. Despite its unique adaptation to Chile's high gradient watersheds and therefore potential application in the investigation of ecosystem integrity and environmental contamination, relatively little is known regarding the molecular biology of this environmental sentinel. Here, we detail the assembly of the Trichomycterus areolatus transcriptome, a molecular resource for the study of this organism and its molecular response to the environment. RNA-Seq reads were obtained by next-generation sequencing with an Illumina® platform and processed using PRINSEQ. The transcriptome assembly was performed using TRINITY assembler. Transcriptome validation was performed by functional characterization with KOG, KEGG, and GO analyses. Additionally, differential expression analysis highlights sex-specific expression patterns, and a list of endocrine and oxidative stress related transcripts are included. PMID:27672404

  1. Gestational tissue transcriptomics in term and preterm human pregnancies: a systematic review and meta-analysis.

    PubMed

    Eidem, Haley R; Ackerman, William E; McGary, Kriston L; Abbot, Patrick; Rokas, Antonis

    2015-06-05

    Preterm birth (PTB), or birth before 37 weeks of gestation, is the leading cause of newborn death worldwide. PTB is a critical area of scientific study not only due to its worldwide toll on human lives and economies, but also due to our limited understanding of its pathogenesis and, therefore, its prevention. This systematic review and meta-analysis synthesizes the landscape of PTB transcriptomics research to further our understanding of the genes and pathways involved in PTB subtypes. We evaluated published genome-wide pregnancy studies across gestational tissues and pathologies, including those that focus on PTB, by performing a targeted PubMed MeSH search and systematically reviewing all relevant studies. Our search yielded 2,361 studies on gestational tissues including placenta, decidua, myometrium, maternal blood, cervix, fetal membranes (chorion and amnion), umbilical cord, fetal blood, and basal plate. Selecting only those original research studies that measured transcription on a genome-wide scale and reported lists of expressed genetic elements identified 93 gene expression, 21 microRNA, and 20 methylation studies. Although 30 % of all PTB cases are due to medical indications, 76 % of the preterm studies focused on them. In contrast, only 18 % of the preterm studies focused on spontaneous onset of labor, which is responsible for 45 % of all PTB cases. Furthermore, only 23 of the 10,993 unique genetic elements reported to be transcriptionally active were recovered 10 or more times in these 134 studies. Meta-analysis of the 93 gene expression studies across 9 distinct gestational tissues and 29 clinical phenotypes showed limited overlap of genes identified as differentially expressed across studies. Overall, profiles of differentially expressed genes were highly heterogeneous both between as well as within clinical subtypes and tissues as well as between studies of the same clinical subtype and tissue. These results suggest that large gaps still exist in

  2. Transcriptome Analysis of Early Fruit Development in Three Seedy Citrus Genotypes and Their Seedless Mutants

    USDA-ARS?s Scientific Manuscript database

    Seedlessness is desirable for most citrus fruit, and identification of spontaneous or irradiated seedless mutants is important in developing citrus cultivars. We conducted a transcriptome analysis in early fruit development of three seedy citrus types (‘Fallglo’, a largely C. reticulata hybrid; ‘Pi...

  3. Comprehensive transcriptome analysis identifies pathways with therapeutic potential in locally advanced cervical cancer.

    PubMed

    Campos-Parra, Alma Delia; Padua-Bracho, Alejandra; Pedroza-Torres, Abraham; Figueroa-González, Gabriela; Fernández-Retana, Jorge; Millan-Catalan, Oliver; Peralta-Zaragoza, Oscar; Cantú de León, David; Herrera, Luis A; Pérez-Plasencia, Carlos

    2016-11-01

    The objective of the present study was to provide genomic and transcriptomic information that may improve clinical outcomes for locally advanced cervical cancer (LACC) patients by searching for therapeutic targets or potential biomarkers through the analysis of significantly altered signaling pathways in LACC. Microarray-based transcriptome profiling of 89 tumor samples from women with LACC was performed. Through Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, significantly over-expressed genes in LACC were identified; these genes were validated by quantitative reverse transcription-polymerase chain reaction in an independent cohort, and the protein expression data were obtained from the Human Protein Atlas. A transcriptome analysis revealed 7530 significantly over-expressed genes in LACC samples. By KEGG analysis, we found 93 dysregulated signaling pathways, including the JAK-STAT, NOTCH and mTOR-autophagy pathways, which were significantly upregulated. We confirmed the overexpression of the relevant genes of each pathway, such as NOTCH1, JAK2, STAM1, SOS1, ADAM17, PSEN1, NCSTN, RPS6, STK11/LKB1 and MLTS8/GBL in LACC compared with normal cervical tissue epithelia. Through comprehensive genomic and transcriptomic analyses, this work provides information regarding signaling pathways with promising therapeutic targets, suggesting novel target therapies to be considered in future clinical trials for LACC patients. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  4. A comprehensive analysis of the human placenta transcriptome.

    PubMed

    Saben, J; Zhong, Y; McKelvey, S; Dajani, N K; Andres, A; Badger, T M; Gomez-Acevedo, H; Shankar, K

    2014-02-01

    As the conduit for nutrients and growth signals, the placenta is critical to establishing an environment sufficient for fetal growth and development. To better understand the mechanisms regulating placental development and gene expression, we characterized the transcriptome of term placenta from 20 healthy women with uncomplicated pregnancies using RNA-seq. To identify genes that were highly expressed and unique to the placenta we compared placental RNA-seq data to data from 7 other tissues (adipose, breast, hear, kidney, liver, lung, and smooth muscle) and identified several genes novel to placental biology (QSOX1, DLG5, and SEMA7A). Semi-quantitative RT-PCR confirmed the RNA-seq results and immunohistochemistry indicated these proteins were highly expressed in the placental syncytium. Additionally, we mined our RNA-seq data to map the relative expression of key developmental gene families (Fox, Sox, Gata, Tead, and Wnt) within the placenta. We identified FOXO4, GATA3, and WNT7A to be amongst the highest expressed members of these families. Overall, these findings provide a new reference for understanding of placental transcriptome and can aid in the identification of novel pathways regulating placenta physiology that may be dysregulated in placental disease. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. A Comprehensive Analysis of the Human Placenta Transcriptome

    PubMed Central

    Saben, Jessica; Zhong, Ying; McKelvey, Samantha; Dajani, Nafisa K.; Andres, Aline; Badger, Thomas M.; Gomez-Acevedo, Horacio; Shankar, Kartik

    2014-01-01

    As the conduit for nutrients and growth signals, the placenta is critical to establishing an environment sufficient for fetal growth and development. To better understand the mechanisms regulating placental development and gene expression, we characterized the transcriptome of term placenta from 20 healthy women with uncomplicated pregnancies using RNA-seq. To identify genes that were highly expressed and unique to the placenta we compared placental RNA-seq data to data from 7 other tissues (adipose, breast, hear, kidney, liver, lung, and smooth muscle) and identified several genes novel to placental biology (QSOX1, DLG5, and SEMA7A). Semi-quantitative RT-PCR confirmed the RNA-seq results and immunohistochemistry indicated these proteins were highly expressed in the placental syncytium. Additionally, we mined our RNA-seq data to map the relative expression of key developmental gene families (Fox, Sox, Gata, Tead, and Wnt) within the placenta. We identified FOXO4, GATA3, and WNT7A to be amongst the highest expressed members of these families. Overall, these findings provide a new reference for understanding of placental transcriptome and can aid in the identification of novel pathways regulating placenta physiology that may be dysregulated in placental disease. PMID:24333048

  6. De Novo Assembly and Transcriptome Analysis of Contrasting Sugarcane Varieties

    PubMed Central

    Mancini, Melina Cristina; Balsalobre, Thiago Willian Almeida; Canesin, Lucas Eduardo Costa; Pinto, Luciana Rossini; Carneiro, Monalisa Sampaio; Garcia, Antonio Augusto Franco; de Souza, Anete Pereira; Vicentini, Renato

    2014-01-01

    Sugarcane is an important crop and a major source of sugar and alcohol. In this study, we performed de novo assembly and transcriptome annotation for six sugarcane genotypes involved in bi-parental crosses. The de novo assembly of the sugarcane transcriptome was performed using short reads generated using the Illumina RNA-Seq platform. We produced more than 400 million reads, which were assembled into 72,269 unigenes. Based on a similarity search, the unigenes showed significant similarity to more than 28,788 sorghum proteins, including a set of 5,272 unigenes that are not present in the public sugarcane EST databases; many of these unigenes are likely putative undescribed sugarcane genes. From this collection of unigenes, a large number of molecular markers were identified, including 5,106 simple sequence repeats (SSRs) and 708,125 single-nucleotide polymorphisms (SNPs). This new dataset will be a useful resource for future genetic and genomic studies in this species. PMID:24523899

  7. Transcriptome and Genome Size Analysis of the Venus Flytrap

    PubMed Central

    Bressendorff, Simon; Seguin-Orlando, Andaine; Petersen, Morten; Sicheritz-Pontén, Thomas; Mundy, John

    2015-01-01

    The insectivorous Venus flytrap (Dionaea muscipula) is renowned from Darwin’s studies of plant carnivory and the origins of species. To provide tools to analyze the evolution and functional genomics of D. muscipula, we sequenced a normalized cDNA library synthesized from mRNA isolated from D. muscipula flowers and traps. Using the Oases transcriptome assembler 79,165,657 quality trimmed reads were assembled into 80,806 cDNA contigs, with an average length of 679 bp and an N50 length of 1,051 bp. A total of 17,047 unique proteins were identified, and assigned to Gene Ontology (GO) and classified into functional categories. A total of 15,547 full-length cDNA sequences were identified, from which open reading frames were detected in 10,941. Comparative GO analyses revealed that D. muscipula is highly represented in molecular functions related to catalytic, antioxidant, and electron carrier activities. Also, using a single copy sequence PCR-based method, we estimated that the genome size of D. muscipula is approx. 3 Gb. Our genome size estimate and transcriptome analyses will contribute to future research on this fascinating, monotypic species and its heterotrophic adaptations. PMID:25886597

  8. Transcriptome and genome size analysis of the Venus flytrap.

    PubMed

    Jensen, Michael Krogh; Vogt, Josef Korbinian; Bressendorff, Simon; Seguin-Orlando, Andaine; Petersen, Morten; Sicheritz-Pontén, Thomas; Mundy, John

    2015-01-01

    The insectivorous Venus flytrap (Dionaea muscipula) is renowned from Darwin's studies of plant carnivory and the origins of species. To provide tools to analyze the evolution and functional genomics of D. muscipula, we sequenced a normalized cDNA library synthesized from mRNA isolated from D. muscipula flowers and traps. Using the Oases transcriptome assembler 79,165,657 quality trimmed reads were assembled into 80,806 cDNA contigs, with an average length of 679 bp and an N50 length of 1,051 bp. A total of 17,047 unique proteins were identified, and assigned to Gene Ontology (GO) and classified into functional categories. A total of 15,547 full-length cDNA sequences were identified, from which open reading frames were detected in 10,941. Comparative GO analyses revealed that D. muscipula is highly represented in molecular functions related to catalytic, antioxidant, and electron carrier activities. Also, using a single copy sequence PCR-based method, we estimated that the genome size of D. muscipula is approx. 3 Gb. Our genome size estimate and transcriptome analyses will contribute to future research on this fascinating, monotypic species and its heterotrophic adaptations.

  9. Transcriptome analysis in cotton boll weevil (Anthonomus grandis) and RNA interference in insect pests.

    PubMed

    Firmino, Alexandre Augusto Pereira; Fonseca, Fernando Campos de Assis; de Macedo, Leonardo Lima Pepino; Coelho, Roberta Ramos; Antonino de Souza, José Dijair; Togawa, Roberto Coiti; Silva-Junior, Orzenil Bonfim; Pappas-Jr, Georgios Joannis; da Silva, Maria Cristina Mattar; Engler, Gilbert; Grossi-de-Sa, Maria Fatima

    2013-01-01

    Cotton plants are subjected to the attack of several insect pests. In Brazil, the cotton boll weevil, Anthonomus grandis, is the most important cotton pest. The use of insecticidal proteins and gene silencing by interference RNA (RNAi) as techniques for insect control are promising strategies, which has been applied in the last few years. For this insect, there are not much available molecular information on databases. Using 454-pyrosequencing methodology, the transcriptome of all developmental stages of the insect pest, A. grandis, was analyzed. The A. grandis transcriptome analysis resulted in more than 500.000 reads and a data set of high quality 20,841 contigs. After sequence assembly and annotation, around 10,600 contigs had at least one BLAST hit against NCBI non-redundant protein database and 65.7% was similar to Tribolium castaneum sequences. A comparison of A. grandis, Drosophila melanogaster and Bombyx mori protein families' data showed higher similarity to dipteran than to lepidopteran sequences. Several contigs of genes encoding proteins involved in RNAi mechanism were found. PAZ Domains sequences extracted from the transcriptome showed high similarity and conservation for the most important functional and structural motifs when compared to PAZ Domains from 5 species. Two SID-like contigs were phylogenetically analyzed and grouped with T. castaneum SID-like proteins. No RdRP gene was found. A contig matching chitin synthase 1 was mined from the transcriptome. dsRNA microinjection of a chitin synthase gene to A. grandis female adults resulted in normal oviposition of unviable eggs and malformed alive larvae that were unable to develop in artificial diet. This is the first study that characterizes the transcriptome of the coleopteran, A. grandis. A new and representative transcriptome database for this insect pest is now available. All data support the state of the art of RNAi mechanism in insects.

  10. Transcriptome Analysis in Cotton Boll Weevil (Anthonomus grandis) and RNA Interference in Insect Pests

    PubMed Central

    Coelho, Roberta Ramos; Antonino de Souza Jr, José Dijair; Togawa, Roberto Coiti; Silva-Junior, Orzenil Bonfim; Pappas-Jr, Georgios Joannis; da Silva, Maria Cristina Mattar; Engler, Gilbert; Grossi-de-Sa, Maria Fatima

    2013-01-01

    Cotton plants are subjected to the attack of several insect pests. In Brazil, the cotton boll weevil, Anthonomus grandis, is the most important cotton pest. The use of insecticidal proteins and gene silencing by interference RNA (RNAi) as techniques for insect control are promising strategies, which has been applied in the last few years. For this insect, there are not much available molecular information on databases. Using 454-pyrosequencing methodology, the transcriptome of all developmental stages of the insect pest, A. grandis, was analyzed. The A. grandis transcriptome analysis resulted in more than 500.000 reads and a data set of high quality 20,841 contigs. After sequence assembly and annotation, around 10,600 contigs had at least one BLAST hit against NCBI non-redundant protein database and 65.7% was similar to Tribolium castaneum sequences. A comparison of A. grandis, Drosophila melanogaster and Bombyx mori protein families’ data showed higher similarity to dipteran than to lepidopteran sequences. Several contigs of genes encoding proteins involved in RNAi mechanism were found. PAZ Domains sequences extracted from the transcriptome showed high similarity and conservation for the most important functional and structural motifs when compared to PAZ Domains from 5 species. Two SID-like contigs were phylogenetically analyzed and grouped with T. castaneum SID-like proteins. No RdRP gene was found. A contig matching chitin synthase 1 was mined from the transcriptome. dsRNA microinjection of a chitin synthase gene to A. grandis female adults resulted in normal oviposition of unviable eggs and malformed alive larvae that were unable to develop in artificial diet. This is the first study that characterizes the transcriptome of the coleopteran, A. grandis. A new and representative transcriptome database for this insect pest is now available. All data support the state of the art of RNAi mechanism in insects. PMID:24386449

  11. Transcriptome analysis of Aedes aegypti transgenic mosquitoes with altered immunity.

    PubMed

    Zou, Zhen; Souza-Neto, Jayme; Xi, Zhiyong; Kokoza, Vladimir; Shin, Sang Woon; Dimopoulos, George; Raikhel, Alexander

    2011-11-01

    The mosquito immune system is involved in pathogen-elicited defense responses. The NF-κB factors REL1 and REL2 are downstream transcription activators of Toll and IMD immune pathways, respectively. We have used genome-wide microarray analyses to characterize fat-body-specific gene transcript repertoires activated by either REL1 or REL2 in two transgenic strains of the mosquito Aedes aegypti. Vitellogenin gene promoter was used in each transgenic strain to ectopically express either REL1 (REL1+) or REL2 (REL2+) in a sex, tissue, and stage specific manner. There was a significant change in the transcript abundance of 297 (79 up- and 218 down-regulated) and 299 (123 up- and 176 down-regulated) genes in fat bodies of REL1+ and REL2+, respectively. Over half of the induced genes had predicted functions in immunity, and a large group of these was co-regulated by REL1 and REL2. By generating a hybrid transgenic strain, which ectopically expresses both REL1 and REL2, we have shown a synergistic action of these NF-κB factors in activating immune genes. The REL1+ immune transcriptome showed a significant overlap with that of cactus (RNAi)-depleted mosquitoes (50%). In contrast, the REL2+ -regulated transcriptome differed from the relatively small group of gene transcripts regulated by RNAi depletion of a putative inhibitor of the IMD pathway, caspar (35 up- and 140 down-regulated), suggesting that caspar contributes to regulation of a subset of IMD-pathway controlled genes. Infections of the wild type Ae. aegypti with Plasmodium gallinaceum elicited the transcription of a distinct subset of immune genes (76 up- and 25 down-regulated) relative to that observed in REL1+ and REL2+ mosquitoes. Considerable overlap was observed between the fat body transcriptome of Plasmodium-infected mosquitoes and that of mosquitoes with transiently depleted PIAS, an inhibitor of the JAK-STAT pathway. PIAS gene silencing reduced Plasmodium proliferation in Ae. aegypti, indicating the

  12. Comparative transcriptome analysis between planarian Dugesia japonica and other platyhelminth species

    PubMed Central

    2012-01-01

    Background Planarians are considered to be among the extant animals close to one of the earliest groups of organisms that acquired a central nervous system (CNS) during evolution. Planarians have a bilobed brain with nine lateral branches from which a variety of external signals are projected into different portions of the main lobes. Various interneurons process different signals to regulate behavior and learning/memory. Furthermore, planarians have robust regenerative ability and are attracting attention as a new model organism for the study of regeneration. Here we conducted large-scale EST analysis of the head region of the planarian Dugesia japonica to construct a database of the head-region transcriptome, and then performed comparative analyses among related species. Results A total of 54,752 high-quality EST reads were obtained from a head library of the planarian Dugesia japonica, and 13,167 unigene sequences were produced by de novo assembly. A new method devised here revealed that proteins related to metabolism and defense mechanisms have high flexibility of amino-acid substitutions within the planarian family. Eight-two CNS-development genes were found in the planarian (cf. C. elegans 3; chicken 129). Comparative analysis revealed that 91% of the planarian CNS-development genes could be mapped onto the schistosome genome, but one-third of these shared genes were not expressed in the schistosome. Conclusions We constructed a database that is a useful resource for comparative planarian transcriptome studies. Analysis comparing homologous genes between two planarian species showed that the potential of genes is important for accumulation of amino-acid substitutions. The presence of many CNS-development genes in our database supports the notion that the planarian has a fundamental brain with regard to evolution and development at not only the morphological/functional, but also the genomic, level. In addition, our results indicate that the planarian CNS

  13. A transcriptome analysis of the Aedes aegypti vitellogenic fat body

    PubMed Central

    Feitosa, Fabiana M.; Calvo, Eric; Merino, Emilio F.; Durham, Alan M.; James, Anthony A.; de Bianchi, Antonio G.; Marinotti, Osvaldo; Capurro, Margareth L.

    2006-01-01

    Aedes (Stegomyia) aegypti is an important dengue vector in tropical and subtropical zones throughout the world. A transcriptome of Ae. aegypti vitellogenic fat bodies is described here. The fat body is a dynamic tissue that participates in multiple biochemical functions of intermediate metabolism. A total of 589 randomly selected cDNAs were assembled into 262 clusters based on their primary sequence similarities. The putative translated proteins were classified into categories based on their function in accordance with significant similarity using the BlastX at NCBI FTP site and Pfam (Bateman et al. 2000) and SMART (Schultz et al. 2000) databases. The characterization of transcripts expressed in the fat body of Ae. aegypti at 24 hours post blood meal provides a basic tool for understanding the processes occurring in this organ and could identify putative new genes whose promoters can be used to specifically express transgenes in the fat bodies of Ae. aegypti. PMID:19537968

  14. Transcriptome-scale homoeolog-specific transcript assemblies of bread wheat

    PubMed Central

    2012-01-01

    Background Bread wheat is one of the world’s most important food crops and considerable efforts have been made to develop genomic resources for this species. This includes an on-going project by the International Wheat Genome Sequencing Consortium to assemble its large and complex genome, which is hexaploid and contains three closely related ‘homoeologous’ copies for each chromosome. This multi-national effort avoids the complications polyploidy entails for correct assembly of the genome by sequencing flow-sorted chromosome arms one at a time. Here we report on an alternate approach, a direct homoeolog-specific assembly of the expressed portion of the genome, the transcriptome. Results After assessment of the ability of various assemblers to generate homoeolog-specific assemblies, we employed a two-stage assembly process to produce a high-quality assembly of the transcriptome of hexaploid wheat from Roche-454 and Illumina GAIIx paired-end sequence reads. The assembly process made use of a rapid partitioning of expressed sequences into homoeologous clusters, followed by a parallel high-fidelity assembly of each cluster on a 1150-processor compute cloud. We assessed assembly quality through comparison to known wheat gene sequences and found that in ca. 98.5% of cases the assembly was sufficiently accurate for homoeologous triplets to be cleanly separated into either two or three separate contigs. Comparison to publicly available transcript collections suggests that the assembly covers ~75-80% of the complete transcriptome. Conclusions This work therefore describes the first homoeolog-specific sequence assembly of the wheat transcriptome and provides a reference transcriptome for future wheat research. Furthermore, our assembly methodology is transferable to other polyploid organisms. PMID:22989011

  15. A Systematic Heritability Analysis of the Human Whole Blood Transcriptome

    PubMed Central

    Huan, Tianxiao; Liu, Chunyu; Joehanes, Roby; Zhang, Xiaoling; Chen, Brian H.; Johnson, Andrew D.; Yao, Chen; Courchesne, Paul; O'Donnell, Christopher J.; Munson, Peter J.; Levy, Daniel

    2015-01-01

    Genome-wide expression quantitative trait locus (eQTL) mapping may reveal common genetic variants regulating gene expression. In addition to mapping eQTLs, we systematically evaluated the heritability of the whole blood transcriptome in 5626 participants from the Framingham Heart Study. Of all gene expression measurements, about 40% exhibit evidence of being heritable (hgeneExp2>0, (p<0.05]), the average heritability was estimated to be 0.13, and 10% display hgeneExp2>0.2. In order to identify the role of eQTLs in promoting phenotype differences and disease susceptibility, we investigated the proportion of cis/trans eQTLs in different heritability categories and discovered that genes with higher heritability are more likely to have cis eQTLs that explain large proportions of variance in the expression of the corresponding genes. Single cis eQTLs explain 0.33–0.53 of variance in transcripts on average, whereas single trans eQTLs only explain 0.02–0.07. The top cis eQTLs tend to explain more variance in the corresponding gene when its hgeneExp2 is greater. Taking body mass index (BMI) as a case study, we cross-linked cis/trans eQTLs with both GWAS SNPs and differentially expressed genes for BMI. We discovered that BMI GWAS SNPs in 16p11.2 (e.g., rs7359397) are associated with several BMI differentially expressed genes in a cis manner (e.g. SULT1A1, SPNS1, and TUFM). These BMI signature genes explain a much larger proportion of variance in BMI than do the GWAS SNPs. Our results shed light the impact of eQTLs on the heritability of the human whole blood transcriptome and its relations to phenotype differences. PMID:25585846

  16. Comparative transcriptome analysis of flower heterosis in two soybean F1 hybrids by RNA-seq

    PubMed Central

    Zhang, Chunbao; Lin, Chunjing; Zhong, Xiaofang; Peng, Bao; Yan, Hao; Zhang, Jingyong; Zhang, Weilong; Wang, Pengnian; Ding, Xiaoyang; Zhang, Wei; Zhao, Limei

    2017-01-01

    Heterosis has been widely exploited as an approach to enhance crop traits during breeding. However, its underlying molecular genetic mechanisms remain unclear. Recent advances in RNA sequencing technology (RNA-seq) have provided an opportunity to conduct transcriptome profiling for heterosis studies. We used RNA-seq to analyze the flower transcriptomes of two F1 hybrid soybeans (HYBSOY-1 and HYBSOY-5) and their parents. More than 385 million high-quality reads were generated and aligned against the soybean reference genome. A total of 681 and 899 genes were identified as being differentially expressed between HYBSOY-1 and HYBSOY-5 and their parents, respectively. These differentially expressed genes (DEGs) were categorized into four major expression categories with 12 expression patterns. Furthermore, gene ontology (GO) term analysis showed that the DEGs were enriched in the categories metabolic process and catalytic activity, while Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis found that metabolic pathway and biosynthesis of secondary metabolites were enriched in the two F1 hybrids. Comparing the DEGs of the two F1 hybrids by GO term and KEGG pathway analyses identified 26 common DEGs that showed transgressive up-regulation, and which could be considered potential candidate genes for heterosis in soybean F1 hybrids. This identification of an extensive transcriptome dataset gives a comprehensive overview of the flower transcriptomes in two F1 hybrids, and provides useful information for soybean hybrid breeding. These findings lay the foundation for future studies on molecular mechanisms underlying soybean heterosis. PMID:28708857

  17. Differential transcriptome analysis supports Rhodnius montenegrensis and Rhodnius robustus (Hemiptera, Reduviidae, Triatominae) as distinct species.

    PubMed

    de Carvalho, Danila Blanco; Congrains, Carlos; Chahad-Ehlers, Samira; Pinotti, Heloisa; Brito, Reinaldo Alves de; da Rosa, João Aristeu

    2017-01-01

    Chagas disease is one of the main parasitic diseases found in Latin America and it is estimated that between six and seven million people are infected worldwide. Its etiologic agent, the protozoan Trypanosoma cruzi, is transmitted by triatomines, some of which from the genus Rhodnius. Twenty species are currently recognized in this genus, including some closely related species with low levels of morphological differentiation, such as Rhodnius montenegrensis and Rhodnius robustus. In order to investigate genetic differences between these two species, we generated large-scale RNA-sequencing data (consisting of four RNA-seq libraries) from the heads and salivary glands of males of R. montenegrensis and R. robustus. Transcriptome assemblies produced for each species resulted in 64,952 contigs for R. montenegrensis and 70,894 contigs for R. robustus, with N50 of approximately 2,100 for both species. SNP calling based on the more complete R. robustus assembly revealed 3,055 fixed interspecific differences and 216 transcripts with high levels of divergence which contained only fixed differences between the two species. A gene ontology enrichment analysis revealed that these highly differentiated transcripts were enriched for eight GO terms related to AP-2 adaptor complex, as well as other interesting genes that could be involved in their differentiation. The results show that R. montenegrensis and R. robustus have a substantial quantity of fixed interspecific polymorphisms, which suggests a high degree of genetic divergence between the two species and likely corroborates the species status of R. montenegrensis.

  18. Transcriptome analysis highlights the conserved difference between embryonic and postnatal-derived alveolar macrophages

    PubMed Central

    Gibbings, Sophie L.; Goyal, Rajni; Desch, A. Nicole; Leach, Sonia M.; Prabagar, Miglena; Atif, Shaikh M.; Bratton, Donna L.; Janssen, William

    2015-01-01

    Alveolar macrophages (AMs) reside on the luminal surfaces of the airways and alveoli where they maintain host defense and promote alveolar homeostasis by ingesting inhaled particulates and regulating inflammatory responses. Recent studies have demonstrated that AMs populate the lungs during embryogenesis and self-renew throughout life with minimal replacement by circulating monocytes, except under extreme conditions of depletion or radiation injury. Here we demonstrate that on a global scale, environment appears to dictate AM development and function. Indeed, transcriptome analysis of embryonic host-derived and postnatal donor-derived AMs coexisting within the same mouse demonstrated >98% correlation and overall functional analyses were similar. However, we also identified several genes whose expression was dictated by origin rather than environment. The most differentially expressed gene not altered by environment was Marco, a gene recently demonstrated to have enhancer activity in embryonic-derived but not postnatal-derived tissue macrophages. Overall, we show that under homeostatic conditions, the environment largely dictates the programming and function of AMs, whereas the expression of a small number of genes remains linked to the origin of the cell. PMID:26232173

  19. mRNA-Seq Analysis of the Pseudoperonospora cubensis Transcriptome During Cucumber (Cucumis sativus L.) Infection

    PubMed Central

    Hamilton, John P.; Vaillancourt, Brieanne; Buell, C. Robin; Day, Brad

    2012-01-01

    Pseudoperonospora cubensis, an oomycete, is the causal agent of cucurbit downy mildew, and is responsible for significant losses on cucurbit crops worldwide. While other oomycete plant pathogens have been extensively studied at the molecular level, Ps. cubensis and the molecular basis of its interaction with cucurbit hosts has not been well examined. Here, we present the first large-scale global gene expression analysis of Ps. cubensis infection of a susceptible Cucumis sativus cultivar, ‘Vlaspik’, and identification of genes with putative roles in infection, growth, and pathogenicity. Using high throughput whole transcriptome sequencing, we captured differential expression of 2383 Ps. cubensis genes in sporangia and at 1, 2, 3, 4, 6, and 8 days post-inoculation (dpi). Additionally, comparison of Ps. cubensis expression profiles with expression profiles from an infection time course of the oomycete pathogen Phytophthora infestans on Solanum tuberosum revealed similarities in expression patterns of 1,576–6,806 orthologous genes suggesting a substantial degree of overlap in molecular events in virulence between the biotrophic Ps. cubensis and the hemi-biotrophic P. infestans. Co-expression analyses identified distinct modules of Ps. cubensis genes that were representative of early, intermediate, and late infection stages. Collectively, these expression data have advanced our understanding of key molecular and genetic events in the virulence of Ps. cubensis and thus, provides a foundation for identifying mechanism(s) by which to engineer or effect resistance in the host. PMID:22545137

  20. Transcriptome Analysis and Gene Expression Profiling of Abortive and Developing Ovules during Fruit Development in Hazelnut

    PubMed Central

    Cheng, Yunqing; Liu, Jianfeng; Zhang, Huidi; Wang, Ju; Zhao, Yixin; Geng, Wanting

    2015-01-01

    Background A high ratio of blank fruit in hazelnut (Corylus heterophylla Fisch) is a very common phenomenon that causes serious yield losses in northeast China. The development of blank fruit in the Corylus genus is known to be associated with embryo abortion. However, little is known about the molecular mechanisms responsible for embryo abortion during the nut development stage. Genomic information for C. heterophylla Fisch is not available; therefore, data related to transcriptome and gene expression profiling of developing and abortive ovules are needed. Methodology/Principal Findings In this study, de novo transcriptome sequencing and RNA-seq analysis were conducted using short-read sequencing technology (Illumina HiSeq 2000). The results of the transcriptome assembly analysis revealed genetic information that was associated with the fruit development stage. Two digital gene expression libraries were constructed, one for a full (normally developing) ovule and one for an empty (abortive) ovule. Transcriptome sequencing and assembly results revealed 55,353 unigenes, including 18,751 clusters and 36,602 singletons. These results were annotated using the public databases NR, NT, Swiss-Prot, KEGG, COG, and GO. Using digital gene expression profiling, gene expression differences in developing and abortive ovules were identified. A total of 1,637 and 715 unigenes were significantly upregulated and downregulated, respectively, in abortive ovules, compared with developing ovules. Quantitative real-time polymerase chain reaction analysis was used in order to verify the differential expression of some genes. Conclusions/Significance The transcriptome and digital gene expression profiling data of normally developing and abortive ovules in hazelnut provide exhaustive information that will improve our understanding of the molecular mechanisms of abortive ovule formation in hazelnut. PMID:25836368

  1. Transcriptome-Wide Analysis Reveals the Role of PPARγ Controlling the Lipid Metabolism in Goat Mammary Epithelial Cells

    PubMed Central

    Zhao, Wangsheng; Zhang, Changhui

    2016-01-01

    To explore the large-scale effect of peroxisome proliferator-activated receptor γ (PPARG) in goat mammary epithelial cells (GMEC), an oligonucleotide microarray platform was used for transcriptome profiling in cells overexpressing PPARG and incubated with or without rosiglitazone (ROSI, a PPARγ agonist). A total of 1143 differentially expressed genes (DEG) due to treatment were detected. The Dynamic Impact Approach (DIA) analysis uncovered the most impacted and induced pathways “fatty acid elongation in mitochondria,” “glycosaminoglycan biosynthesis-keratan sulfate,” and “pentose phosphate pathway.” The data highlights the central role of PPARG in milk fatty acid metabolism via controlling fatty acid elongation, biosynthesis of unsaturated fatty acid, lipid formation, and lipid secretion; furthermore, its role related to carbohydrate metabolism promotes the production of intermediates required for milk fat synthesis. Analysis of upstream regulators indicated that PPARG participates in multiple physiological processes via controlling or cross talking with other key transcription factors such as PPARD and NR1H3 (also known as liver-X-receptor-α). This transcriptome-wide analysis represents the first attempt to better understand the biological relevance of PPARG expression in ruminant mammary cells. Overall, the data underscored the importance of PPARG in mammary lipid metabolism and transcription factor control. PMID:27818678

  2. Transcriptome Sequencing, De Novo Assembly and Differential Gene Expression Analysis of the Early Development of Acipenser baeri

    PubMed Central

    Song, Wei; Jiang, Keji; Zhang, Fengying; Lin, Yu; Ma, Lingbo

    2015-01-01

    The molecular mechanisms that drive the development of the endangered fossil fish species Acipenser baeri are difficult to study due to the lack of genomic data. Recent advances in sequencing technologies and the reducing cost of sequencing offer exclusive opportunities for exploring important molecular mechanisms underlying specific biological processes. This manuscript describes the large scale sequencing and analyses of mRNA from Acipenser baeri collected at five development time points using the Illumina Hiseq2000 platform. The sequencing reads were de novo assembled and clustered into 278167 unigenes, of which 57346 (20.62%) had 45837 known homologues proteins in Uniprot protein databases while 11509 proteins matched with at least one sequence of assembled unigenes. The remaining 79.38% of unigenes could stand for non-coding unigenes or unigenes specific to A. baeri. A number of 43062 unigenes were annotated into functional categories via Gene Ontology (GO) annotation whereas 29526 unigenes were associated with 329 pathways by mapping to KEGG database. Subsequently, 3479 differentially expressed genes were scanned within developmental stages and clustered into 50 gene expression profiles. Genes preferentially expressed at each stage were also identified. Through GO and KEGG pathway enrichment analysis, relevant physiological variations during the early development of A. baeri could be better cognized. Accordingly, the present study gives insights into the transcriptome profile of the early development of A. baeri, and the information contained in this large scale transcriptome will provide substantial references for A. baeri developmental biology and promote its aquaculture research. PMID:26359664

  3. Insights into Sexual Precocity of Female Oriental River Prawn Macrobrachium nipponense through Transcriptome Analysis

    PubMed Central

    Jiang, Hongxia; Li, Xilian; Sun, Yuhang; Hou, Fujun; Zhang, Yufei; Li, Fei; Gu, Zhimin; Liu, Xiaolin

    2016-01-01

    Background The oriental river prawn (Macrobrachium nipponense) is the most prevalent aquaculture species in China. The sexual precocity in this species has received considerable attention in recent years because more and more individuals matured at a small size, which devalues the commercial production. In this study, we developed deep-coverage transcriptomic sequencing data for the ovaries of sexually precocious and normal sexually mature M. nipponense using next-generation RNA sequencing technology and attempted to provide the first insight into the molecular regulatory mechanism of sexual precocity in this species. Results A total of 63,336 unigenes were produced from the ovarian cDNA libraries of sexually precocious and normal sexually mature M. nipponense using Illumina HiSeq 2500 platform. Through BLASTX searches against the NR, STRING, Pfam, Swissprot and KEGG databases, 15,134 unigenes were annotated, accounting for 23.89% of the total unigenes. 5,195 and 3,227 matched unigenes were categorized by GO and COG analysis respectively. 15,908 unigenes were consequently mapped into 332 KEGG pathways, and many reproduction-related pathways and genes were identified. Moreover, 26,008 SSRs were identified from 18,133 unigenes. 80,529 and 80,516 SNPs were yielded from ovarian libraries of sexually precocious and normal sexually mature prawn, respectively, and 29,851 potential SNPs between these two groups were also predicted. After comparing the ovarian libraries of sexually precocious and normal sexually mature prawn, 549 differentially expressed genes (DEGs) and 9 key DEGs that may be related to sexual precocity of M. nipponense were identified. 20 DEGs were selected for validation by quantitative real-time PCR (QPCR) and 19 DEGs show consistent expression between QPCR and RNAseq-based differential expression analysis datasets. Conclusion This is the first report on the large-scale RNA sequencing of ovaries of sexually precocious and normal sexually mature M

  4. 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

  5. Profile Analysis: Multidimensional Scaling Approach.

    ERIC Educational Resources Information Center

    Ding, Cody S.

    2001-01-01

    Outlines an exploratory multidimensional scaling-based approach to profile analysis called Profile Analysis via Multidimensional Scaling (PAMS) (M. Davison, 1994). The PAMS model has the advantages of being applied to samples of any size easily, classifying persons on a continuum, and using person profile index for further hypothesis studies, but…

  6. De novo characterization of a whitefly transcriptome and analysis of its gene expression during development.

    PubMed

    Wang, Xiao-Wei; Luan, Jun-Bo; Li, Jun-Min; Bao, Yan-Yuan; Zhang, Chuan-Xi; Liu, Shu-Sheng

    2010-06-24

    Whitefly (Bemisia tabaci) causes extensive crop damage throughout the world by feeding directly on plants and by vectoring hundreds of species of begomoviruses. Yet little is understood about its genes involved in development, insecticide resistance, host range plasticity and virus transmission. To facilitate research on whitefly, we present a method for de novo assembly of whitefly transcriptome using short read sequencing technology (Illumina). In a single run, we produced more than 43 million sequencing reads. These reads were assembled into 168,900 unique sequences (mean size = 266 bp) which represent more than 10-fold of all the whitefly sequences deposited in the GenBank (as of March 2010). Based on similarity search with known proteins, these analyses identified 27,290 sequences with a cut-off E-value above 10-5. Assembled sequences were annotated with gene descriptions, gene ontology and clusters of orthologous group terms. In addition, we investigated the transcriptome changes during whitefly development using a tag-based digital gene expression (DGE) system. We obtained a sequencing depth of over 2.5 million tags per sample and identified a large number of genes associated with specific developmental stages and insecticide resistance. Our data provides the most comprehensive sequence resource available for whitefly study and demonstrates that the Illumina sequencing allows de novo transcriptome assembly and gene expression analysis in a species lacking genome information. We anticipate that next generation sequencing technologies hold great potential for the study of the transcriptome in other non-model organisms.

  7. Transcriptomic and Proteomic Analysis of Arion vulgaris—Proteins for Probably Successful Survival Strategies?

    PubMed Central

    Bulat, Tanja; Smidak, Roman; Sialana, Fernando J.; Jung, Gangsoo; Rattei, Thomas; Bilban, Martin; Sattmann, Helmut; Lubec, Gert; Aradska, Jana

    2016-01-01

    The Spanish slug, Arion vulgaris, is considered one of the hundred most invasive species in Central Europe. The immense and very successful adaptation and spreading of A. vulgaris suggest that it developed highly effective mechanisms to deal with infections and natural predators. Current transcriptomic and proteomic studies on gastropods have been restricted mainly to marine and freshwater gastropods. No transcriptomic or proteomic study on A. vulgaris has been carried out so far, and in the current study, the first transcriptomic database from adult specimen of A. vulgaris is reported. To facilitate and enable proteomics in this non-model organism, a mRNA-derived protein database was constructed for protein identification. A gel-based proteomic approach was used to obtain the first generation of a comprehensive slug mantle proteome. A total of 2128 proteins were unambiguously identified; 48 proteins represent novel proteins with no significant homology in NCBI non-redundant database. Combined transcriptomic and proteomic analysis revealed an extensive repertoire of novel proteins with a role in innate immunity including many associated pattern recognition, effector proteins and cytokine-like proteins. The number and diversity in gene families encoding lectins point to a complex defense system, probably as a result of adaptation to a pathogen-rich environment. These results are providing a fundamental and important resource for subsequent studies on molluscs as well as for putative antimicrobial compounds for drug discovery and biomedical applications. PMID:26986963

  8. Transcriptomic and Proteomic Analysis of Arion vulgaris--Proteins for Probably Successful Survival Strategies?

    PubMed

    Bulat, Tanja; Smidak, Roman; Sialana, Fernando J; Jung, Gangsoo; Rattei, Thomas; Bilban, Martin; Sattmann, Helmut; Lubec, Gert; Aradska, Jana

    2016-01-01

    The Spanish slug, Arion vulgaris, is considered one of the hundred most invasive species in Central Europe. The immense and very successful adaptation and spreading of A. vulgaris suggest that it developed highly effective mechanisms to deal with infections and natural predators. Current transcriptomic and proteomic studies on gastropods have been restricted mainly to marine and freshwater gastropods. No transcriptomic or proteomic study on A. vulgaris has been carried out so far, and in the current study, the first transcriptomic database from adult specimen of A. vulgaris is reported. To facilitate and enable proteomics in this non-model organism, a mRNA-derived protein database was constructed for protein identification. A gel-based proteomic approach was used to obtain the first generation of a comprehensive slug mantle proteome. A total of 2128 proteins were unambiguously identified; 48 proteins represent novel proteins with no significant homology in NCBI non-redundant database. Combined transcriptomic and proteomic analysis revealed an extensive repertoire of novel proteins with a role in innate immunity including many associated pattern recognition, effector proteins and cytokine-like proteins. The number and diversity in gene families encoding lectins point to a complex defense system, probably as a result of adaptation to a pathogen-rich environment. These results are providing a fundamental and important resource for subsequent studies on molluscs as well as for putative antimicrobial compounds for drug discovery and biomedical applications.

  9. Genome and transcriptome analysis of surfactin biosynthesis in Bacillus amyloliquefaciens MT45

    PubMed Central

    Zhi, Yan; Wu, Qun; Xu, Yan

    2017-01-01

    Natural Bacillus isolates generate limited amounts of surfactin (<10% of their biomass), which functions as an antibiotic or signalling molecule in inter-/intra-specific interactions. However, overproduction of surfactin in Bacillus amyloliquefaciens MT45 was observed at a titre of 2.93 g/l, which is equivalent to half of the maximum biomass. To systemically unravel this efficient biosynthetic process, the genome and transcriptome of this bacterium were compared with those of B. amyloliquefaciens type strain DSM7T. MT45 possesses a smaller genome while containing more unique transporters and resistance-associated genes. Comparative transcriptome analysis revealed notable enrichment of the surfactin synthesis pathway in MT45, including central carbon metabolism and fatty acid biosynthesis to provide sufficient quantities of building precursors. Most importantly, the modular surfactin synthase overexpressed (9 to 49-fold) in MT45 compared to DSM7T suggested efficient surfactin assembly and resulted in the overproduction of surfactin. Furthermore, based on the expression trends observed in the transcriptome, there are multiple potential regulatory genes mediating the expression of surfactin synthase. Thus, the results of the present study provide new insights regarding the synthesis and regulation of surfactin in high-producing strain and enrich the genomic and transcriptomic resources available for B. amyloliquefaciens. PMID:28112210

  10. Transcriptomic and proteomic analysis of mouse radiation-induced acute myeloid leukaemia (AML)

    PubMed Central

    Badie, Christophe; Blachowicz, Agnieszka; Barjaktarovic, Zarko; Finnon, Rosemary; Michaux, Arlette; Sarioglu, Hakan; Brown, Natalie; Manning, Grainne; Benotmane, M. Abderrafi; Tapio, Soile; Polanska, Joanna; Bouffler, Simon D.

    2016-01-01

    A combined transcriptome and proteome analysis of mouse radiation-induced AMLs using two primary AMLs, cell lines from these primaries, another cell line and its in vivo passage is reported. Compared to haematopoietic progenitor and stem cells (HPSC), over 5000 transcriptome alterations were identified, 2600 present in all materials. 55 and 3 alterations were detected in the proteomes of the cell lines and primary/in vivo passage material respectively, with one common to all materials. In cell lines, approximately 50% of the transcriptome changes are related to adaptation to cell culture, and in the proteome this proportion was higher. An AML ‘signature’ of 17 genes/proteins commonly deregulated in primary AMLs and cell lines compared to HPSCs was identified and validated using human AML transcriptome data. This also distinguishes primary AMLs from cell lines and includes proteins such as Coronin 1, pontin/RUVBL1 and Myeloperoxidase commonly implicated in human AML. C-Myc was identified as having a key role in radiation leukaemogenesis. These data identify novel candidates relevant to mouse radiation AML pathogenesis, and confirm that pathways of leukaemogenesis in the mouse and human share substantial commonality. PMID:27250028

  11. Comparative Transcriptome Analysis Identifies Putative Genes Involved in the Biosynthesis of Xanthanolides in Xanthium strumarium L.

    PubMed Central

    Li, Yuanjun; Gou, Junbo; Chen, Fangfang; Li, Changfu; Zhang, Yansheng

    2016-01-01

    Xanthium strumarium L. is a traditional Chinese herb belonging to the Asteraceae family. The major bioactive components of this plant are sesquiterpene lactones (STLs), which include the xanthanolides. To date, the biogenesis of xanthanolides, especially their downstream pathway, remains largely unknown. In X. strumarium, xanthanolides primarily accumulate in its glandular trichomes. To identify putative gene candidates involved in the biosynthesis of xanthanolides, three X. strumarium transcriptomes, which were derived from the young leaves of two different cultivars and the purified glandular trichomes from one of the cultivars, were constructed in this study. In total, 157 million clean reads were generated and assembled into 91,861 unigenes, of which 59,858 unigenes were successfully annotated. All the genes coding for known enzymes in the upstream pathway to the biosynthesis of xanthanolides were present in the X. strumarium transcriptomes. From a comparative analysis of the X. strumarium transcriptomes, this study identified a number of gene candidates that are putatively involved in the downstream pathway to the synthesis of xanthanolides, such as four unigenes encoding CYP71 P450s, 50 unigenes for dehydrogenases, and 27 genes for acetyltransferases. The possible functions of these four CYP71 candidates are extensively discussed. In addition, 116 transcription factors that are highly expressed in X. strumarium glandular trichomes were also identified. Their possible regulatory roles in the biosynthesis of STLs are discussed. The global transcriptomic data for X. strumarium should provide a valuable resource for further research into the biosynthesis of xanthanolides. PMID:27625674

  12. Fish-T1K (Transcriptomes of 1,000 Fishes) Project: large-scale transcriptome data for fish evolution studies.

    PubMed

    Sun, Ying; Huang, Yu; Li, Xiaofeng; Baldwin, Carole C; Zhou, Zhuocheng; Yan, Zhixiang; Crandall, Keith A; Zhang, Yong; Zhao, Xiaomeng; Wang, Min; Wong, Alex; Fang, Chao; Zhang, Xinhui; Huang, Hai; Lopez, Jose V; Kilfoyle, Kirk; Zhang, Yong; Ortí, Guillermo; Venkatesh, Byrappa; Shi, Qiong

    2016-01-01

    Ray-finned fishes (Actinopterygii) represent more than 50 % of extant vertebrates and are of great evolutionary, ecologic and economic significance, but they are relatively underrepresented in 'omics studies. Increased availability of transcriptome data for these species will allow researchers to better understand changes in gene expression, and to carry out functional analyses. An international project known as the "Transcriptomes of 1,000 Fishes" (Fish-T1K) project has been established to generate RNA-seq transcriptome sequences for 1,000 diverse species of ray-finned fishes. The first phase of this project has produced transcriptomes from more than 180 ray-finned fishes, representing 142 species and covering 51 orders and 109 families. Here we provide an overview of the goals of this project and the work done so far.

  13. Use of prior knowledge for the analysis of high-throughput transcriptomics and metabolomics data

    PubMed Central

    2014-01-01

    Background High-throughput omics technologies have enabled the measurement of many genes or metabolites simultaneously. The resulting high dimensional experimental data poses significant challenges to transcriptomics and metabolomics data analysis methods, which may lead to spurious instead of biologically relevant results. One strategy to improve the results is the incorporation of prior biological knowledge in the analysis. This strategy is used to reduce the solution space and/or to focus the analysis on biological meaningful regions. In this article, we review a selection of these methods used in transcriptomics and metabolomics. We combine the reviewed methods in three groups based on the underlying mathematical model: exploratory methods, supervised methods and estimation of the covariance matrix. We discuss which prior knowledge has been used, how it is incorporated and how it modifies the mathematical properties of the underlying methods. PMID:25033193

  14. Computational analysis of conserved RNA secondary structure in transcriptomes and genomes

    PubMed Central

    Eddy, Sean R.

    2017-01-01

    Transcriptomics experiments and computational predictions both enable systematic discovery of new functional RNAs, but many putative noncoding transcripts arise instead from artifacts and biological noise, and current computational prediction methods have high false positive rates. I discuss prospects for improving computational methods for analyzing and identifying functional RNAs, with a focus on detecting signatures of conserved RNA secondary structure. An interesting new front is the application of chemical and enzymatic RNA structure probing experiments on a transcriptome-wide scale. I review several proposed approaches for incorporating structure probing data into computational RNA secondary structure prediction. Using probabilistic inference formalisms, I show how all these approaches can be unified in a well-principled framework. Using that framework, RNA probing data can easily be integrated into a wide range of different analyses that depend on RNA secondary structure inference, including homology search and genome-wide detection of new structural RNAs. PMID:24895857

  15. Transcriptome-scale similarities between mouse and human skeletal muscles with normal and myopathic phenotypes

    PubMed Central

    Kho, Alvin T; Kang, Peter B; Kohane, Isaac S; Kunkel, Louis M

    2006-01-01

    Background Mouse and human skeletal muscle transcriptome profiles vary by muscle type, raising the question of which mouse muscle groups have the greatest molecular similarities to human skeletal muscle. Methods Orthologous (whole, sub-) transcriptome profiles were compared among four mouse-human transcriptome datasets: (M) six muscle groups obtained from three mouse strains (wildtype, mdx, mdx5cv); (H1) biopsied human quadriceps from controls and Duchenne muscular dystrophy patients; (H2) four different control human muscle types obtained at autopsy; and (H3) 12 different control human tissues (ten non-muscle). Results Of the six mouse muscles examined, mouse soleus bore the greatest molecular similarities to human skeletal muscles, independent of the latters' anatomic location/muscle type, disease state, age and sampling method (autopsy versus biopsy). Significant similarity to any one mouse muscle group was not observed for non-muscle human tissues (dataset H3), indicating this finding to be muscle specific. Conclusion This observation may be partly explained by the higher type I fiber content of soleus relative to the other mouse muscles sampled. PMID:16522209

  16. Transcriptome analysis elucidates key developmental components of bryozoan lophophore development.

    PubMed

    Wong, Yue Him; Ryu, Taewoo; Seridi, Loqmane; Ghosheh, Yanal; Bougouffa, Salim; Qian, Pei-Yuan; Ravasi, Timothy

    2014-10-10

    The most recent phylogenomic study suggested that Bryozoa (Ectoprocta), Brachiopoda, and Phoronida are monophyletic, implying that the lophophore of bryozoans, phoronids and brachiopods is a synapomorphy. Understanding the molecular mechanisms of the lophophore development of the Lophophorata clade can therefore provide us a new insight into the formation of the diverse morphological traits in metazoans. In the present study, we profiled the transcriptome of the Bryozoan (Ectoproct) Bugula neritina during the swimming larval stage (SW) and the early (4 h) and late (24 h) metamorphic stages using the Illumina HiSeq2000 platform. Various genes that function in development, the immune response and neurogenesis showed differential expression levels during metamorphosis. In situ hybridization of 23 genes that participate in the Wnt, BMP, Notch, and Hedgehog signaling pathways revealed their regulatory roles in the development of the lophophore and the ancestrula digestive tract. Our findings support the hypothesis that developmental precursors of the lophophore and the ancestrula digestive tract are pre-patterned by the differential expression of key developmental genes according to their fate. This study provides a foundation to better understand the developmental divergence and/or convergence among developmental precursors of the lophophore of bryozoans, branchiopods and phoronids.

  17. Transcriptome analysis elucidates key developmental components of bryozoan lophophore development

    PubMed Central

    Wong, Yue Him; Ryu, Taewoo; Seridi, Loqmane; Ghosheh, Yanal; Bougouffa, Salim; Qian, Pei-Yuan; Ravasi, Timothy

    2014-01-01

    The most recent phylogenomic study suggested that Bryozoa (Ectoprocta), Brachiopoda, and Phoronida are monophyletic, implying that the lophophore of bryozoans, phoronids and brachiopods is a synapomorphy. Understanding the molecular mechanisms of the lophophore development of the Lophophorata clade can therefore provide us a new insight into the formation of the diverse morphological traits in metazoans. In the present study, we profiled the transcriptome of the Bryozoan (Ectoproct) Bugula neritina during the swimming larval stage (SW) and the early (4 h) and late (24 h) metamorphic stages using the Illumina HiSeq2000 platform. Various genes that function in development, the immune response and neurogenesis showed differential expression levels during metamorphosis. In situ hybridization of 23 genes that participate in the Wnt, BMP, Notch, and Hedgehog signaling pathways revealed their regulatory roles in the development of the lophophore and the ancestrula digestive tract. Our findings support the hypothesis that developmental precursors of the lophophore and the ancestrula digestive tract are pre-patterned by the differential expression of key developmental genes according to their fate. This study provides a foundation to better understand the developmental divergence and/or convergence among developmental precursors of the lophophore of bryozoans, branchiopods and phoronids. PMID:25300304

  18. Comparative transcriptomic analysis of Clostridium perfringens biofilms and planktonic cells.

    PubMed

    Charlebois, Audrey; Jacques, Mario; Archambault, Marie

    2016-10-01

    Clostridium perfringens is an opportunistic pathogen that can cause food poisoning in humans and various enterotoxaemias in animal species. Recently, C. perfringens was shown to form biofilms, a structured community of bacterial cells enclosed in a self-produced extracellular matrix. However, very little is known on the subject and no information is available on gene expression in C. perfringens biofilms. To gain insights into the differences between free-living C. perfringens cells and those in biofilms, we used RNA sequencing. In total, 25.7% of genes showed differential expression in the two growth modes; about 12.8% of genes were up-regulated and about 12.9% were down-regulated in biofilms. We show that 772 genes were significantly differentially expressed between biofilms and planktonic cells from the supernatant of biofilms. Genes that were down-regulated in biofilm cells, relative to planktonic cells, included those involved in virulence, energy production, amino acid, nucleotide and carbohydrate metabolism, and in translation and ribosomal structure. Genes up-regulated in biofilm cells were mainly involved in amino acid and carbohydrate metabolism, transcription, inorganic ion metabolism and in defence mechanisms. This study provides new insights into the transcriptomic response of C. perfringens during biofilm formation.

  19. Comparative transcriptomic analysis of human and Drosophila extracellular vesicles

    PubMed Central

    Lefebvre, Fabio Alexis; Benoit Bouvrette, Louis Philip; Perras, Lilyanne; Blanchet-Cohen, Alexis; Garnier, Delphine; Rak, Janusz; Lécuyer, Éric

    2016-01-01

    Extracellular vesicles (EVs) are membrane-enclosed nanoparticles containing specific repertoires of genetic material. In mammals, EVs can mediate the horizontal transfer of various cargos and signaling molecules, notably miRNA and mRNA species. Whether this form of intercellular communication prevails in other metazoans remains unclear. Here, we report the first parallel comparative morphologic and transcriptomic characterization of EVs from Drosophila and human cellular models. Electronic microscopy revealed that human and Drosophila cells release similar EVs with diameters ranging from 30 to 200 nm, which contain complex populations of transcripts. RNA-seq identified abundant ribosomal RNAs, related pseudogenes and retrotransposons in human and Drosophila EVs. Vault RNAs and Y RNAs abounded in human samples, whereas small nucleolar RNAs involved in pseudouridylation were most prevalent in Drosophila EVs. Numerous mRNAs were identified, largely consisting of exonic sequences displaying full-length read coverage and enriched for translation and electronic transport chain functions. By analogy with human systems, these sizeable similarities suggest that EVs could potentially enable RNA-mediated intercellular communication in Drosophila. PMID:27282340

  20. Transcriptome response analysis of Arabidopsis thaliana to leafminer (Liriomyza huidobrensis)

    PubMed Central

    2012-01-01

    Background Plants have evolved a complicated resistance system and exhibit a variety of defense patterns in response to different attackers. Previous studies have shown that responses of plants to chewing insects and phloem-feeding insects are significantly different. Less is known, however, regarding molecular responses to leafminer insects. To investigate plant transcriptome response to leafminers, we selected the leafminer Liriomyza huidobrensis, which has a special feeding pattern more similar to pathogen damage than that of chewing insects, as a model insect, and Arabidopsis thaliana as a response plant. Results We first investigated local and systemic responses of A. thaliana to leafminer feeding using an Affymetrix ATH1 genome array. Genes related to metabolic processes and stimulus responses were highly regulated. Most systemically-induced genes formed a subset of the local response genes. We then downloaded gene expression data from online databases and used hierarchical clustering to explore relationships among gene expression patterns in A. thaliana damaged by different attackers. Conclusions Our results demonstrate that plant response patterns are strongly coupled to damage patterns of attackers. PMID:23231622

  1. Integrated proteomic and transcriptomic analysis of the Aedes aegypti eggshell

    PubMed Central

    2014-01-01

    Background Mosquito eggshells show remarkable diversity in physical properties and structure consistent with adaptations to the wide variety of environments exploited by these insects. We applied proteomic, transcriptomic, and hybridization in situ techniques to identify gene products and pathways that participate in the assembly of the Aedes aegypti eggshell. Aedes aegypti population density is low during cold and dry seasons and increases immediately after rainfall. The survival of embryos through unfavorable periods is a key factor in the persistence of their populations. The work described here supports integrated vector control approaches that target eggshell formation and result in Ae. aegypti drought-intolerant phenotypes for public health initiatives directed to reduce mosquito-borne diseases. Results A total of 130 proteins were identified from the combined mass spectrometric analyses of eggshell preparations. Conclusions Classification of proteins according to their known and putative functions revealed the complexity of the eggshell structure. Three novel Ae. aegypti vitelline membrane proteins were discovered. Odorant-binding and cysteine-rich proteins that may be structural components of the eggshell were identified. Enzymes with peroxidase, laccase and phenoloxidase activities also were identified, and their likely involvements in cross-linking reactions that stabilize the eggshell structure are discussed. PMID:24707823

  2. Transcriptome analysis of human cumulus cells reveals hypoxia as the main determinant of follicular senescence

    PubMed Central

    Molinari, E.; Bar, H.; Pyle, A.M.; Patrizio, P.

    2016-01-01

    Study question Can RNA sequencing of human cumulus cells (CC) reveal molecular pathways involved in the physiology of reproductive aging? Study finding Senescent but not young CC activate gene pathways associated with hypoxia and oxidative stress. What is known already Shifts in socioeconomic norms are resulting in larger numbers of women postponing childbearing. The reproductive potential is sharply decreased with aging, and the reasons are poorly understood. Since CCs play an integral role in oocyte maturation and direct access to human oocytes is limited, we used whole transcriptome analysis of these somatic cells to gain insights into the molecular mechanisms playing a role in follicular senescence. Study design, samples/materials, methods Twenty CC samples (from a total of 15 patients) were obtained from oocytes of either male factor or egg donor patients. RNA sequencing and bioinformatic tools were used to identify differentially expressed genes between CCs from seven aged and eight young patients (<35 (years old) y.o. vs >40 y.o.). Quantitative-PCR and immunoflourescent staining were used for validation. Main results and the role of chance RNA sequencing identified 11 572 genes expressed in CC of both age cohorts, 45 of which were differentially expressed. In CC collected from patients >40 y.o., genes involved in the hypoxia stress response (NOS2, RORA and NR4A3), vasculature development (NR2F2, PTHLH), glycolysis (RALGAPA2 and TBC1D4) and cAMP turnover (PDE4D) were significantly overexpressed when compared with CC of patients younger than 35 y.o. Limitations, reasons for caution This study focused almost exclusively on assessing the genetic differences in CC transcriptome between young and older women. These genetic findings were not fully correlated with embryonic development and clinical outcome. Wider implications of the findings Our data provide a new hypothesis—follicular hypoxia—as the main mechanism leading to ovarian follicular senescence and

  3. Comprehensive analysis of RNA-seq data reveals the complexity of the transcriptome in Brassica rapa.

    PubMed

    Tong, Chaobo; Wang, Xiaowu; Yu, Jingyin; Wu, Jian; Li, Wanshun; Huang, Junyan; Dong, Caihua; Hua, Wei; Liu, Shengyi

    2013-10-07

    The species Brassica rapa (2n=20, AA) is an important vegetable and oilseed crop, and serves as an excellent model for genomic and evolutionary research in Brassica species. With the availability of whole genome sequence of B. rapa, it is essential to further determine the activity of all functional elements of the B. rapa genome and explore the transcriptome on a genome-wide scale. Here, RNA-seq data was employed to provide a genome-wide transcriptional landscape and characterization of the annotated and novel transcripts and alternative splicing events across tissues. RNA-seq reads were generated using the Illumina platform from six different tissues (root, stem, leaf, flower, silique and callus) of the B. rapa accession Chiifu-401-42, the same line used for whole genome sequencing. First, these data detected the widespread transcription of the B. rapa genome, leading to the identification of numerous novel transcripts and definition of 5'/3' UTRs of known genes. Second, 78.8% of the total annotated genes were detected as expressed and 45.8% were constitutively expressed across all tissues. We further defined several groups of genes: housekeeping genes, tissue-specific expressed genes and co-expressed genes across tissues, which will serve as a valuable repository for future crop functional genomics research. Third, alternative splicing (AS) is estimated to occur in more than 29.4% of intron-containing B. rapa genes, and 65% of them were commonly detected in more than two tissues. Interestingly, genes with high rate of AS were over-represented in GO categories relating to transcriptional regulation and signal transduction, suggesting potential importance of AS for playing regulatory role in these genes. Further, we observed that intron retention (IR) is predominant in the AS events and seems to preferentially occurred in genes with short introns. The high-resolution RNA-seq analysis provides a global transcriptional landscape as a complement to the B. rapa genome

  4. Transcriptomic analysis of mononuclear phagocyte differentiation and activation.

    PubMed

    Hume, David A; Freeman, Tom C

    2014-11-01

    Monocytes and macrophages differentiate from progenitor cells under the influence of colony-stimulating factors. Genome-scale data have enabled the identification of the sets of genes that are associated with specific functions and the mechanisms by which thousands of genes are regulated in response to pathogen challenge. In large datasets, it is possible to identify large sets of genes that are coregulated with the transcription factors that regulate them. They include macrophage-specific genes, interferon-responsive genes, early inflammatory genes, and those associated with endocytosis. Such analyses can also extract macrophage-associated signatures from large cancer tissue datasets. However, cluster analysis provides no support for a signature that distinguishes macrophages from antigen-presenting dendritic cells, nor the classification of macrophage activation states as classical versus alternative, or M1 versus M2. Although there has been a focus on a small subset of lineage-enriched transcription factors, such as PU.1, more than half of the transcription factors in the genome can be expressed in macrophage lineage cells under some state of activation, and they interact in a complex network. The network architecture is conserved across species, but many of the target genes evolve rapidly and differ between mouse and human. The data and publication deluge related to macrophage biology require the development of new analytical tools and ways of presenting information in an accessible form.

  5. Transcriptome Analysis of Differentially Expressed Genes Relevant to Variegation in Peach Flowers

    PubMed Central

    Yu, Faxin; Li, Shuxian; Yin, Tongming

    2014-01-01

    Background Variegation in flower color is commonly observed in many plant species and also occurs on ornamental peaches (Prunus persica f. versicolor [Sieb.] Voss). Variegated plants are highly valuable in the floricultural market. To gain a global perspective on genes differentially expressed in variegated peach flowers, we performed large-scale transcriptome sequencing of white and red petals separately collected from a variegated peach tree. Results A total of 1,556,597 high-quality reads were obtained, with an average read length of 445 bp. The ESTs were assembled into 16,530 contigs and 42,050 singletons. The resulting unigenes covered about 60% of total predicted genes in the peach genome. These unigenes were further subjected to functional annotation and biochemical pathway analysis. Digital expression analysis identified a total of 514 genes differentially expressed between red and white flower petals. Since peach flower coloration is determined by the expression and regulation of structural genes relevant to flavonoid biosynthesis, a detailed examination detected four key structural genes, including C4H, CHS, CHI and F3H, expressed at a significantly higher level in red than in white petal. Except for the structural genes, we also detected 11 differentially expressed regulatory genes relating to flavonoid biosynthesis. Using the differentially expressed structural genes as the test objects, we validated the digital expression results by using quantitative real-time PCR, and the differential expression of C4H, CHS and F3H were confirmed. Conclusion In this study, we generated a large EST collection from flower petals of a variegated peach. By digital expression analysis, we identified an informative list of candidate genes associated with variegation in peach flowers, which offered a unique opportunity to uncover the genetic mechanisms underlying flower color variegation. PMID:24603808

  6. De novo assembly and analysis of the transcriptome of Ocimum americanum var. pilosum under cold stress.

    PubMed

    Zhan, Xiangqiang; Yang, Lan; Wang, Dong; Zhu, Jian Kang; Lang, Zhaobo

    2016-03-09

    Ocimum americanum var. pilosum is a chilling-sensitive, widely distributed plant that is consumed as a vegetable in central and southern China. To increase our understanding of cold stress responses in this species, we performed de novo transcriptome assembly for O. americanum var. pilosum and compared the transcriptomes of plants grown under normal and low temperatures. A total of 115,022,842 high quality, clean reads were obtained from four libraries (two replicates of control samples and two replicates of chilling-treated samples) and were used to perform de novo transcriptome assembly. After isoforms were considered, 42,816 unigenes were generated, 30,748 of which were similar to known proteins as determined by a BLASTx search (E-value < =1.0E-05) against NCBI non-redundant, Swiss-Prot, Gene Ontology, KEGG, and Cluster of COG databases. Comparative analysis of transcriptomes revealed that 5179 unigenes were differentially expressed (with at least 2-fold changes, FDR < 0.01) in chilling-treated samples, and that 2344 and 2835 unigenes were up- and down-regulated by chilling stress, respectively. Expression of the 10 most up-regulated and the five most down-regulated unigenes was validated by qRT-PCR. To increase our understanding of these differentially expressed unigenes, we performed Gene ontology and KEGG pathway enrichment analyses. The CBF-mediated transcriptional cascade, a well-known cold tolerance pathway, was reconstructed using our de novo assembled transcriptome. Our study has generated a genome-wide transcript profile of O. americanum var. pilosum and a de novo assembled transcriptome, which can be used to characterize genes related to diverse biological processes. This is the first study to assess the cold-responsive transcriptome in an Ocimum species. Our results suggest that cold temperature significantly affects genes related to protein translation and cellular metabolism in this chilling sensitive species. Although most of the CBF

  7. 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

  8. Comprehensive analysis of the Corynebacterium glutamicum transcriptome using an improved RNAseq technique.

    PubMed

    Pfeifer-Sancar, Katharina; Mentz, Almut; Rückert, Christian; Kalinowski, Jörn

    2013-12-17

    The use of RNAseq to resolve the transcriptional organization of an organism was established in recent years and also showed the complexity and dynamics of bacterial transcriptomes. The aim of this study was to comprehensively investigate the transcriptome of the industrially relevant amino acid producer and model organism Corynebacterium glutamicum by RNAseq in order to improve its genome annotation and to describe important features for transcription and translation. RNAseq data sets were obtained by two methods, one that focuses on 5'-ends of primary transcripts and another that provides the overall transcriptome with an improved resolution of 3'-ends of transcripts. Subsequent data analysis led to the identification of more than 2,000 transcription start sites (TSSs), the definition of 5'-UTRs (untranslated regions) for annotated protein-coding genes, operon structures and many novel transcripts located between or in antisense orientation to protein-coding regions. Interestingly, a high number of mRNAs (33%) is transcribed as leaderless transcripts. From the data, consensus promoter and ribosome binding site (RBS) motifs were identified and it was shown that the majority of genes in C. glutamicum are transcribed monocistronically, but operons containing up to 16 genes are also present. The comprehensive transcriptome map of C. glutamicum established in this study represents a major step forward towards a complete definition of genetic elements (e.g. promoter regions, gene starts and stops, 5'-UTRs, RBSs, transcript starts and ends) and provides the ideal basis for further analyses on transcriptional regulatory networks in this organism. The methods developed are easily applicable for other bacteria and have the potential to be used also for quantification of transcriptomes, replacing microarrays in the near future.

  9. Comprehensive analysis of the Corynebacterium glutamicum transcriptome using an improved RNAseq technique

    PubMed Central

    2013-01-01

    Background The use of RNAseq to resolve the transcriptional organization of an organism was established in recent years and also showed the complexity and dynamics of bacterial transcriptomes. The aim of this study was to comprehensively investigate the transcriptome of the industrially relevant amino acid producer and model organism Corynebacterium glutamicum by RNAseq in order to improve its genome annotation and to describe important features for transcription and translation. Results RNAseq data sets were obtained by two methods, one that focuses on 5′-ends of primary transcripts and another that provides the overall transcriptome with an improved resolution of 3′-ends of transcripts. Subsequent data analysis led to the identification of more than 2,000 transcription start sites (TSSs), the definition of 5′-UTRs (untranslated regions) for annotated protein-coding genes, operon structures and many novel transcripts located between or in antisense orientation to protein-coding regions. Interestingly, a high number of mRNAs (33%) is transcribed as leaderless transcripts. From the data, consensus promoter and ribosome binding site (RBS) motifs were identified and it was shown that the majority of genes in C. glutamicum are transcribed monocistronically, but operons containing up to 16 genes are also present. Conclusions The comprehensive transcriptome map of C. glutamicum established in this study represents a major step forward towards a complete definition of genetic elements (e.g. promoter regions, gene starts and stops, 5′-UTRs, RBSs, transcript starts and ends) and provides the ideal basis for further analyses on transcriptional regulatory networks in this organism. The methods developed are easily applicable for other bacteria and have the potential to be used also for quantification of transcriptomes, replacing microarrays in the near future. PMID:24341750

  10. Sequencing and analysis of the transcriptome of the acorn worm Ptychodera flava, an indirect developing hemichordate.

    PubMed

    Chen, Shu-Hwa; Li, Kun-Lin; Lu, I-Hsuan; Wang, Yu-Bin; Tung, Che-Huang; Ting, Hsiu-Chi; Lin, Ching-Yi; Lin, Chung-Yen; Su, Yi-Hsien; Yu, Jr-Kai

    2014-06-01

    Hemichordates are the sister group of echinoderms, and together they are closely related to chordates within the deuterostome lineage. Therefore, hemichordates represent an important animal group for the understanding of both the evolution of developmental mechanisms in deuterostome animals and the origin of chordates. Recently, the majority of studies investigating hemichordates have focused on the direct-developing enteropneust hemichordate Saccoglossus kowalevskii; few have focused on the indirect-developing hemichordates, partly because of the lack of extensive genomic resources in these animals. In this study, we report the sequencing and analysis of a transcriptome from an indirect-developing enteropneust hemichordate Ptychodera flava. We sequenced a mixed cDNA library from six developmental stages using the Roche GS FLX Titanium System to generate more than 879,000 reads. These reads were assembled into 17,990 contigs with an average length of 1316bp. We found that 60% of the assembled contigs, along with 28% of the unassembled singleton reads, had significant hits to sequences in the NCBI database by a BLASTx search, and we also annotated these sequences and obtained Gene Ontology (GO) terms for 6744 contigs and 5802 singletons. We further identified candidate P. flava transcripts corresponding to genes involved in major developmental signaling pathways, including the Wnt, Notch and TGF-β signaling pathways. Using available genome/transcriptome datasets from the direct-developing hemichordate S. kowalevskii, the echinoderm Strongylocentrotus purpuratus and the chordate Branchiostoma floridae, we found that 90%, 80% and 73% of the annotated protein sequences in these respective species matched our P. flava transcriptome in a homology search. We also constructed a database for the P. flava transcriptome, and researchers can easily access this dataset online. Our dataset significantly increases the amount of available P. flava sequence data and can serve as a

  11. Robust-LongSAGE (RL-SAGE): an improved LongSAGE method for high-throughput transcriptome analysis.

    PubMed

    Gowda, Malali; Wang, Guo-Liang

    2008-01-01

    Serial analysis of gene expression (SAGE) is a powerful technique for large-scale transcriptome analysis in eukaryotes. However, technical difficulties in the SAGE library construction, such as low concatemer cloning efficiency, small concatemer size, and a high level of empty clones, has prohibited its widespread use as a routine technique for expression profiling in many laboratories. We recently improved the LongSAGE library construction method considerably and developed a modified version called Robust-LongSAGE, or RL-SAGE. In RL-SAGE, concatemer cloning efficiency and clone insert size were increased significantly. About 20 PCR reactions are sufficient to make a library with more than 150,000 clones. Using RL-SAGE, we have made 10 libraries of rice, maize, and the rice blast fungus Magnaporthe grisea.

  12. Transcriptome analysis of genes involved in secondary cell wall biosynthesis in developing internodes of Miscanthus lutarioriparius.

    PubMed

    Hu, Ruibo; Xu, Yan; Yu, Changjiang; He, Kang; Tang, Qi; Jia, Chunlin; He, Guo; Wang, Xiaoyu; Kong, Yingzhen; Zhou, Gongke

    2017-08-22

    Miscanthus is a promising lignocellulosic bioenergy crop for bioethanol production. To identify candidate genes and regulation networks involved in secondary cell wall (SCW) development in Miscanthus, we performed de novo transcriptome analysis of a developing internode. According to the histological and in-situ histochemical analysis, an elongating internode of M. lutarioriparius can be divided into three distinct segments, the upper internode (UI), middle internode (MI) and basal internode (BI), each representing a different stage of SCW development. The transcriptome analysis generated approximately 300 million clean reads, which were de novo assembled into 79,705 unigenes. Nearly 65% of unigenes was annotated in seven public databases. Comparative profiling among the UI, MI and BI revealed four distinct clusters. Moreover, detailed expression profiling was analyzed for gene families and transcription factors (TFs) involved in SCW biosynthesis, assembly and modification. Based on the co-expression patterns, putative regulatory networks between TFs and SCW-associated genes were constructed. The work provided the first transcriptome analysis of SCW development in M. lutarioriparius. The results obtained provide novel insights into the biosynthesis and regulation of SCW in Miscanthus. In addition, the genes identified represent good candidates for further functional studies to unravel their roles in SCW biosynthesis and modification.

  13. Visual Analysis of Transcriptome Data in the Context of Anatomical Structures and Biological Networks

    PubMed Central

    Junker, Astrid; Rohn, Hendrik; Schreiber, Falk

    2012-01-01

    The complexity and temporal as well as spatial resolution of transcriptome datasets is constantly increasing due to extensive technological developments. Here we present methods for advanced visualization and intuitive exploration of transcriptomics data as necessary prerequisites in order to facilitate the gain of biological knowledge. Color-coding of structural images based on the expression level enables a fast visual data analysis in the background of the examined biological system. The network-based exploration of these visualizations allows for comparative analysis of genes with specific transcript patterns and supports the extraction of functional relationships even from large datasets. In order to illustrate the presented methods, the tool HIVE was applied for visualization and exploration of database-retrieved expression data for master regulators of Arabidopsis thaliana flower and seed development in the context of corresponding tissue-specific regulatory networks. PMID:23162564

  14. Comprehensive analysis of the venom gland transcriptome of the spider Dolomedes fimbriatus

    PubMed Central

    Kozlov, Sergey A.; Lazarev, Vassili N.; Kostryukova, Elena S.; Selezneva, Oksana V.; Ospanova, Elena A.; Alexeev, Dmitry G.; Govorun, Vadim M.; Grishin, Eugene V.

    2014-01-01

    A comprehensive transcriptome analysis of an expressed sequence tag (EST) database of the spider Dolomedes fimbriatus venom glands using single-residue distribution analysis (SRDA) identified 7,169 unique sequences. Mature chains of 163 different toxin-like polypeptides were predicted on the basis of well-established methodology. The number of protein precursors of these polypeptides was appreciably numerous than the number of mature polypeptides. A total of 451 different polypeptide precursors, translated from 795 unique nucleotide sequences, were deduced. A homology search divided the 163 mature polypeptide sequences into 16 superfamilies and 19 singletons. The number of mature toxins in a superfamily ranged from 2 to 49, whereas the diversity of the original nucleotide sequences was greater (2–261 variants). We observed a predominance of inhibitor cysteine knot toxin-like polypeptides among the cysteine-containing structures in the analyzed transcriptome bank. Uncommon spatial folds were also found. PMID:25977780

  15. 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.

  16. Transcriptome analysis of scorpion species belonging to the Vaejovis genus.

    PubMed

    Quintero-Hernández, Verónica; Ramírez-Carreto, Santos; Romero-Gutiérrez, María Teresa; Valdez-Velázquez, Laura L; Becerril, Baltazar; Possani, Lourival D; Ortiz, Ernesto

    2015-01-01

    Scorpions belonging to the Buthidae family have traditionally drawn much of the biochemist's attention due to the strong toxicity of their venoms. Scorpions not toxic to mammals, however, also have complex venoms. They have been shown to be an important source of bioactive peptides, some of them identified as potential drug candidates for the treatment of several emerging diseases and conditions. It is therefore important to characterize the large diversity of components found in the non-Buthidae venoms. As a contribution to this goal, this manuscript reports the construction and characterization of cDNA libraries from four scorpion species belonging to the Vaejovis genus of the Vaejovidae family: Vaejovis mexicanus, V. intrepidus, V. subcristatus and V. punctatus. Some sequences coding for channel-acting toxins were found, as expected, but the main transcribed genes in the glands actively producing venom were those coding for non disulfide-bridged peptides. The ESTs coding for putative channel-acting toxins, corresponded to sodium channel β toxins, to members of the potassium channel-acting α or κ families, and to calcium channel-acting toxins of the calcin family. Transcripts for scorpine-like peptides of two different lengths were found, with some of the species coding for the two kinds. One sequence coding for La1-like peptides, of yet unknown function, was found for each species. Finally, the most abundant transcripts corresponded to peptides belonging to the long chain multifunctional NDBP-2 family and to the short antimicrobials of the NDBP-4 family. This apparent venom composition is in correspondence with the data obtained to date for other non-Buthidae species. Our study constitutes the first approach to the characterization of the venom gland transcriptome for scorpion species belonging to the Vaejovidae family.

  17. A preliminary transcriptomic analysis of lichen Dirinaria sp.

    NASA Astrophysics Data System (ADS)

    Nurhani, A. R. Siti; Munir, A. M. Abdul; Wahid, S. Mohd; Diba, A. B. Farah

    2013-11-01

    Lichen is a slow-growing symbiotic organism that consists of a fungus and a photobiont, comprising either an algae or a cyanobacterium living together in a single composite body, known as a thallus. Lichens have a remarkable ability to survive in extreme environmental conditions on earth that makes them a great biological indicator of air quality. The primary goal of this study is to discover the genes that may unravel the mechanism behind the tolerance of this lichen towards air pollution. Lichen samples of Dirinaria sp. were collected from two sites - Jerantut (J) as having a relatively good air quality and Klang (K), an area of bad air quality. Total RNA extraction was carried out, followed by sample preparation prior to transcriptomic sequencing. Altogether 21.7 million and 30.5 million high quality sequence reads from samples J and K, respectively were de novo assembled into 106884 and 88116 transcripts. The assembled sequences were annotated by BLASTX comparison against a non-redundant protein sequence database with 59403 sequences (67.4%) of sample K and 68972 sequences (64.5%) of sample J had a match in the database with a cut-off value of 1e-06. A total of 42175 sequences (47.8%) of sample K and 25648 sequences (24%) of sample J had a Gene Ontology term match. The sequences were assigned to Kyoto Encyclopedia of Genes and Genome (KEGG) pathways, resulting in 129 KEGG pathways generated from sample K, whilst 123 KEGG pathways were produced from sample J.

  18. Transcriptome Analysis of Scorpion Species Belonging to the Vaejovis Genus

    PubMed Central

    Quintero-Hernández, Verónica; Ramírez-Carreto, Santos; Romero-Gutiérrez, María Teresa; Valdez-Velázquez, Laura L.; Becerril, Baltazar; Possani, Lourival D.; Ortiz, Ernesto

    2015-01-01

    Scorpions belonging to the Buthidae family have traditionally drawn much of the biochemist’s attention due to the strong toxicity of their venoms. Scorpions not toxic to mammals, however, also have complex venoms. They have been shown to be an important source of bioactive peptides, some of them identified as potential drug candidates for the treatment of several emerging diseases and conditions. It is therefore important to characterize the large diversity of components found in the non-Buthidae venoms. As a contribution to this goal, this manuscript reports the construction and characterization of cDNA libraries from four scorpion species belonging to the Vaejovis genus of the Vaejovidae family: Vaejovis mexicanus, V. intrepidus, V. subcristatus and V. punctatus. Some sequences coding for channel-acting toxins were found, as expected, but the main transcribed genes in the glands actively producing venom were those coding for non disulfide-bridged peptides. The ESTs coding for putative channel-acting toxins, corresponded to sodium channel β toxins, to members of the potassium channel-acting α or κ families, and to calcium channel-acting toxins of the calcin family. Transcripts for scorpine-like peptides of two different lengths were found, with some of the species coding for the two kinds. One sequence coding for La1-like peptides, of yet unknown function, was found for each species. Finally, the most abundant transcripts corresponded to peptides belonging to the long chain multifunctional NDBP-2 family and to the short antimicrobials of the NDBP-4 family. This apparent venom composition is in correspondence with the data obtained to date for other non-Buthidae species. Our study constitutes the first approach to the characterization of the venom gland transcriptome for scorpion species belonging to the Vaejovidae family. PMID:25659089

  19. Genome-wide transcriptome and expression profile analysis of Phalaenopsis during explant browning.

    PubMed

    Xu, Chuanjun; Zeng, Biyu; Huang, Junmei; Huang, Wen; Liu, Yumei

    2015-01-01

    Explant browning presents a major problem for in vitro culture, and can lead to the death of the explant and failure of regeneration. Considerable work has examined the physiological mechanisms underlying Phalaenopsis leaf explant browning, but the molecular mechanisms of browning remain elusive. In this study, we used whole genome RNA sequencing to examine Phalaenopsis leaf explant browning at genome-wide level. We first used Illumina high-throughput technology to sequence the transcriptome of Phalaenopsis and then performed de novo transcriptome assembly. We assembled 79,434,350 clean reads into 31,708 isogenes and generated 26,565 annotated unigenes. We assigned Gene Ontology (GO) terms, Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations, and potential Pfam domains to each transcript. Using the transcriptome data as a reference, we next analyzed the differential gene expression of explants cultured for 0, 3, and 6 d, respectively. We then identified differentially expressed genes (DEGs) before and after Phalaenopsis explant browning. We also performed GO, KEGG functional enrichment and Pfam analysis of all DEGs. Finally, we selected 11 genes for quantitative real-time PCR (qPCR) analysis to confirm the expression profile analysis. Here, we report the first comprehensive analysis of transcriptome and expression profiles during Phalaenopsis explant browning. Our results suggest that Phalaenopsis explant browning may be due in part to gene expression changes that affect the secondary metabolism, such as: phenylpropanoid pathway and flavonoid biosynthesis. Genes involved in photosynthesis and ATPase activity have been found to be changed at transcription level; these changes may perturb energy metabolism and thus lead to the decay of plant cells and tissues. This study provides comprehensive gene expression data for Phalaenopsis browning. Our data constitute an important resource for further functional studies to prevent explant browning.

  20. Genome-Wide Transcriptome and Expression Profile Analysis of Phalaenopsis during Explant Browning

    PubMed Central

    Xu, Chuanjun; Zeng, Biyu; Huang, Junmei; Huang, Wen; Liu, Yumei

    2015-01-01

    Background Explant browning presents a major problem for in vitro culture, and can lead to the death of the explant and failure of regeneration. Considerable work has examined the physiological mechanisms underlying Phalaenopsis leaf explant browning, but the molecular mechanisms of browning remain elusive. In this study, we used whole genome RNA sequencing to examine Phalaenopsis leaf explant browning at genome-wide level. Methodology/Principal Findings We first used Illumina high-throughput technology to sequence the transcriptome of Phalaenopsis and then performed de novo transcriptome assembly. We assembled 79,434,350 clean reads into 31,708 isogenes and generated 26,565 annotated unigenes. We assigned Gene Ontology (GO) terms, Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations, and potential Pfam domains to each transcript. Using the transcriptome data as a reference, we next analyzed the differential gene expression of explants cultured for 0, 3, and 6 d, respectively. We then identified differentially expressed genes (DEGs) before and after Phalaenopsis explant browning. We also performed GO, KEGG functional enrichment and Pfam analysis of all DEGs. Finally, we selected 11 genes for quantitative real-time PCR (qPCR) analysis to confirm the expression profile analysis. Conclusions/Significance Here, we report the first comprehensive analysis of transcriptome and expression profiles during Phalaenopsis explant browning. Our results suggest that Phalaenopsis explant browning may be due in part to gene expression changes that affect the secondary metabolism, such as: phenylpropanoid pathway and flavonoid biosynthesis. Genes involved in photosynthesis and ATPase activity have been found to be changed at transcription level; these changes may perturb energy metabolism and thus lead to the decay of plant cells and tissues. This study provides comprehensive gene expression data for Phalaenopsis browning. Our data constitute an important resource for further

  1. Transcriptome and exoproteome analysis of utilization of plant-derived biomass by Myceliophthora thermophila.

    PubMed

    Kolbusz, Magdalena Anna; Di Falco, Marcos; Ishmael, Nadeeza; Marqueteau, Sandrine; Moisan, Marie-Claude; Baptista, Cassio da Silva; Powlowski, Justin; Tsang, Adrian

    2014-11-01

    Myceliophthora thermophila is a thermophilic fungus whose genome encodes a wide range of carbohydrate-active enzymes (CAZymes) involved in plant biomass degradation. Such enzymes have potential applications in turning different kinds of lignocellulosic feedstock into sugar precursors for biofuels and chemicals. The present study examined and compared the transcriptomes and exoproteomes of M. thermophila during cultivation on different types of complex biomass to gain insight into how its secreted enzymatic machinery varies with different sources of lignocellulose. In the transcriptome analysis three monocot (barley, oat, triticale) and three dicot (alfalfa, canola, flax) plants were used whereas in the proteome analysis additional substrates, i.e. wood and corn stover pulps, were included. A core set of 59 genes encoding CAZymes was up-regulated in response to both monocot and dicot straws, including nine polysaccharide monooxygenases and GH10, but not GH11, xylanases. Genes encoding additional xylanolytic enzymes were up-regulated during growth on monocot straws, while genes encoding additional pectinolytic enzymes were up-regulated in response to dicot biomass. Exoproteome analysis was generally consistent with the conclusions drawn from transcriptome analysis, but additional CAZymes that accumulated to high levels were identified. Despite the wide variety of biomass sources tested some CAZy family members were not expressed under any condition. The results of this study provide a comprehensive view from both transcriptome and exoproteome levels, of how M. thermophila responds to a wide range of biomass sources using its genomic resources. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Spiritual Competency Scale: Further Analysis

    ERIC Educational Resources Information Center

    Dailey, Stephanie F.; Robertson, Linda A.; Gill, Carman S.

    2015-01-01

    This article describes a follow-up analysis of the Spiritual Competency Scale, which initially validated ASERVIC's (Association for Spiritual, Ethical and Religious Values in Counseling) spiritual competencies. The study examined whether the factor structure of the Spiritual Competency Scale would be supported by participants (i.e., ASERVIC…

  3. Spiritual Competency Scale: Further Analysis

    ERIC Educational Resources Information Center

    Dailey, Stephanie F.; Robertson, Linda A.; Gill, Carman S.

    2015-01-01

    This article describes a follow-up analysis of the Spiritual Competency Scale, which initially validated ASERVIC's (Association for Spiritual, Ethical and Religious Values in Counseling) spiritual competencies. The study examined whether the factor structure of the Spiritual Competency Scale would be supported by participants (i.e., ASERVIC…

  4. Quantitative proteome and transcriptome analysis of the archaeon Thermoplasma acidophilum cultured under aerobic and anaerobic conditions.

    PubMed

    Sun, Na; Pan, Cuiping; Nickell, Stephan; Mann, Matthias; Baumeister, Wolfgang; Nagy, István

    2010-09-03

    A comparative proteome and transcriptome analysis of Thermoplasma acidophilum cultured under aerobic and anaerobic conditions has been performed. One-thousand twenty-five proteins were identified covering 88% of the cytosolic proteome. Using a label-free quantitation method, we found that approximately one-quarter of the identified proteome (263 proteins) were significantly induced (>2 fold) under anaerobic conditions. Thirty-nine macromolecular complexes were identified, of which 28 were quantified and 15 were regulated under anaerobiosis. In parallel, a whole genome cDNA microarray analysis was performed showing that the expression levels of 445 genes were influenced by the absence of oxygen. Interestingly, more than 40% of the membrane protein-encoding genes (145 out of 335 ORFs) were up- or down-regulated at the mRNA level. Many of these proteins are functionally associated with extracellular protein or peptide degradation or ion and amino acid transport. Comparison of the transcriptome and proteome showed only a weak positive correlation between mRNA and protein expression changes, which is indicative of extensive post-transcriptional regulatory mechanisms in T. acidophilum. Integration of transcriptomics and proteomics data generated hypotheses for physiological adaptations of the cells to anaerobiosis, and the quantitative proteomics data together with quantitative analysis of protein complexes provide a platform for correlation of MS-based proteomics studies with cryo-electron tomography-based visual proteomics approaches.

  5. Genetic signals of high-altitude adaptation in amphibians: a comparative transcriptome analysis.

    PubMed

    Yang, Weizhao; Qi, Yin; Fu, Jinzhong

    2016-10-03

    High-altitude adaptation provides an excellent system for studying how organisms cope with multiple environmental stressors and interacting genetic modifications. To explore the genetic basis of high-altitude adaptation in poikilothermic animals, we acquired transcriptome sequences from a high-altitude population and a low-altitude population of the Asiatic toad (Bufo gargarizans). Transcriptome data from another high-altitude amphibian, Rana kukunoris and its low-altitude relative R. chensiensis, which are from a previous study, were also incorporated into our comparative analysis. More than 40,000 transcripts were obtained from each transcriptome, and 5107 one-to-one orthologs were identified among the four taxa for comparative analysis. A total of 29 (Bufo) and 33 (Rana) putative positively selected genes were identified for the two high-altitude species, which were mainly concentrated in nutrient metabolism related functions. Using SNP-tagging and FST outlier analysis, we further tested 89 other nutrient metabolism related genes for signatures of natural selection, and found that two genes, CAPN2 and ITPR1, were likely under balancing selection. We did not detect any positively selected genes associated with response to hypoxia. Amphibians clearly employ different genetic mechanisms for high-altitude adaptation compared to endotherms. Modifications of genes associated with nutrient metabolism feature prominently while genes related to hypoxia tolerance appear to be insignificant. Poikilotherms represent the majority of animal diversity, and we hope that our results will provide useful directions for future studies of amphibians as well as other poikilotherms.

  6. Transcriptomic Analysis of Cadmium Stress Response in the Heavy Metal Hyperaccumulator Sedum alfredii Hance

    PubMed Central

    Yang, Xiaoe; Liu, Jian-Xiang

    2013-01-01

    The Sedum alfredii Hance hyperaccumulating ecotype (HE) has the ability to hyperaccumulate cadmium (Cd), as well as zinc (Zn) and lead (Pb) in above-ground tissues. Although many physiological studies have been conducted with these plants, the molecular mechanisms underlying their hyper-tolerance to heavy metals are largely unknown. Here we report on the generation of 9.4 gigabases of adaptor-trimmed raw sequences and the assembly of 57,162 transcript contigs in S. alfredii Hance (HE) shoots by the combination of Roche 454 and Illumina/Solexa deep sequencing technologies. We also have functionally annotated the transcriptome and analyzed the transcriptome changes upon Cd hyperaccumulation in S. alfredii Hance (HE) shoots. There are 110 contigs and 123 contigs that were up-regulated (Fold Change ≧2.0) and down-regulated (Fold Change ≦0.5) by chronic Cd treatment in S. alfredii Hance (HE) at q-value cutoff of 0.005, respectively. Quantitative RT-PCR was employed to compare gene expression patterns between S. alfredii Hance (HE) and non-hyperaccumulating ecotype (NHE). Our results demonstrated that several genes involved in cell wall modification, metal translocation and remobilization were more induced or constitutively expressed at higher levels in HE shoots than that in NHE shoots in response to Cd exposure. Together, our study provides large-scale expressed sequence information and genome-wide transcriptome profiling of Cd responses in S. alfredii Hance (HE) shoots. PMID:23755133

  7. De novo assembly and comparative transcriptome analysis of Euglena gracilis in response to anaerobic conditions.

    PubMed

    Yoshida, Yuta; Tomiyama, Takuya; Maruta, Takanori; Tomita, Masaru; Ishikawa, Takahiro; Arakawa, Kazuharu

    2016-03-03

    The phytoflagellated protozoan, Euglena gracilis, has been proposed as an attractive feedstock for the accumulation of valuable compounds such as β-1,3-glucan, also known as paramylon, and wax esters. The production of wax esters proceeds under anaerobic conditions, designated as wax ester fermentation. In spite of the importance and usefulness of Euglena, the genome and transcriptome data are currently unavailable, though another research group has recently published E.gracilis transcriptome study during our submission. We herein performed an RNA-Seq analysis to provide a comprehensive sequence resource and some insights into the regulation of genes including wax ester metabolism by comparative transcriptome analysis of E.gracilis under aerobic and anaerobic conditions. The E.gracilis transcriptome analysis was performed using the Illumina platform and yielded 90.3 million reads after the filtering steps. A total of 49,826 components were assembled and identified as a reference sequence of E.gracilis, of which 26,479 sequences were considered to be potentially expressed (having FPKM value of greater than 1). Approximately half of all components were estimated to be regulated in a trans-splicing manner, with the addition of protruding spliced leader sequences. Nearly 40 % of 26,479 sequences were annotated by similarity to Swiss-Prot database using the BLASTX program. A total of 2080 transcripts were identified as differentially expressed genes (DEGs) in response to anaerobic treatment for 24 h. A comprehensive pathway enrichment analysis using the KEGG pathway revealed that the majority of DEGs were involved in photosynthesis, nucleotide metabolism, oxidative phosphorylation, fatty acid metabolism. We successfully identified a candidate gene set of paramylon and wax esters, including novel β-1,3-glucan and wax ester synthases. A comparative expression analysis of aerobic- and anaerobic-treated E.gracilis cells indicated that gene expression changes in these

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

    PubMed

    Degletagne, Cyril; Keime, Céline; Rey, Benjamin; de Dinechin, Marc; Forcheron, Fabien; Chuchana, Paul; Jouventin, Pierre; Gautier, Christian; Duchamp, Claude

    2010-05-31

    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. 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. MAXRS appears to be an appropriate method for gene expression analysis under heterologous hybridization conditions.

  9. 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

  10. jNMFMA: a joint non-negative matrix factorization meta-analysis of transcriptomics data.

    PubMed

    Wang, Hong-Qiang; Zheng, Chun-Hou; Zhao, Xing-Ming

    2015-02-15

    Tremendous amount of omics data being accumulated poses a pressing challenge of meta-analyzing the heterogeneous data for mining new biological knowledge. Most existing methods deal with each gene independently, thus often resulting in high false positive rates in detecting differentially expressed genes (DEG). To our knowledge, no or little effort has been devoted to methods that consider dependence structures underlying transcriptomics data for DEG identification in meta-analysis context. This article proposes a new meta-analysis method for identification of DEGs based on joint non-negative matrix factorization (jNMFMA). We mathematically extend non-negative matrix factorization (NMF) to a joint version (jNMF), which is used to simultaneously decompose multiple transcriptomics data matrices into one common submatrix plus multiple individual submatrices. By the jNMF, the dependence structures underlying transcriptomics data can be interrogated and utilized, while the high-dimensional transcriptomics data are mapped into a low-dimensional space spanned by metagenes that represent hidden biological signals. jNMFMA finally identifies DEGs as genes that are associated with differentially expressed metagenes. The ability of extracting dependence structures makes jNMFMA more efficient and robust to identify DEGs in meta-analysis context. Furthermore, jNMFMA is also flexible to identify DEGs that are consistent among various types of omics data, e.g. gene expression and DNA methylation. Experimental results on both simulation data and real-world cancer data demonstrate the effectiveness of jNMFMA and its superior performance over other popular approaches. R code for jNMFMA is available for non-commercial use via http://micblab.iim.ac.cn/Download/. hqwang@ustc.edu Supplementary data are available at Bioinformatics online. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. Perigone Lobe Transcriptome Analysis Provides Insights into Rafflesia cantleyi Flower Development

    PubMed Central

    Lee, Xin-Wei; Mat-Isa, Mohd-Noor; Mohd-Elias, Nur-Atiqah; Aizat-Juhari, Mohd Afiq; Goh, Hoe-Han; Dear, Paul H.; Chow, Keng-See; Haji Adam, Jumaat; Mohamed, Rahmah; Firdaus-Raih, Mohd; Wan, Kiew-Lian

    2016-01-01

    Rafflesia is a biologically enigmatic species that is very rare in occurrence and possesses an extraordinary morphology. This parasitic plant produces a gigantic flower up to one metre in diameter with no leaves, stem or roots. However, little is known about the floral biology of this species especially at the molecular level. In an effort to address this issue, we have generated and characterised the transcriptome of the Rafflesia cantleyi flower, and performed a comparison with the transcriptome of its floral bud to predict genes that are expressed and regulated during flower development. Approximately 40 million sequencing reads were generated and assembled de novo into 18,053 transcripts with an average length of 641 bp. Of these, more than 79% of the transcripts had significant matches to annotated sequences in the public protein database. A total of 11,756 and 7,891 transcripts were assigned to Gene Ontology categories and clusters of orthologous groups respectively. In addition, 6,019 transcripts could be mapped to 129 pathways in Kyoto Encyclopaedia of Genes and Genomes Pathway database. Digital abundance analysis identified 52 transcripts with very high expression in the flower transcriptome of R. cantleyi. Subsequently, analysis of differential expression between developing flower and the floral bud revealed a set of 105 transcripts with potential role in flower development. Our work presents a deep transcriptome resource analysis for the developing flower of R. cantleyi. Genes potentially involved in the growth and development of the R. cantleyi flower were identified and provide insights into biological processes that occur during flower development. PMID:27977777

  12. Perigone Lobe Transcriptome Analysis Provides Insights into Rafflesia cantleyi Flower Development.

    PubMed

    Lee, Xin-Wei; Mat-Isa, Mohd-Noor; Mohd-Elias, Nur-Atiqah; Aizat-Juhari, Mohd Afiq; Goh, Hoe-Han; Dear, Paul H; Chow, Keng-See; Haji Adam, Jumaat; Mohamed, Rahmah; Firdaus-Raih, Mohd; Wan, Kiew-Lian

    2016-01-01

    Rafflesia is a biologically enigmatic species that is very rare in occurrence and possesses an extraordinary morphology. This parasitic plant produces a gigantic flower up to one metre in diameter with no leaves, stem or roots. However, little is known about the floral biology of this species especially at the molecular level. In an effort to address this issue, we have generated and characterised the transcriptome of the Rafflesia cantleyi flower, and performed a comparison with the transcriptome of its floral bud to predict genes that are expressed and regulated during flower development. Approximately 40 million sequencing reads were generated and assembled de novo into 18,053 transcripts with an average length of 641 bp. Of these, more than 79% of the transcripts had significant matches to annotated sequences in the public protein database. A total of 11,756 and 7,891 transcripts were assigned to Gene Ontology categories and clusters of orthologous groups respectively. In addition, 6,019 transcripts could be mapped to 129 pathways in Kyoto Encyclopaedia of Genes and Genomes Pathway database. Digital abundance analysis identified 52 transcripts with very high expression in the flower transcriptome of R. cantleyi. Subsequently, analysis of differential expression between developing flower and the floral bud revealed a set of 105 transcripts with potential role in flower development. Our work presents a deep transcriptome resource analysis for the developing flower of R. cantleyi. Genes potentially involved in the growth and development of the R. cantleyi flower were identified and provide insights into biological processes that occur during flower development.

  13. MicroRNAs shape circadian hepatic gene expression on a transcriptome-wide scale

    PubMed Central

    Du, Ngoc-Hien; Arpat, Alaaddin Bulak; De Matos, Mara; Gatfield, David

    2014-01-01

    A considerable proportion of mammalian gene expression undergoes circadian oscillations. Post-transcriptional mechanisms likely make important contributions to mRNA abundance rhythms. We have investigated how microRNAs (miRNAs) contribute to core clock and clock-controlled gene expression using mice in which miRNA biogenesis can be inactivated in the liver. While the hepatic core clock was surprisingly resilient to miRNA loss, whole transcriptome sequencing uncovered widespread effects on clock output gene expression. Cyclic transcription paired with miRNA-mediated regulation was thus identified as a frequent phenomenon that affected up to 30% of the rhythmic transcriptome and served to post-transcriptionally adjust the phases and amplitudes of rhythmic mRNA accumulation. However, only few mRNA rhythms were actually generated by miRNAs. Overall, our study suggests that miRNAs function to adapt clock-driven gene expression to tissue-specific requirements. Finally, we pinpoint several miRNAs predicted to act as modulators of rhythmic transcripts, and identify rhythmic pathways particularly prone to miRNA regulation. DOI: http://dx.doi.org/10.7554/eLife.02510.001 PMID:24867642

  14. Comparative transcriptomic analysis between an artificially induced SIRS in healthy individuals and spontaneous sepsis.

    PubMed

    Monteiro Sousa, Claudio; Boissel, Jean-Pierre; Gueyffier, François; Olivera-Botello, Gustavo

    2015-10-01

    Sepsis is defined as a syndrome combining a systemic inflammatory response with a documented infection. It may progress to more serious cases such as septic shock following the failure of one or more organs and the emergence of hemodynamic defects. Assuming that the emergence of serious septic syndromes may be partially explained by the early loss of regulation of the inflammatory response, we decided to compare, in a transcriptomic perspective, the biological mechanisms expressed during an induced systemic inflammatory response with those expressed during severe septic syndromes. By using open-access transcriptomic databases, we first studied the kinetics of an induced inflammatory response. The use of functional analysis helped us identify discriminating biological mechanisms, such as the mTOR signaling pathway, between the pathological cases of sepsis and non-pathological (i.e., the artificially induced SIRS) cases. Copyright © 2015 Académie des sciences. Published by Elsevier SAS. All rights reserved.

  15. Longitudinal analysis of the group A Streptococcus transcriptome in experimental pharyngitis in cynomolgus macaques

    PubMed Central

    Virtaneva, Kimmo; Porcella, Stephen F.; Graham, Morag R.; Ireland, Robin M.; Johnson, Claire A.; Ricklefs, Stacy M.; Babar, Imran; Parkins, Larye D.; Romero, Romina A.; Corn, G. Judson; Gardner, Don J.; Bailey, John R.; Parnell, Michael J.; Musser, James M.

    2005-01-01

    Identification of the genetic events that contribute to host–pathogen interactions is important for understanding the natural history of infectious diseases and developing therapeutics. Transcriptome studies conducted on pathogens have been central to this goal in recent years. However, most of these investigations have focused on specific end points or disease phases, rather than analysis of the entire time course of infection. To gain a more complete understanding of how bacterial gene expression changes over time in a primate host, the transcriptome of group A Streptococcus (GAS) was analyzed during an 86-day infection protocol in 20 cynomolgus macaques with experimental pharyngitis. The study used 260 custom Affymetrix (Santa Clara, CA) chips, and data were confirmed by TaqMan analysis. Colonization, acute, and asymptomatic phases of disease were identified. Successful colonization and severe inflammation were significantly correlated with an early onset of superantigen gene expression. The differential expression of two-component regulators covR and spy0680 (M1_spy0874) was significantly associated with GAS colony-forming units, inflammation, and phases of disease. Prophage virulence gene expression and prophage induction occurred predominantly during high pathogen cell densities and acute inflammation. We discovered that temporal changes in the GAS transcriptome were integrally linked to the phase of clinical disease and host-defense response. Knowledge of the gene expression patterns characterizing each phase of pathogen–host interaction provides avenues for targeted investigation of proven and putative virulence factors and genes of unknown function and will assist vaccine research. PMID:15956184

  16. Analysis of upland cotton (Gossypium hirsutum) response to Verticillium dahliae inoculation by transcriptome sequencing.

    PubMed

    Shao, B X; Zhao, Y L; Chen, W; Wang, H M; Guo, Z J; Gong, H Y; Sang, X H; Cui, Y L; Wang, C H

    2015-10-27

    Verticillium wilt is one of the main diseases in cotton (Gossypium hirsutum), severely reduces yield and fiber quality, and is difficult to be con-trolled effectively. At present, the molecular mechanism that confers resistance to this disease is unclear. Transcriptome sequencing is an important method to detect resistance genes, explore metabolic pathways, and study resistance mechanisms. In this study, the transcriptome of a disease-resistant inbred cot-ton line inoculated with Verticillium dahliae was sequenced. A total of 126,402 unigenes were obtained using de novo assembly and data analysis, 99,712 (78.88%) of which were annotated into the Nr, Nt, Swiss-Prot, KEGG, COG, and GO databases. The expression patterns of 16 candidate disease-resis-tance genes showed that some genes were upregulated soon after V. dahliae inoculation and others were upregulated later, which may indicate instanta-neous basal defense and lagged specific defense, respectively. We conducted a preliminary analysis of the transcriptome database, which will contribute to further research regarding the cloning of disease-resistance genes.

  17. Comparative Transcriptome Analysis Reveals Sex-Biased Gene Expression in Juvenile Chinese Mitten Crab Eriocheir sinensis

    PubMed Central

    Liu, Yuan; Hui, Min; Cui, Zhaoxia; Luo, Danli; Song, Chengwen; Li, Yingdong; Liu, Lei

    2015-01-01

    Sex-biased genes are considered to account for most of phenotypic differences between males and females. In order to explore the sex-biased gene expression in crab, we performed the whole-body transcriptome analysis in male and female juveniles of the Chinese mitten crab Eriocheir sinensis using next-generation sequencing technology. Of the 23,349 annotated unigenes, 148 were identified as sex-related genes. A total of 29 candidate genes involved in primary sex determination pathways were detected, indicating the sex determination cascade of the mitten crab might be more complex than previously supposed. Differential expression analysis showed 448 differentially expressed genes (DEGs) between the two transcriptomes. Most of DEGs were involved in processes such as metabolism and immunity, and not associated with obvious sexual function. The pathway predominantly enriched for DEGs were related to lysosome, which might reflect the differences in metabolism between males and females. Of the immune DGEs, 18 up-regulated genes in females were humoral immune factors, and eight up-regulated genes in males were pattern recognition receptors, suggesting sex differences of immune defense might exist in the mitten crab. In addition, two reproduction-related genes, vitellogenin and insulin-like androgenic gland factor, were identified to express in both sexes but with significantly higher level in males. Our research provides the first whole-body RNA sequencing of sex-specific transcriptomes for juvenile E. sinensis and will facilitate further studies on molecular mechanisms of crab sexual dimorphism. PMID:26193085

  18. Comparative Transcriptome Analysis Reveals Sex-Biased Gene Expression in Juvenile Chinese Mitten Crab Eriocheir sinensis.

    PubMed

    Liu, Yuan; Hui, Min; Cui, Zhaoxia; Luo, Danli; Song, Chengwen; Li, Yingdong; Liu, Lei

    2015-01-01

    Sex-biased genes are considered to account for most of phenotypic differences between males and females. In order to explore the sex-biased gene expression in crab, we performed the whole-body transcriptome analysis in male and female juveniles of the Chinese mitten crab Eriocheir sinensis using next-generation sequencing technology. Of the 23,349 annotated unigenes, 148 were identified as sex-related genes. A total of 29 candidate genes involved in primary sex determination pathways were detected, indicating the sex determination cascade of the mitten crab might be more complex than previously supposed. Differential expression analysis showed 448 differentially expressed genes (DEGs) between the two transcriptomes. Most of DEGs were involved in processes such as metabolism and immunity, and not associated with obvious sexual function. The pathway predominantly enriched for DEGs were related to lysosome, which might reflect the differences in metabolism between males and females. Of the immune DGEs, 18 up-regulated genes in females were humoral immune factors, and eight up-regulated genes in males were pattern recognition receptors, suggesting sex differences of immune defense might exist in the mitten crab. In addition, two reproduction-related genes, vitellogenin and insulin-like androgenic gland factor, were identified to express in both sexes but with significantly higher level in males. Our research provides the first whole-body RNA sequencing of sex-specific transcriptomes for juvenile E. sinensis and will facilitate further studies on molecular mechanisms of crab sexual dimorphism.

  19. Transcriptome analysis of Cucumis sativus infected by Cucurbit chlorotic yellows virus.

    PubMed

    Sun, Xinyan; Wang, Zhenyue; Gu, Qinsheng; Li, Honglian; Han, Weili; Shi, Yan

    2017-02-02

    Cucurbit chlorotic yellows virus (CCYV) is a recently reported bipartite crinivirus that causes chlorotic leaf spots and yellowing symptoms on the leaves of cucurbit plants. The virus-host interaction of CCYV remains to be elucidated, and the influence of criniviruses on the host gene transcriptome requires analysis. We used transcriptome sequencing to analyse the differentially expressed genes (DEGs) caused by CCYV infection. CCYV infection resulted in 865 DEGs. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis identified 67 pathways, and the three major enrichment pathways (according to the P-values) were photosynthesis-antenna proteins (KO00196), phenylalanine metabolism (KO00360a), and phenylpropanoid biosynthesis (KO00940). Of the 13 DEGs identified in phenylalanine metabolism, 11 genes encode disease resistance-related phenylalanine ammonia-lyase (PAL) genes. Using quantitative real-time PCR, we validated the differential expression of 12 genes. Our study based on the CCYV-cucumber interaction provides comprehensive transcriptomic information, and will improve our understanding of host-crinivirus interactions.

  20. 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.

  1. Transcriptome sequencing and analysis of leaf tissue of Avicennia marina using the Illumina platform.

    PubMed

    Huang, Jianzi; Lu, Xiang; Zhang, Wanke; Huang, Rongfeng; Chen, Shouyi; Zheng, Yizhi

    2014-01-01

    Avicennia marina is a widely distributed mangrove species that thrives in high-salinity habitats. It plays a significant role in supporting coastal ecosystem and holds unique potential for studying molecular mechanisms underlying ecological adaptation. Despite and sometimes because of its numerous merits, this species is facing increasing pressure of exploitation and deforestation. Both study on adaptation mechanisms and conservation efforts necessitate more genomic resources for A. marina. In this study, we used Illumina sequencing of an A. marina foliar cDNA library to generate a transcriptome dataset for gene and marker discovery. We obtained 40 million high-quality reads and assembled them into 91,125 unigenes with a mean length of 463 bp. These unigenes covered most of the publicly available A. marina Sanger ESTs and greatly extended the repertoire of transcripts for this species. A total of 54,497 and 32,637 unigenes were annotated based on homology to sequences in the NCBI non-redundant and the Swiss-prot protein databases, respectively. Both Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed some transcriptomic signatures of stress adaptation for this halophytic species. We also detected an extraordinary amount of transcripts derived from fungal endophytes and demonstrated the utility of transcriptome sequencing in surveying endophyte diversity without isolating them out of plant tissues. Additionally, we identified 3,423 candidate simple sequence repeats (SSRs) from 3,141 unigenes with a density of one SSR locus every 8.25 kb sequence. Our transcriptomic data will provide valuable resources for ecological, genetic and evolutionary studies in A. marina.

  2. Transcriptome Sequencing and Analysis of Leaf Tissue of Avicennia marina Using the Illumina Platform

    PubMed Central

    Zhang, Wanke; Huang, Rongfeng; Chen, Shouyi; Zheng, Yizhi

    2014-01-01

    Avicennia marina is a widely distributed mangrove species that thrives in high-salinity habitats. It plays a significant role in supporting coastal ecosystem and holds unique potential for studying molecular mechanisms underlying ecological adaptation. Despite and sometimes because of its numerous merits, this species is facing increasing pressure of exploitation and deforestation. Both study on adaptation mechanisms and conservation efforts necessitate more genomic resources for A. marina. In this study, we used Illumina sequencing of an A. marina foliar cDNA library to generate a transcriptome dataset for gene and marker discovery. We obtained 40 million high-quality reads and assembled them into 91,125 unigenes with a mean length of 463 bp. These unigenes covered most of the publicly available A. marina Sanger ESTs and greatly extended the repertoire of transcripts for this species. A total of 54,497 and 32,637 unigenes were annotated based on homology to sequences in the NCBI non-redundant and the Swiss-prot protein databases, respectively. Both Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed some transcriptomic signatures of stress adaptation for this halophytic species. We also detected an extraordinary amount of transcripts derived from fungal endophytes and demonstrated the utility of transcriptome sequencing in surveying endophyte diversity without isolating them out of plant tissues. Additionally, we identified 3,423 candidate simple sequence repeats (SSRs) from 3,141 unigenes with a density of one SSR locus every 8.25 kb sequence. Our transcriptomic data will provide valuable resources for ecological, genetic and evolutionary studies in A. marina. PMID:25265387

  3. Transcriptome Analysis of Hamelia patens (Rubiaceae) Anthers Reveals Candidate Genes for Tapetum and Pollen Wall Development

    PubMed Central

    Yue, Lin; Twell, David; Kuang, Yanfeng; Liao, Jingping; Zhou, Xianqiang

    2017-01-01

    Studies of the anther transcriptome on non-model plants without a known genome are surprisingly scarce. RNA-Seq and digital gene expression (DGE) profiling provides a comprehensive approach to identify candidate genes contributing to developmental processes in non-model species. Here we built a transcriptome library of developing anthers of Hamelia patens and analyzed DGE profiles from each stage to identify genes that regulate tapetum and pollen development. In total 7,720 putative differentially expressed genes across four anther stages were identified. The number of putative stage-specific genes was: 776 at microspore mother cell stage, 807 at tetrad stage, 322 at uninucleate microspore stage, and the highest number (1,864) at bicellular pollen stage. GO enrichment analysis revealed 243 differentially expressed and 108 stage-specific genes that are potentially related to tapetum development, sporopollenin synthesis, and pollen wall. The number of expressed genes, their function and expression profiles were all significantly correlated with anther developmental processes. Overall comparisons of anther and pollen transcriptomes with those of rice and Arabidopsis together with the expression profiles of homologs of known anther-expressed genes, revealed conserved patterns and also divergence. The divergence may reflect taxon-specific differences in gene expression, the use RNA-seq as a more sensitive methodology, variation in tissue composition and sampling strategies. Given the lack of genomic sequence, this study succeeded in assigning putative identity to a significant proportion of anther-expressed genes and genes relevant to tapetum and pollen development in H. patens. The anther transcriptome revealed a molecular distinction between developmental stages, serving as a resource to unravel the functions of genes involved in anther development in H. patens and informing the analysis of other members of the Rubiaceae. PMID:28119704

  4. Proteomic and transcriptomic analysis of saliva components from the hematophagous reduviid Triatoma pallidipennis.

    PubMed

    Hernández-Vargas, María J; Gil, Jeovanis; Lozano, Luis; Pedraza-Escalona, Martha; Ortiz, Ernesto; Encarnación-Guevara, Sergio; Alagón, Alejandro; Corzo, Gerardo

    2017-06-06

    Species belonging to the Triatominae subfamily are commonly associated with Chagas disease, as they are potential vectors of the parasite Trypanosoma cruzi. However, their saliva contains a cocktail of diverse anti-hemostatic proteins that prevent blood coagulation, vasodilation and platelet aggregation of blood; components with indisputable therapeutic potential. We performed a transcriptomic and proteomic analyses of salivary glands and protein spots from 2DE gels of milked saliva, respectively, from the Mexican Triatoma pallidipennis. Massive sequencing techniques were used to reveal this protein diversity. A total of 78 out of 233 transcripts were identified as proteins in the saliva, divided among 43 of 55 spots from 2DE gels of saliva, identified by LC-MS/MS analysis. Some of the annotated transcripts putatively code for anti-hemostatic proteins, which share sequence similarities with proteins previously described for South American triatomines. The most abundant as well as diverse transcripts and proteins in the saliva were the anti-hemostatic triabins. For the first time, a transcriptomic analysis uncovered other unrelated but relevant components in triatomines, including antimicrobial and thrombolytic polypeptides. Likewise, unique proteins such as the angiotensin-converting enzyme were identified not just in the salivary gland transcriptome but also at saliva proteome of this North American bloodsucking insect. This manuscript is the first report of the correlation between proteome and transcriptome of Triatoma pallidipennis, which shows for the first time the presence of proteins in this insect that have not been characterized in other species of this family. This information contributes to a better understanding of the multiple host defense mechanisms that are being affected at the moment of blood ingestion by the insect. Furthermore, this report gives a repertoire of possible therapeutic proteins. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. OM-FBA: Integrate Transcriptomics Data with Flux Balance Analysis to Decipher the Cell Metabolism.

    PubMed

    Guo, Weihua; Feng, Xueyang

    2016-01-01

    Constraint-based metabolic modeling such as flux balance analysis (FBA) has been widely used to simulate cell metabolism. Thanks to its simplicity and flexibility, numerous algorithms have been developed based on FBA and successfully predicted the phenotypes of various biological systems. However, their phenotype predictions may not always be accurate in FBA because of using the objective function that is assumed for cell metabolism. To overcome this challenge, we have developed a novel computational framework, namely omFBA, to integrate multi-omics data (e.g. transcriptomics) into FBA to obtain omics-guided objective functions with high accuracy. In general, we first collected transcriptomics data and phenotype data from published database (e.g. GEO database) for different microorganisms such as Saccharomyces cerevisiae. We then developed a "Phenotype Match" algorithm to derive an objective function for FBA that could lead to the most accurate estimation of the known phenotype (e.g. ethanol yield). The derived objective function was next correlated with the transcriptomics data via regression analysis to generate the omics-guided objective function, which was next used to accurately simulate cell metabolism at unknown conditions. We have applied omFBA in studying sugar metabolism of S. cerevisiae and found that the ethanol yield could be accurately predicted in most of the cases tested (>80%) by using transcriptomics data alone, and revealed valuable metabolic insights such as the dynamics of flux ratios. Overall, omFBA presents a novel platform to potentially integrate multi-omics data simultaneously and could be incorporated with other FBA-derived tools by replacing the arbitrary objective function with the omics-guided objective functions.

  6. Acclimation of Antarctic Chlamydomonas to the sea-ice environment: a transcriptomic analysis.

    PubMed

    Liu, Chenlin; Wang, Xiuliang; Wang, Xingna; Sun, Chengjun

    2016-07-01

    The Antarctic green alga Chlamydomonas sp. ICE-L was isolated from sea ice. As a psychrophilic microalga, it can tolerate the environmental stress in the sea-ice brine, such as freezing temperature and high salinity. We performed a transcriptome analysis to identify freezing stress responding genes and explore the extreme environmental acclimation-related strategies. Here, we show that many genes in ICE-L transcriptome that encoding PUFA synthesis enzymes, molecular chaperon proteins, and cell membrane transport proteins have high similarity to the gens from Antarctic bacteria. These ICE-L genes are supposed to be acquired through horizontal gene transfer from its symbiotic microbes in the sea-ice brine. The presence of these genes in both sea-ice microalgae and bacteria indicated the biological processes they involved in are possibly contributing to ICE-L success in sea ice. In addition, the biological pathways were compared between ICE-L and its closely related sister species, Chlamydomonas reinhardtii and Volvox carteri. In ICE-L transcripome, many sequences homologous to the plant or bacteria proteins in the post-transcriptional, post-translational modification, and signal-transduction KEGG pathways, are absent in the nonpsychrophilic green algae. These complex structural components might imply enhanced stress adaptation capacity. At last, differential gene expression analysis at the transcriptome level of ICE-L indicated that genes that associated with post-translational modification, lipid metabolism, and nitrogen metabolism are responding to the freezing treatment. In conclusion, the transcriptome of Chlamydomonas sp. ICE-L is very useful for exploring the mutualistic interaction between microalgae and bacteria in sea ice; and discovering the specific genes and metabolism pathways responding to the freezing acclimation in psychrophilic microalgae.

  7. Characterization of the Floral Transcriptome of Moso Bamboo (Phyllostachys edulis) at Different Flowering Developmental Stages by Transcriptome Sequencing and RNA-Seq Analysis

    PubMed Central

    Zhang, Chunling; Qi, Feiyan; Li, Xueping; Mu, Shaohua; Peng, Zhenhua

    2014-01-01

    Background As an arborescent and perennial plant, Moso bamboo (Phyllostachys edulis (Carrière) J. Houzeau, synonym Phyllostachys heterocycla Carrière) is characterized by its infrequent sexual reproduction with flowering intervals ranging from several to more than a hundred years. However, little bamboo genomic research has been conducted on this due to a variety of reasons. Here, for the first time, we investigated the transcriptome of developing flowers in Moso bamboo by using high-throughput Illumina GAII sequencing and mapping short reads to the Moso bamboo genome and reference genes. We performed RNA-seq analysis on four important stages of flower development, and obtained extensive gene and transcript abundance data for the floral transcriptome of this key bamboo species. Results We constructed a cDNA library using equal amounts of RNA from Moso bamboo leaf samples from non-flowering plants (CK) and mixed flower samples (F) of four flower development stages. We generated more than 67 million reads from each of the CK and F samples. About 70% of the reads could be uniquely mapped to the Moso bamboo genome and the reference genes. Genes detected at each stage were categorized to putative functional categories based on their expression patterns. The analysis of RNA-seq data of bamboo flowering tissues at different developmental stages reveals key gene expression properties during the flower development of bamboo. Conclusion We showed that a combination of transcriptome sequencing and RNA-seq analysis was a powerful approach to identifying candidate genes related to floral transition and flower development in bamboo species. The results give a better insight into the mechanisms of Moso bamboo flowering and ageing. This transcriptomic data also provides an important gene resource for improving breeding for Moso bamboo. PMID:24915141

  8. Characterization of the floral transcriptome of Moso bamboo (Phyllostachys edulis) at different flowering developmental stages by transcriptome sequencing and RNA-seq analysis.

    PubMed

    Gao, Jian; Zhang, Ying; Zhang, Chunling; Qi, Feiyan; Li, Xueping; Mu, Shaohua; Peng, Zhenhua

    2014-01-01

    As an arborescent and perennial plant, Moso bamboo (Phyllostachys edulis (Carrière) J. Houzeau, synonym Phyllostachys heterocycla Carrière) is characterized by its infrequent sexual reproduction with flowering intervals ranging from several to more than a hundred years. However, little bamboo genomic research has been conducted on this due to a variety of reasons. Here, for the first time, we investigated the transcriptome of developing flowers in Moso bamboo by using high-throughput Illumina GAII sequencing and mapping short reads to the Moso bamboo genome and reference genes. We performed RNA-seq analysis on four important stages of flower development, and obtained extensive gene and transcript abundance data for the floral transcriptome of this key bamboo species. We constructed a cDNA library using equal amounts of RNA from Moso bamboo leaf samples from non-flowering plants (CK) and mixed flower samples (F) of four flower development stages. We generated more than 67 million reads from each of the CK and F samples. About 70% of the reads could be uniquely mapped to the Moso bamboo genome and the reference genes. Genes detected at each stage were categorized to putative functional categories based on their expression patterns. The analysis of RNA-seq data of bamboo flowering tissues at different developmental stages reveals key gene expression properties during the flower development of bamboo. We showed that a combination of transcriptome sequencing and RNA-seq analysis was a powerful approach to identifying candidate genes related to floral transition and flower development in bamboo species. The results give a better insight into the mechanisms of Moso bamboo flowering and ageing. This transcriptomic data also provides an important gene resource for improving breeding for Moso bamboo.

  9. Comprehensive transcriptome analysis reveals accelerated genic evolution in a Tibet fish, Gymnodiptychus pachycheilus.

    PubMed

    Yang, Liandong; Wang, Ying; Zhang, Zhaolei; He, Shunping

    2014-12-26

    Elucidating the genetic mechanisms of organismal adaptation to the Tibetan Plateau at a genomic scale can provide insights into the process of adaptive evolution. Many highland species have been investigated and various candidate genes that may be responsible for highland adaptation have been identified. However, we know little about the genomic basis of adaptation to Tibet in fishes. Here, we performed transcriptome sequencing of a schizothoracine fish (Gymnodiptychus pachycheilus) and used it to identify potential genetic mechanisms of highland adaptation. We obtained totally 66,105 assembled unigenes, of which 7,232 were assigned as putative one-to-one orthologs in zebrafish. Comparative gene annotations from several species indicated that at least 350 genes lost and 41 gained since the divergence between G. pachycheilus and zebrafish. An analysis of 6,324 orthologs among zebrafish, fugu, medaka, and spotted gar identified consistent evidence for genome-wide accelerated evolution in G. pachycheilus and only the terminal branch of G. pachycheilus had an elevated Ka/Ks ratio than the ancestral branch. Many functional categories related to hypoxia and energy metabolism exhibited rapid evolution in G. pachycheilus relative to zebrafish. Genes showing signature of rapid evolution and positive selection in the G. pachycheilus lineage were also enriched in functions associated with energy metabolism and hypoxia. The first genomic resources for fish in the Tibetan Plateau and evolutionary analyses provided some novel insights into highland adaptation in fishes and served as a foundation for future studies aiming to identify candidate genes underlying the genetic bases of adaptation to Tibet in fishes.

  10. Comprehensive Transcriptome Analysis Reveals Accelerated Genic Evolution in a Tibet Fish, Gymnodiptychus pachycheilus

    PubMed Central

    Yang, Liandong; Wang, Ying; Zhang, Zhaolei; He, Shunping

    2015-01-01

    Elucidating the genetic mechanisms of organismal adaptation to the Tibetan Plateau at a genomic scale can provide insights into the process of adaptive evolution. Many highland species have been investigated and various candidate genes that may be responsible for highland adaptation have been identified. However, we know little about the genomic basis of adaptation to Tibet in fishes. Here, we performed transcriptome sequencing of a schizothoracine fish (Gymnodiptychus pachycheilus) and used it to identify potential genetic mechanisms of highland adaptation. We obtained totally 66,105 assembled unigenes, of which 7,232 were assigned as putative one-to-one orthologs in zebrafish. Comparative gene annotations from several species indicated that at least 350 genes lost and 41 gained since the divergence between G. pachycheilus and zebrafish. An analysis of 6,324 orthologs among zebrafish, fugu, medaka, and spotted gar identified consistent evidence for genome-wide accelerated evolution in G. pachycheilus and only the terminal branch of G. pachycheilus had an elevated Ka/Ks ratio than the ancestral branch. Many functional categories related to hypoxia and energy metabolism exhibited rapid evolution in G. pachycheilus relative to zebrafish. Genes showing signature of rapid evolution and positive selection in the G. pachycheilus lineage were also enriched in functions associated with energy metabolism and hypoxia. The first genomic resources for fish in the Tibetan Plateau and evolutionary analyses provided some novel insights into highland adaptation in fishes and served as a foundation for future studies aiming to identify candidate genes underlying the genetic bases of adaptation to Tibet in fishes. PMID:25543049

  11. Transcriptomic Analysis and Meta-Analysis of Human Granulosa and Cumulus Cells

    PubMed Central

    Burnik Papler, Tanja; Vrtacnik Bokal, Eda; Maver, Ales; Kopitar, Andreja Natasa; Lovrečić, Luca

    2015-01-01

    Specific gene expression in oocytes and its surrounding cumulus (CC) and granulosa (GC) cells is needed for successful folliculogenesis and oocyte maturation. The aim of the present study was to compare genome-wide gene expression and biological functions of human GC and CC. Individual GC and CC were derived from 37 women undergoing IVF procedures. Gene expression analysis was performed using microarrays, followed by a meta-analysis. Results were validated using quantitative real-time PCR. There were 6029 differentially expressed genes (q < 10−4); of which 650 genes had a log2 FC ≥ 2. After the meta-analysis there were 3156 genes differentially expressed. Among these there were genes that have previously not been reported in human somatic follicular cells, like prokineticin 2 (PROK2), higher expressed in GC, and pregnancy up-regulated nonubiquitous CaM kinase (PNCK), higher expressed in CC. Pathways like inflammatory response and angiogenesis were enriched in GC, whereas in CC, cell differentiation and multicellular organismal development were among enriched pathways. In conclusion, transcriptomes of GC and CC as well as biological functions, are distinctive for each cell subpopulation. By describing novel genes like PROK2 and PNCK, expressed in GC and CC, we upgraded the existing data on human follicular biology. PMID:26313571

  12. Comprehensive Tissue-Specific Transcriptome Analysis Reveals Distinct Regulatory Programs during Early Tomato Fruit Development.

    PubMed

    Pattison, Richard J; Csukasi, Fabiana; Zheng, Yi; Fei, Zhangjun; van der Knaap, Esther; Catalá, Carmen

    2015-08-01

    Fruit formation and early development involve a range of physiological and morphological transformations of the various constituent tissues of the ovary. These developmental changes vary considerably according to tissue type, but molecular analyses at an organ-wide level inevitably obscure many tissue-specific phenomena. We used laser-capture microdissection coupled to high-throughput RNA sequencing to analyze the transcriptome of ovaries and fruit tissues of the wild tomato species Solanum pimpinellifolium. This laser-capture microdissection-high-throughput RNA sequencing approach allowed quantitative global profiling of gene expression at previously unobtainable levels of spatial resolution, revealing numerous contrasting transcriptome profiles and uncovering rare and cell type-specific transcripts. Coexpressed gene clusters linked specific tissues and stages to major transcriptional changes underlying the ovary-to-fruit transition and provided evidence of regulatory modules related to cell division, photosynthesis, and auxin transport in internal fruit tissues, together with parallel specialization of the pericarp transcriptome in stress responses and secondary metabolism. Analysis of transcription factor expression and regulatory motifs indicated putative gene regulatory modules that may regulate the development of different tissues and hormonal processes. Major alterations in the expression of hormone metabolic and signaling components illustrate the complex hormonal control underpinning fruit formation, with intricate spatiotemporal variations suggesting separate regulatory programs.

  13. Transcriptome profiling and digital gene expression analysis of Nile tilapia (Oreochromis niloticus) infected by Streptococcus agalactiae.

    PubMed

    Zhang, Rui; Zhang, Li-li; Ye, Xing; Tian, Yuan-yuan; Sun, Cheng-fei; Lu, Mai-xin; Bai, Jun-jie

    2013-10-01

    Tilapia is an important freshwater aquaculture species worldwide. In recent years, streptococcal diseases have severely threatened development of tilapia aquaculture, while effective prevention and control methods have not yet been established. In order to understand the immunological response of tilapia to infection by Streptococcus agalactiae (S. agalactiae), this study employed Solexa/Illumina RNA-seq and digital gene expression (DGE) technology to investigate changes in the tilapia transcriptome before and after S. agalactiae infection. We obtained 82,799 unigenes (mean size: 618 bp) using de novo assembly. Unigenes were annotated by comparing against databases including Nr, Swissprot, cluster of orthologous groups of proteins, Kyoto encyclopedia of genes and genomes, and gene ontology. Combined with DGE technology, transcriptomic changes in tilapia before and after bacteria challenging were examined. A total of 774 significantly up-regulated and 625 significantly down-regulated unigenes were identified, among which 293 were mapped to 181 signaling pathways including 17 immune-related pathways involving 65 differentially expressed genes. We observed a change in the expression of six genes in the Toll-like receptor signaling pathway, and this was subsequently confirmed via quantitative real-time PCR. This comparative study of the tilapia transcriptome before and after S. agalactiae infection identified important differentially-expressed immune-related genes and signaling pathways that will provide useful insights for further analysis of the mechanisms of tilapia defense against S. agalactiae infection.

  14. De Novo Transcriptome Analysis of Allium cepa L. (Onion) Bulb to Identify Allergens and Epitopes.

    PubMed

    Rajkumar, Hemalatha; Ramagoni, Ramesh Kumar; Anchoju, Vijayendra Chary; Vankudavath, Raju Naik; Syed, Arshi Uz Zaman

    2015-01-01

    Allium cepa (onion) is a diploid plant with one of the largest nuclear genomes among all diploids. Onion is an example of an under-researched crop which has a complex heterozygous genome. There are no allergenic proteins and genomic data available for onions. This study was conducted to establish a transcriptome catalogue of onion bulb that will enable us to study onion related genes involved in medicinal use and allergies. Transcriptome dataset generated from onion bulb using the Illumina HiSeq 2000 technology showed a total of 99,074,309 high quality raw reads (~20 Gb). Based on sequence homology onion genes were categorized into 49 different functional groups. Most of the genes however, were classified under 'unknown' in all three gene ontology categories. Of the categorized genes, 61.2% showed metabolic functions followed by cellular components such as binding, cellular processes; catalytic activity and cell part. With BLASTx top hit analysis, a total of 2,511 homologous allergenic sequences were found, which had 37-100% similarity with 46 different types of allergens existing in the database. From the 46 contigs or allergens, 521 B-cell linear epitopes were identified using BepiPred linear epitope prediction tool. This is the first comprehensive insight into the transcriptome of onion bulb tissue using the NGS technology, which can be used to map IgE epitopes and prediction of structures and functions of various proteins.

  15. Comparative analysis of the brain transcriptome in a hyper-aggressive fruit fly, Drosophila prolongata.

    PubMed

    Kudo, Ayumi; Shigenobu, Shuji; Kadota, Koji; Nozawa, Masafumi; Shibata, Tomoko F; Ishikawa, Yukio; Matsuo, Takashi

    2017-03-01

    Aggressive behavior is observed in many animals, but its intensity differs between species. In a model animal of genetics, Drosophila melanogaster, genetic basis of aggressive behavior has been studied intensively, including transcriptome analyses to identify genes whose expression level was associated with intra-species variation in aggressiveness. However, whether these genes are also involved in the evolution of aggressiveness among different species has not been examined. In this study, we performed de novo transcriptome analysis in the brain of Drosophila prolongata to identify genes associated with the evolution of aggressiveness. Males of D. prolongata were hyper-aggressive compared with closely related species. Comparison of the brain transcriptomes identified 21 differentially expressed genes in males of D. prolongata. They did not overlap with the list of aggression-related genes identified in D. melanogaster, suggesting that genes involved in the evolution of aggressiveness were independent of those associated with the intra-species variation in aggressiveness in Drosophila. Although females of D. prolongata were not aggressive as the males, expression levels of the 21 genes identified in this study were more similar between sexes than between species.

  16. Transcriptomic Analysis of Laribacter hongkongensis Reveals Adaptive Response Coupled with Temperature

    PubMed Central

    Kong, Hoi-Kuan; Law, Hon-Wai; Liu, Xuan; Law, Carmen O. K.; Pan, Qing; Gao, Lin; Xiong, Lifeng; Lau, Susanna K. P.; Woo, Patrick C. Y.; Lau, Terrence chi kong

    2017-01-01

    Bacterial adaptation to different hosts requires transcriptomic alteration in response to the environmental conditions. Laribacter hongkongensis is a gram-negative, facultative anaerobic, urease-positive bacillus caused infections in liver cirrhosis patients and community-acquired gastroenteritis. It was also found in intestine from commonly consumed freshwater fishes and drinking water reservoirs. Since L. hongkongensis could survive as either fish or human pathogens, their survival mechanisms in two different habitats should be temperature-regulated and highly complex. Therefore, we performed transcriptomic analysis of L. hongkongensis at body temperatures of fish and human in order to elucidate the versatile adaptation mechanisms coupled with the temperatures. We identified numerous novel temperature-induced pathways involved in host pathogenesis, in addition to the shift of metabolic equilibriums and overexpression of stress-related proteins. Moreover, these pathways form a network that can be activated at a particular temperature, and change the physiology of the bacteria to adapt to the environments. In summary, the dynamic of transcriptomes in L. hongkongensis provides versatile strategies for the bacterial survival at different habitats and this alteration prepares the bacterium for the challenge of host immunity. PMID:28085929

  17. De Novo Transcriptome Analysis of Allium cepa L. (Onion) Bulb to Identify Allergens and Epitopes

    PubMed Central

    Rajkumar, Hemalatha; Ramagoni, Ramesh Kumar; Anchoju, Vijayendra Chary; Vankudavath, Raju Naik; Syed, Arshi Uz Zaman

    2015-01-01

    Allium cepa (onion) is a diploid plant with one of the largest nuclear genomes among all diploids. Onion is an example of an under-researched crop which has a complex heterozygous genome. There are no allergenic proteins and genomic data available for onions. This study was conducted to establish a transcriptome catalogue of onion bulb that will enable us to study onion related genes involved in medicinal use and allergies. Transcriptome dataset generated from onion bulb using the Illumina HiSeq 2000 technology showed a total of 99,074,309 high quality raw reads (~20 Gb). Based on sequence homology onion genes were categorized into 49 different functional groups. Most of the genes however, were classified under 'unknown' in all three gene ontology categories. Of the categorized genes, 61.2% showed metabolic functions followed by cellular components such as binding, cellular processes; catalytic activity and cell part. With BLASTx top hit analysis, a total of 2,511 homologous allergenic sequences were found, which had 37–100% similarity with 46 different types of allergens existing in the database. From the 46 contigs or allergens, 521 B-cell linear epitopes were identified using BepiPred linear epitope prediction tool. This is the first comprehensive insight into the transcriptome of onion bulb tissue using the NGS technology, which can be used to map IgE epitopes and prediction of structures and functions of various proteins. PMID:26284934

  18. Transcriptome analysis of leaf tissue of Raphanus sativus by RNA sequencing.

    PubMed

    Zhang, Libin; Jia, Haibo; Yin, Yongtai; Wu, Gang; Xia, Heng; Wang, Xiaodong; Fu, Chunhua; Li, Maoteng; Wu, Jiangsheng

    2013-01-01

    Raphanus sativus is not only a popular edible vegetable but also an important source of medicinal compounds. However, the paucity of knowledge about the transcriptome of R. sativus greatly impedes better understanding of the functional genomics and medicinal potential of R. sativus. In this study, the transcriptome sequencing of leaf tissues in R. sativus was performed for the first time. Approximately 22 million clean reads were generated and used for transcriptome assembly. The generated unigenes were subsequently annotated against gene ontology (GO) database. KEGG analysis further revealed two important pathways in the bolting stage of R.sativus including spliceosome assembly and alkaloid synthesis. In addition, a total of 6,295 simple sequence repeats (SSRs) with various motifs were identified in the unigene library of R. sativus. Finally, four unigenes of R. sativus were selected for alignment with their homologs from other plants, and phylogenetic trees for each of the genes were constructed. Taken together, this study will provide a platform to facilitate gene discovery and advance functional genomic research of R. sativus.

  19. Multiplexed transcriptome analysis to detect ALK, ROS1 and RET rearrangements in lung cancer

    PubMed Central

    Rogers, Toni-Maree; Arnau, Gisela Mir; Ryland, Georgina L.; Huang, Stephen; Lira, Maruja E.; Emmanuel, Yvette; Perez, Omar D.; Irwin, Darryl; Fellowes, Andrew P.; Wong, Stephen Q.; Fox, Stephen B.

    2017-01-01

    ALK, ROS1 and RET gene fusions are important predictive biomarkers for tyrosine kinase inhibitors in lung cancer. Currently, the gold standard method for gene fusion detection is Fluorescence In Situ Hybridization (FISH) and while highly sensitive and specific, it is also labour intensive, subjective in analysis, and unable to screen a large numbers of gene fusions. Recent developments in high-throughput transcriptome-based methods may provide a suitable alternative to FISH as they are compatible with multiplexing and diagnostic workflows. However, the concordance between these different methods compared with FISH has not been evaluated. In this study we compared the results from three transcriptome-based platforms (Nanostring Elements, Agena LungFusion panel and ThermoFisher NGS fusion panel) to those obtained from ALK, ROS1 and RET FISH on 51 clinical specimens. Overall agreement of results ranged from 86–96% depending on the platform used. While all platforms were highly sensitive, both the Agena panel and Thermo Fisher NGS fusion panel reported minor fusions that were not detectable by FISH. Our proof–of–principle study illustrates that transcriptome-based analyses are sensitive and robust methods for detecting actionable gene fusions in lung cancer and could provide a robust alternative to FISH testing in the diagnostic setting. PMID:28181564

  20. A novel meta-analysis approach of cancer transcriptomes reveals prevailing transcriptional networks in cancer cells.

    PubMed

    Niida, Atsushi; Imoto, Seiya; Nagasaki, Masao; Yamaguchi, Rui; Miyano, Satoru

    2010-01-01

    Although microarray technology has revealed transcriptomic diversities underlining various cancer phenotypes, transcriptional programs controlling them have not been well elucidated. To decode transcriptional programs governing cancer transcriptomes, we have recently developed a computational method termed EEM, which searches for expression modules from prescribed gene sets defined by prior biological knowledge like TF binding motifs. In this paper, we extend our EEM approach to predict cancer transcriptional networks. Starting from functional TF binding motifs and expression modules identified by EEM, we predict cancer transcriptional networks containing regulatory TFs, associated GO terms, and interactions between TF binding motifs. To systematically analyze transcriptional programs in broad types of cancer, we applied our EEM-based network prediction method to 122 microarray datasets collected from public databases. The data sets contain about 15000 experiments for tumor samples of various tissue origins including breast, colon, lung etc. This EEM based meta-analysis successfully revealed a prevailing cancer transcriptional network which functions in a large fraction of cancer transcriptomes; they include cell-cycle and immune related sub-networks. This study demonstrates broad applicability of EEM, and opens a way to comprehensive understanding of transcriptional networks in cancer cells.

  1. Transcriptome Analysis of Thapsia laciniata Rouy Provides Insights into Terpenoid Biosynthesis and Diversity in Apiaceae

    PubMed Central

    Drew, Damian Paul; Dueholm, Bjørn; Weitzel, Corinna; Zhang, Ye; Sensen, Christoph W.; Simonsen, Henrik Toft

    2013-01-01

    Thapsia laciniata Rouy (Apiaceae) produces irregular and regular sesquiterpenoids with thapsane and guaiene carbon skeletons, as found in other Apiaceae species. A transcriptomic analysis utilizing Illumina next-generation sequencing enabled the identification of novel genes involved in the biosynthesis of terpenoids in Thapsia. From 66.78 million HQ paired-end reads obtained from T. laciniata roots, 64.58 million were assembled into 76,565 contigs (N50: 1261 bp). Seventeen contigs were annotated as terpene synthases and five of these were predicted to be sesquiterpene synthases. Of the 67 contigs annotated as cytochromes P450, 18 of these are part of the CYP71 clade that primarily performs hydroxylations of specialized metabolites. Three contigs annotated as aldehyde dehydrogenases grouped phylogenetically with the characterized ALDH1 from Artemisia annua and three contigs annotated as alcohol dehydrogenases grouped with the recently described ADH1 from A. annua. ALDH1 and ADH1 were characterized as part of the artemisinin biosynthesis. We have produced a comprehensive EST dataset for T. laciniata roots, which contains a large sample of the T. laciniata transcriptome. These transcriptome data provide the foundation for future research into the molecular basis for terpenoid biosynthesis in Thapsia and on the evolution of terpenoids in Apiaceae. PMID:23698765

  2. Integrated quantitative analysis of the phosphoproteome and transcriptome in tamoxifen-resistant breast cancer.

    PubMed

    Oyama, Masaaki; Nagashima, Takeshi; Suzuki, Takashi; Kozuka-Hata, Hiroko; Yumoto, Noriko; Shiraishi, Yuichi; Ikeda, Kazuhiro; Kuroki, Yoko; Gotoh, Noriko; Ishida, Takanori; Inoue, Satoshi; Kitano, Hiroaki; Okada-Hatakeyama, Mariko

    2011-01-07

    Quantitative phosphoproteome and transcriptome analysis of ligand-stimulated MCF-7 human breast cancer cells was performed to understand the mechanisms of tamoxifen resistance at a system level. Phosphoproteome data revealed that WT cells were more enriched with phospho-proteins than tamoxifen-resistant cells after stimulation with ligands. Surprisingly, decreased phosphorylation after ligand perturbation was more common than increased phosphorylation. In particular, 17β-estradiol induced down-regulation in WT cells at a very high rate. 17β-Estradiol and the ErbB ligand heregulin induced almost equal numbers of up-regulated phospho-proteins in WT cells. Pathway and motif activity analyses using transcriptome data additionally suggested that deregulated activation of GSK3β (glycogen-synthase kinase 3β) and MAPK1/3 signaling might be associated with altered activation of cAMP-responsive element-binding protein and AP-1 transcription factors in tamoxifen-resistant cells, and this hypothesis was validated by reporter assays. An examination of clinical samples revealed that inhibitory phosphorylation of GSK3β at serine 9 was significantly lower in tamoxifen-treated breast cancer patients that eventually had relapses, implying that activation of GSK3β may be associated with the tamoxifen-resistant phenotype. Thus, the combined phosphoproteome and transcriptome data set analyses revealed distinct signal transcription programs in tumor cells and provided a novel molecular target to understand tamoxifen resistance.

  3. Transcriptome Sequencing and De Novo Analysis of the Copepod Calanus sinicus Using 454 GS FLX

    PubMed Central

    Ning, Juan; Wang, Minxiao; Li, Chaolun; Sun, Song

    2013-01-01

    Background Despite their species abundance and primary economic importance, genomic information about copepods is still limited. In particular, genomic resources are lacking for the copepod Calanus sinicus, which is a dominant species in the coastal waters of East Asia. In this study, we performed de novo transcriptome sequencing to produce a large number of expressed sequence tags for the copepod C. sinicus. Results Copepodid larvae and adults were used as the basic material for transcriptome sequencing. Using 454 pyrosequencing, a total of 1,470,799 reads were obtained, which were assembled into 56,809 high quality expressed sequence tags. Based on their sequence similarity to known proteins, about 14,000 different genes were identified, including members of all major conserved signaling pathways. Transcripts that were putatively involved with growth, lipid metabolism, molting, and diapause were also identified among these genes. Differentially expressed genes related to several processes were found in C. sinicus copepodid larvae and adults. We detected 284,154 single nucleotide polymorphisms (SNPs) that provide a resource for gene function studies. Conclusion Our data provide the most comprehensive transcriptome resource available for C. sinicus. This resource allowed us to identify genes associated with primary physiological processes and SNPs in coding regions, which facilitated the quantitative analysis of differential gene expression. These data should provide foundation for future genetic and genomic studies of this and related species. PMID:23671698

  4. De Novo Transcriptome Analysis and Detection of Antimicrobial Peptides of the American Cockroach Periplaneta americana (Linnaeus)

    PubMed Central

    Subramaniyam, Sathiyamoorthy; Yun, Eun-Young; Kim, Iksoo; Park, Junhyung; Hwang, Jae Sam

    2016-01-01

    Cockroaches are surrogate hosts for microbes that cause many human diseases. In spite of their generally destructive nature, cockroaches have recently been found to harbor potentially beneficial and medically useful substances such as drugs and allergens. However, genomic information for the American cockroach (Periplaneta americana) is currently unavailable; therefore, transcriptome and gene expression profiling is needed as an important resource to better understand the fundamental biological mechanisms of this species, which would be particularly useful for the selection of novel antimicrobial peptides. Thus, we performed de novo transcriptome analysis of P. americana that were or were not immunized with Escherichia coli. Using an Illumina HiSeq sequencer, we generated a total of 9.5 Gb of sequences, which were assembled into 85,984 contigs and functionally annotated using Basic Local Alignment Search Tool (BLAST), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) database terms. Finally, using an in silico antimicrobial peptide prediction method, 86 antimicrobial peptide candidates were predicted from the transcriptome, and 21 of these peptides were experimentally validated for their antimicrobial activity against yeast and gram positive and -negative bacteria by a radial diffusion assay. Notably, 11 peptides showed strong antimicrobial activities against these organisms and displayed little or no cytotoxic effects in the hemolysis and cell viability assay. This work provides prerequisite baseline data for the identification and development of novel antimicrobial peptides, which is expected to provide a better understanding of the phenomenon of innate immunity in similar species. PMID:27167617

  5. Multiplexed transcriptome analysis to detect ALK, ROS1 and RET rearrangements in lung cancer.

    PubMed

    Rogers, Toni-Maree; Arnau, Gisela Mir; Ryland, Georgina L; Huang, Stephen; Lira, Maruja E; Emmanuel, Yvette; Perez, Omar D; Irwin, Darryl; Fellowes, Andrew P; Wong, Stephen Q; Fox, Stephen B

    2017-02-09

    ALK, ROS1 and RET gene fusions are important predictive biomarkers for tyrosine kinase inhibitors in lung cancer. Currently, the gold standard method for gene fusion detection is Fluorescence In Situ Hybridization (FISH) and while highly sensitive and specific, it is also labour intensive, subjective in analysis, and unable to screen a large numbers of gene fusions. Recent developments in high-throughput transcriptome-based methods may provide a suitable alternative to FISH as they are compatible with multiplexing and diagnostic workflows. However, the concordance between these different methods compared with FISH has not been evaluated. In this study we compared the results from three transcriptome-based platforms (Nanostring Elements, Agena LungFusion panel and ThermoFisher NGS fusion panel) to those obtained from ALK, ROS1 and RET FISH on 51 clinical specimens. Overall agreement of results ranged from 86-96% depending on the platform used. While all platforms were highly sensitive, both the Agena panel and Thermo Fisher NGS fusion panel reported minor fusions that were not detectable by FISH. Our proof-of-principle study illustrates that transcriptome-based analyses are sensitive and robust methods for detecting actionable gene fusions in lung cancer and could provide a robust alternative to FISH testing in the diagnostic setting.

  6. Transcriptome profile analysis of adipose tissues from fat and short-tailed sheep.

    PubMed

    Wang, Xiaolong; Zhou, Guangxian; Xu, Xiaochun; Geng, Rongqing; Zhou, Jiping; Yang, Yuxin; Yang, Zhaoxia; Chen, Yulin

    2014-10-10

    Recent studies in domestic animals have used RNA-seq to explore the transcriptome of different tissues in a limited number of individuals. In the present study, de novo transcriptome sequencing was used to compare sheep adipose tissue transcriptome profiles between a fat-tailed breed (Kazak sheep; KS) and a short-tailed (Tibetan sheep; TS). The RNA-seq data from these two groups revealed that 646 genes were differentially expressed between the KS and TS groups, including 280 up-regulated and 366 down-regulated genes. We identified genes relevant to fat metabolism in adipose tissues, including two top genes with the largest fold change (NELL1 and FMO3). Pathway analysis revealed that the differentially expressed genes between the KS and TS breeds belong to fatty acid metabolism relevant pathways (e.g. fat digestion and absorption, glycine, serine, and threonine metabolism) and cell junction-related pathways (e.g. cell adhesion molecules) which contribute to fat deposition. This work highlighted potential genes and gene networks that affect fat deposition and meat quality in sheep. Copyright © 2014. Published by Elsevier B.V.

  7. Transcriptome Analysis of Leaf Tissue of Raphanus sativus by RNA Sequencing

    PubMed Central

    Yin, Yongtai; Wu, Gang; Xia, Heng; Wang, Xiaodong; Fu, Chunhua; Li, Maoteng; Wu, Jiangsheng

    2013-01-01

    Raphanus sativus is not only a popular edible vegetable but also an important source of medicinal compounds. However, the paucity of knowledge about the transcriptome of R. sativus greatly impedes better understanding of the functional genomics and medicinal potential of R. sativus. In this study, the transcriptome sequencing of leaf tissues in R. sativus was performed for the first time. Approximately 22 million clean reads were generated and used for transcriptome assembly. The generated unigenes were subsequently annotated against gene ontology (GO) database. KEGG analysis further revealed two important pathways in the bolting stage of R.sativus including spliceosome assembly and alkaloid synthesis. In addition, a total of 6,295 simple sequence repeats (SSRs) with various motifs were identified in the unigene library of R. sativus. Finally, four unigenes of R. sativus were selected for alignment with their homologs from other plants, and phylogenetic trees for each of the genes were constructed. Taken together, this study will provide a platform to facilitate gene discovery and advance functional genomic research of R. sativus. PMID:24265813

  8. De Novo Transcriptome Analysis and Detection of Antimicrobial Peptides of the American Cockroach Periplaneta americana (Linnaeus).

    PubMed

    Kim, In-Woo; Lee, Joon Ha; Subramaniyam, Sathiyamoorthy; Yun, Eun-Young; Kim, Iksoo; Park, Junhyung; Hwang, Jae Sam

    2016-01-01

    Cockroaches are surrogate hosts for microbes that cause many human diseases. In spite of their generally destructive nature, cockroaches have recently been found to harbor potentially beneficial and medically useful substances such as drugs and allergens. However, genomic information for the American cockroach (Periplaneta americana) is currently unavailable; therefore, transcriptome and gene expression profiling is needed as an important resource to better understand the fundamental biological mechanisms of this species, which would be particularly useful for the selection of novel antimicrobial peptides. Thus, we performed de novo transcriptome analysis of P. americana that were or were not immunized with Escherichia coli. Using an Illumina HiSeq sequencer, we generated a total of 9.5 Gb of sequences, which were assembled into 85,984 contigs and functionally annotated using Basic Local Alignment Search Tool (BLAST), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) database terms. Finally, using an in silico antimicrobial peptide prediction method, 86 antimicrobial peptide candidates were predicted from the transcriptome, and 21 of these peptides were experimentally validated for their antimicrobial activity against yeast and gram positive and -negative bacteria by a radial diffusion assay. Notably, 11 peptides showed strong antimicrobial activities against these organisms and displayed little or no cytotoxic effects in the hemolysis and cell viability assay. This work provides prerequisite baseline data for the identification and development of novel antimicrobial peptides, which is expected to provide a better understanding of the phenomenon of innate immunity in similar species.

  9. De Novo Characterization of the Mung Bean Transcriptome and Transcriptomic Analysis of Adventitious Rooting in Seedlings Using RNA-Seq

    PubMed Central

    Li, Shi-Weng; Shi, Rui-Fang; Leng, Yan

    2015-01-01

    Adventitious rooting is the most important mechanism underlying vegetative propagation and an important strategy for plant propagation under environmental stress. The present study was conducted to obtain transcriptomic data and examine gene expression using RNA-Seq and bioinformatics analysis, thereby providing a foundation for understanding the molecular mechanisms controlling adventitious rooting. Three cDNA libraries constructed from mRNA samples from mung bean hypocotyls during adventitious rooting were sequenced. These three samples generated a total of 73 million, 60 million, and 59 million 100-bp reads, respectively. These reads were assembled into 78,697 unigenes with an average length of 832 bp, totaling 65 Mb. The unigenes were aligned against six public protein databases, and 29,029 unigenes (36.77%) were annotated using BLASTx. Among them, 28,225 (35.75%) and 28,119 (35.62%) unigenes had homologs in the TrEMBL and NCBI non-redundant (Nr) databases, respectively. Of these unigenes, 21,140 were assigned to gene ontology classes, and a total of 11,990 unigenes were classified into 25 KOG functional categories. A total of 7,357 unigenes were annotated to 4,524 KOs, and 4,651 unigenes were mapped onto 342 KEGG pathways using BLAST comparison against the KEGG database. A total of 11,717 unigenes were differentially expressed (fold change>2) during the root induction stage, with 8,772 unigenes down-regulated and 2,945 unigenes up-regulated. A total of 12,737 unigenes were differentially expressed during the root initiation stage, with 9,303 unigenes down-regulated and 3,434 unigenes up-regulated. A total of 5,334 unigenes were differentially expressed between the root induction and initiation stage, with 2,167 unigenes down-regulated and 3,167 unigenes up-regulated. qRT-PCR validation of the 39 genes with known functions indicated a strong correlation (92.3%) with the RNA-Seq data. The GO enrichment, pathway mapping, and gene expression profiles reveal

  10. De novo assembly of a tadpole shrimp (Triops newberryi) transcriptome and preliminary differential gene expression analysis.

    PubMed

    Horn, Rebekah L; Ramaraj, Thiruvarangan; Devitt, Nicholas P; Schilkey, Faye D; Cowley, David E

    2017-03-01

    Next-generation sequencing techniques, such as RNA sequencing, have provided a wealth of genomic information for nonmodel species. Transcriptomic information can be used to quantify the patterns of gene expression, which can identify how environmental differences invoke organismal stress responses and provide a gauge in predicting species adaptability. In our study, we used RNA sequencing to characterize the first transcriptome from a naupliar tadpole shrimp (Triops newberryi) to identify the genes expressed during the early life history stages and which could be important for future genomic studies. RNA was extracted from naupliar T. newberryi that were reared in a laboratory-controlled setting and in two different water types, a native and a non-native condition. A total of six replicates, three per condition, were sequenced with the Illumina Hi-Seq 2000 achieving 365 M 50-nt reads. High-quality reads were produced and de novo assembly was used to construct a T. newberryi transcriptome that was approximately 24.8 M base pairs. More than 10 000 peptides were predicted from the assembly, and genes were sorted into gene ontology categories. The use of different water conditions allowed for a preliminary differential gene expression analysis in order to compare the changes in gene expression between conditions. There were 299 differentially expressed genes between water conditions that might serve as a focal point for future genomic studies of Triops acclimation to different environments. The Triops transcriptome could serve as vital genomic information for additional studies on Branchiopod crustaceans. © 2016 John Wiley & Sons Ltd.

  11. Differential transcriptome analysis of glandular and filamentous trichomes in Artemisia annua

    PubMed Central

    2013-01-01

    Background The medicinal plant Artemisia annua is covered with filamentous trichomes and glandular, artemisinin producing trichomes. A high artemisinin supply is needed at a reduced cost for treating malaria. Artemisinin production in bioreactors can be facilitated if a better insight is obtained in the biosynthesis of artemisinin and other metabolites. Therefore, metabolic activities of glandular and filamentous trichomes were investigated at the transcriptome level. Results By laser pressure catapulting, glandular and filamentous trichomes as well as apical and sub-apical cells from glandular trichomes were collected and their transcriptome was sequenced using Illumina RNA-Seq. A de novo transcriptome was assembled (Trinity) and studied with a differential expression analysis (edgeR). A comparison of the transcriptome from glandular and filamentous trichomes shows that MEP, MVA, most terpene and lipid biosynthesis pathways are significantly upregulated in glandular trichomes. Conversely, some transcripts coding for specific sesquiterpenoid and triterpenoid enzymes such as 8-epi-cedrol synthase and an uncharacterized oxidosqualene cyclase were significantly upregulated in filamentous trichomes. All known artemisinin biosynthesis genes are upregulated in glandular trichomes and were detected in both the apical and sub-apical cells of the glandular trichomes. No significant differential expression could be observed between the apical and sub-apical cells. Conclusions Our results underscore the vast metabolic capacities of A. annua glandular trichomes but nonetheless point to the existence of specific terpene metabolic pathways in the filamentous trichomes. Candidate genes that might be involved in artemisinin biosynthesis are proposed based on their putative function and their differential expression level. PMID:24359620

  12. Transcriptome-based analysis of the saprophytic fungus Abortiporus biennis - response to oxalic acid.

    PubMed

    Grąz, Marcin; Jarosz-Wilkołazka, Anna; Janusz, Grzegorz; Mazur, Andrzej; Wielbo, Jerzy; Koper, Piotr; Żebracki, Kamil; Kubik-Komar, Agnieszka

    2017-06-01

    In this study, the transcriptomic-based response of the white rot fungus Abortiporus biennis to oxalic acid induction was reported. The whole transcriptome of A. biennis was analysed using the RNA-based sequencing technology and Solid 5500 platform. De novo assembly of reads generated 37,719 contigs. A molecular function for 26,280 unique transcripts was assigned. The analysis of the A. biennis transcriptome predicted 635 hypothetical open reading frames encoding carbohydrate active enzymes distributed in 122 families. 82 genes were identified, whose expression level was significantly changed after oxalic acid addition. Among them, 18 genes were up-regulated and 64 genes were down-regulated. Genes coding for putative cellulose and hemicellulose degrading enzymes were predominantly up-regulated in the mycelium induced with oxalic acid; it was in the case of cellulases and xylanases (hemicellulases), in particular, β-glucosidase and endo-1,4-β-xylanases. On the contrary, several genes coding for lignolytic enzymes were down-regulated, with the significant repression level in the case of versatile peroxidase. Finally, we identified putative genes involved in oxalate metabolism. Among the transcripts detected in the A. biennis transcriptome, one was annotated as coding for putative oxalate decarboxylase (ODC) and nine transcripts were annotated as formate dehydrogenases (FDH). The addition of oxalic acid to the culture caused upregulation of the gene coding for ODC and three genes for FDH. Amongst the transcripts of putative FDH genes, one designated as NODE_36057, demonstrated the highest induction level recorded in this study after the oxalic acid addition. Copyright © 2017 Elsevier GmbH. All rights reserved.

  13. Integrative Analysis of Transcriptomic and Epigenomic Data to Reveal Regulation Patterns for BMD Variation.

    PubMed

    Zhang, Ji-Gang; Tan, Li-Jun; Xu, Chao; He, Hao; Tian, Qing; Zhou, Yu; Qiu, Chuan; Chen, Xiang-Ding; Deng, Hong-Wen

    2015-01-01

    Integration of multiple profiling data and construction of functional gene networks may provide additional insights into the molecular mechanisms of complex diseases. Osteoporosis is a worldwide public health problem, but the complex gene-gene interactions, post-transcriptional modifications and regulation of functional networks are still unclear. To gain a comprehensive understanding of osteoporosis etiology, transcriptome gene expression microarray, epigenomic miRNA microarray and methylome sequencing were performed simultaneously in 5 high hip BMD (Bone Mineral Density) subjects and 5 low hip BMD subjects. SPIA (Signaling Pathway Impact Analysis) and PCST (Prize Collecting Steiner Tree) algorithm were used to perform pathway-enrichment analysis and construct the interaction networks. Through integrating the transcriptomic and epigenomic data, firstly we identified 3 genes (FAM50A, ZNF473 and TMEM55B) and one miRNA (hsa-mir-4291) which showed the consistent association evidence from both gene expression and methylation data; secondly in network analysis we identified an interaction network module with 12 genes and 11 miRNAs including AKT1, STAT3, STAT5A, FLT3, hsa-mir-141 and hsa-mir-34a which have been associated with BMD in previous studies. This module revealed the crosstalk among miRNAs, mRNAs and DNA methylation and showed four potential regulatory patterns of gene expression to influence the BMD status. In conclusion, the integration of multiple layers of omics can yield in-depth results than analysis of individual omics data respectively. Integrative analysis from transcriptomics and epigenomic data improves our ability to identify causal genetic factors, and more importantly uncover functional regulation pattern of multi-omics for osteoporosis etiology.

  14. Transcriptome analysis reveals manifold mechanisms of cyst development in ADPKD.

    PubMed

    de Almeida, Rita M C; Clendenon, Sherry G; Richards, William G; Boedigheimer, Michael; Damore, Michael; Rossetti, Sandro; Harris, Peter C; Herbert, Britney-Shea; Xu, Wei Min; Wandinger-Ness, Angela; Ward, Heather H; Glazier, James A; Bacallao, Robert L

    2016-11-21

    Autosomal dominant polycystic kidney disease (ADPKD) causes progressive loss of renal function in adults as a consequence of the accumulation of cysts. ADPKD is the most common genetic cause of end-stage renal disease. Mutations in polycystin-1 occur in 87% of cases of ADPKD and mutations in polycystin-2 are found in 12% of ADPKD patients. The complexity of ADPKD has hampered efforts to identify the mechanisms underlying its pathogenesis. No current FDA (Federal Drug Administration)-approved therapies ameliorate ADPKD progression. We used the de Almeida laboratory's sensitive new transcriptogram method for whole-genome gene expression data analysis to analyze microarray data from cell lines developed from cell isolates of normal kidney and of both non-cystic nephrons and cysts from the kidney of a patient with ADPKD. We compared results obtained using standard Ingenuity Volcano plot analysis, Gene Set Enrichment Analysis (GSEA) and transcriptogram analysis. Transcriptogram analysis confirmed the findings of Ingenuity, GSEA, and published analysis of ADPKD kidney data and also identified multiple new expression changes in KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways related to cell growth, cell death, genetic information processing, nucleotide metabolism, signal transduction, immune response, response to stimulus, cellular processes, ion homeostasis and transport and cofactors, vitamins, amino acids, energy, carbohydrates, drugs, lipids, and glycans. Transcriptogram analysis also provides significance metrics which allow us to prioritize further study of these pathways. Transcriptogram analysis identifies novel pathways altered in ADPKD, providing new avenues to identify both ADPKD's mechanisms of pathogenesis and pharmaceutical targets to ameliorate the progression of the disease.

  15. Effect of rosemary polyphenols on human colon cancer cells: transcriptomic profiling and functional enrichment analysis.

    PubMed

    Valdés, Alberto; García-Cañas, Virginia; Rocamora-Reverte, Lourdes; Gómez-Martínez, Angeles; Ferragut, José Antonio; Cifuentes, Alejandro

    2013-01-01

    In this work, the effect of rosemary extracts rich on polyphenols obtained using pressurized fluids was investigated on the gene expression of human SW480 and HT29 colon cancer cells. The application of transcriptomic profiling and functional enrichment analysis was done via two computational approaches, Ingenuity Pathway Analysis and Gene Set Enrichment Analysis. These two approaches were used for functional enrichment analysis as a previous step for a reliable interpretation of the data obtained from microarray analysis. Reverse transcription quantitative-PCR was used to confirm relative changes in mRNA levels of selected genes from microarrays. The selection of genes was based on their expression change, adjusted p value, and known biological function. According to genome-wide transcriptomics analysis, rosemary polyphenols altered the expression of ~4 % of the genes covered by the Affymetrix Human Gene 1.0ST chip in both colon cancer cells. However, only ~18 % of the differentially expressed genes were common to both cell lines, indicating markedly different expression profiles in response to the treatment. Differences in induction of G2/M arrest observed by rosemary polyphenols in the two colon adenocarcinoma cell lines suggest that the extract may be differentially effective against tumors with specific mutational pattern. From our results, it is also concluded that rosemary polyphenols induced a low degree of apoptosis indicating that other multiple signaling pathways may contribute to colon cancer cell death.

  16. Transcriptome Analysis of Sunflower Genotypes with Contrasting Oxidative Stress Tolerance Reveals Individual- and Combined- Biotic and Abiotic Stress Tolerance Mechanisms.

    PubMed

    Ramu, Vemanna S; Paramanantham, Anjugam; Ramegowda, Venkategowda; Mohan-Raju, Basavaiah; Udayakumar, Makarla; Senthil-Kumar, Muthappa

    2016-01-01

    In nature plants are often simultaneously challenged by different biotic and abiotic stresses. Although the mechanisms underlying plant responses against single stress have been studied considerably, plant tolerance mechanisms under combined stress is not understood. Also, the mechanism used to combat independently and sequentially occurring many number of biotic and abiotic stresses has also not systematically studied. From this context, in this study, we attempted to explore the shared response of sunflower plants to many independent stresses by using meta-analysis of publically available transcriptome data and transcript profiling by quantitative PCR. Further, we have also analyzed the possible role of the genes so identified in contributing to combined stress tolerance. Meta-analysis of transcriptomic data from many abiotic and biotic stresses indicated the common representation of oxidative stress responsive genes. Further, menadione-mediated oxidative stress in sunflower seedlings showed similar pattern of changes in the oxidative stress related genes. Based on this a large scale screening of 55 sunflower genotypes was performed under menadione stress and those contrasting in oxidative stress tolerance were identified. Further to confirm the role of genes identified in individual and combined stress tolerance the contrasting genotypes were individually and simultaneously challenged with few abiotic and biotic stresses. The tolerant hybrid showed reduced levels of stress damage both under combined stress and few independent stresses. Transcript profiling of the genes identified from meta-analysis in the tolerant hybrid also indicated that the selected genes were up-regulated under individual and combined stresses. Our results indicate that menadione-based screening can identify genotypes not only tolerant to multiple number of individual biotic and abiotic stresses, but also the combined stresses.

  17. Transcriptome Analysis of Sunflower Genotypes with Contrasting Oxidative Stress Tolerance Reveals Individual- and Combined- Biotic and Abiotic Stress Tolerance Mechanisms

    PubMed Central

    Ramu, Vemanna S.; Paramanantham, Anjugam; Ramegowda, Venkategowda; Mohan-Raju, Basavaiah; Udayakumar, Makarla

    2016-01-01

    In nature plants are often simultaneously challenged by different biotic and abiotic stresses. Although the mechanisms underlying plant responses against single stress have been studied considerably, plant tolerance mechanisms under combined stress is not understood. Also, the mechanism used to combat independently and sequentially occurring many number of biotic and abiotic stresses has also not systematically studied. From this context, in this study, we attempted to explore the shared response of sunflower plants to many independent stresses by using meta-analysis of publically available transcriptome data and transcript profiling by quantitative PCR. Further, we have also analyzed the possible role of the genes so identified in contributing to combined stress tolerance. Meta-analysis of transcriptomic data from many abiotic and biotic stresses indicated the common representation of oxidative stress responsive genes. Further, menadione-mediated oxidative stress in sunflower seedlings showed similar pattern of changes in the oxidative stress related genes. Based on this a large scale screening of 55 sunflower genotypes was performed under menadione stress and those contrasting in oxidative stress tolerance were identified. Further to confirm the role of genes identified in individual and combined stress tolerance the contrasting genotypes were individually and simultaneously challenged with few abiotic and biotic stresses. The tolerant hybrid showed reduced levels of stress damage both under combined stress and few independent stresses. Transcript profiling of the genes identified from meta-analysis in the tolerant hybrid also indicated that the selected genes were up-regulated under individual and combined stresses. Our results indicate that menadione-based screening can identify genotypes not only tolerant to multiple number of individual biotic and abiotic stresses, but also the combined stresses. PMID:27314499

  18. Transcriptome analysis of feline infectious peritonitis virus infection.

    PubMed

    Mehrbod, Parvaneh; Harun, Mohammad Syamsul Reza; Shuid, Ahmad Naqib; Omar, Abdul Rahman

    2015-01-01

    Feline infectious peritonitis (FIP) is a lethal systemic disease caused by FIP virus (FIPV). There are no effective vaccines or treatment available, and the virus virulence determinants and pathogenesis are not fully understood. Here, we describe the sequencing of RNA extracted from Crandell Rees Feline Kidney (CRFK) cells infected with FIPV using the Illumina next-generation sequencing approach. Bioinformatics analysis, based on Felis catus 2X annotated shotgun reference genome, using CLC bio Genome Workbench is used to map both control and infected cells. Kal's Z test statistical analysis is used to analyze the differentially expressed genes from the infected CRFK cells. In addition, RT-qPCR analysis is used for further transcriptional profiling of selected genes in infected CRFK cells and Peripheral Blood Mononuclear Cells (PBMCs) from healthy and FIP-diagnosed cats.

  19. Transcriptome analysis of epithelial and stromal contributions to mammogenesis in three week prepartum cows.

    PubMed

    Casey, Theresa; Dover, Heather; Liesman, James; DeVries, Lindsey; Kiupel, Matti; Vandehaar, Michael; Plaut, Karen

    2011-01-01

    Transcriptome analysis of bovine mammary development has provided insight into regulation of mammogenesis. However, previous studies primarily examined expression of epithelial and stromal tissues combined, and consequently did not account for tissue specific contribution to mammary development. Our objective was to identify differences in gene expression in epithelial and intralobular stromal compartments. Tissue was biopsied from non-lactating dairy cows 3 weeks prepartum, cut into explants and incubated for 2 hr with insulin and hydrocortisone. Epithelial and intralobular stromal tissues were isolated with laser capture microdissection. Global gene expression was measured with Bovine Affymetrix GeneChips, and data were preprocessed using RMA method. Moderated t-tests from gene-specific linear model analysis with cell type as a fixed effect showed more than 3,000 genes were differentially expressed between tissues (P<0.05; FDR<0.17). Analysis of epithelial and stromal transcriptomes using Database for Annotation, Visualization and Integrated Discovery (DAVID) and Ingenuity Pathways Analysis (IPA) showed that epithelial and stromal cells contributed distinct molecular signatures. Epithelial signatures were enriched with gene sets for protein synthesis, metabolism and secretion. Stromal signatures were enriched with genes that encoded molecules important to signaling, extracellular matrix composition and remodeling. Transcriptome differences also showed evidence for paracrine interactions between tissues in stimulation of IGF1 signaling pathway, stromal reaction, angiogenesis, neurogenesis, and immune response. Molecular signatures point to the dynamic role the stroma plays in prepartum mammogenesis and highlight the importance of examining the roles of cell types within the mammary gland when targeting therapies and studying mechanisms that affect milk production.

  20. Transcriptome Analysis of Epithelial and Stromal Contributions to Mammogenesis in Three Week Prepartum Cows

    PubMed Central

    Casey, Theresa; Dover, Heather; Liesman, James; DeVries, Lindsey; Kiupel, Matti; VandeHaar, Michael; Plaut, Karen

    2011-01-01

    Transcriptome analysis of bovine mammary development has provided insight into regulation of mammogenesis. However, previous studies primarily examined expression of epithelial and stromal tissues combined, and consequently did not account for tissue specific contribution to mammary development. Our objective was to identify differences in gene expression in epithelial and intralobular stromal compartments. Tissue was biopsied from non-lactating dairy cows 3 weeks prepartum, cut into explants and incubated for 2 hr with insulin and hydrocortisone. Epithelial and intralobular stromal tissues were isolated with laser capture microdissection. Global gene expression was measured with Bovine Affymetrix GeneChips, and data were preprocessed using RMA method. Moderated t-tests from gene-specific linear model analysis with cell type as a fixed effect showed more than 3,000 genes were differentially expressed between tissues (P<0.05; FDR<0.17). Analysis of epithelial and stromal transcriptomes using Database for Annotation, Visualization and Integrated Discovery (DAVID) and Ingenuity Pathways Analysis (IPA) showed that epithelial and stromal cells contributed distinct molecular signatures. Epithelial signatures were enriched with gene sets for protein synthesis, metabolism and secretion. Stromal signatures were enriched with genes that encoded molecules important to signaling, extracellular matrix composition and remodeling. Transcriptome differences also showed evidence for paracrine interactions between tissues in stimulation of IGF1 signaling pathway, stromal reaction, angiogenesis, neurogenesis, and immune response. Molecular signatures point to the dynamic role the stroma plays in prepartum mammogenesis and highlight the importance of examining the roles of cell types within the mammary gland when targeting therapies and studying mechanisms that affect milk production. PMID:21829467

  1. Next-generation sequencing facilitates quantitative analysis of wild-type and Nrl(-/-) retinal transcriptomes.

    PubMed

    Brooks, Matthew J; Rajasimha, Harsha K; Roger, Jerome E; Swaroop, Anand

    2011-01-01

    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. 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. 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 (R(2)) 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. Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA

  2. 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.

  3. Rice transcriptome analysis to identify possible herbicide quinclorac detoxification genes

    PubMed Central

    Xu, Wenying; Di, Chao; Zhou, Shaoxia; Liu, Jia; Li, Li; Liu, Fengxia; Yang, Xinling; Ling, Yun; Su, Zhen

    2015-01-01

    Quinclorac is a highly selective auxin-type herbicide and is widely used in the effective control of barnyard grass in paddy rice fields, improving the world's rice yield. The herbicide mode of action of quinclorac has been proposed, and hormone interactions affecting quinclorac signaling has been identified. Because of widespread use, quinclorac may be transported outside rice fields with the drainage waters, leading to soil and water pollution and other environmental health problems. In this study, we used 57K Affymetrix rice whole-genome array to identify quinclorac signaling response genes to study the molecular mechanisms of action and detoxification of quinclorac in rice plants. Overall, 637 probe sets were identified with differential expression levels under either 6 or 24 h of quinclorac treatment. Auxin-related genes such as GH3 and OsIAAs responded to quinclorac treatment. Gene Ontology analysis showed that genes of detoxification-related family genes were significantly enriched, including cytochrome P450, GST, UGT, and ABC and drug transporter genes. Moreover, real-time RT-PCR analysis showed that top candidate genes of P450 families such as CYP81, CYP709C, and CYP72A were universally induced by different herbicides. Some Arabidopsis genes of the same P450 family were up-regulated under quinclorac treatment. We conducted rice whole-genome GeneChip analysis and the first global identification of quinclorac response genes. This work may provide potential markers for detoxification of quinclorac and biomonitors of environmental chemical pollution. PMID:26483837

  4. Transcriptome analysis of Japanese pear (Pyrus pyrifolia Nakai) flower buds transitioning through endodormancy.

    PubMed

    Bai, Songling; Saito, Takanori; Sakamoto, Daisuke; Ito, Akiko; Fujii, Hiroshi; Moriguchi, Takaya

    2013-07-01

    The transcriptomes of endodormant and ecodormant Japanese pear (Pyrus pyrifolia Nakai 'Kosui') flower buds were analyzed using RNA-seq technology and compared. Among de novo assembly of 114,191 unigenes, 76,995 unigenes were successfully annotated by BLAST searches against various databases. Gene Ontology (GO) enrichment analysis revealed that oxidoreductases were enriched in the molecular function category, a result consistent with previous observations of notable changes in hydrogen peroxide concentration during endodormancy release. In the GO categories related to biological process, the abundance of DNA methylation-related gene transcripts also significantly changed during endodormancy release, indicating the involvement of epigenetic regulation. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis also showed the changes in transcript abundance of genes involved in the metabolism of various phytohormones. Genes for both ABA and gibberellin biosynthesis were down-regulated, whereas the genes encoding their degradation enzymes were up-regulated during endodormancy release. In the ethylene pathway, 1-aminocyclopropane-1-carboxylate synthase (ACS), a gene encoding the rate-limiting enzyme for ethylene biosynthesis, was induced towards endodormancy release. All of these results indicated the involvement of phytohormones in endodormancy release. Furthermore, the expression of dormancy-associated MADS-box (DAM) genes was down-regulated concomitant with endodormancy release, although changes in the abundance of these gene transcripts were not as significant as those identified by transcriptome analysis. Consequently, characterization of the Japanese pear transcriptome during the transition from endormancy to ecodormancy will provide researchers with useful information for data mining and will facilitate further experiments on endodormancy especially in rosaceae fruit trees.

  5. A transcriptomic analysis of the adult stage of the bovine lungworm, Dictyocaulus viviparus

    PubMed Central

    2007-01-01

    Background Lungworms of the genus Dictyocaulus (family Dictyocaulidae) are parasitic nematodes of major economic importance. They cause pathological effects and clinical disease in various ruminant hosts, particularly in young animals. Dictyocaulus viviparus, called the bovine lungworm, is a major pathogen of cattle, with severe infections being fatal. In this study, we provide first insights into the transcriptome of the adult stage of D. viviparus through the analysis of expressed sequence tags (ESTs). Results Using our EST analysis pipeline, we estimate that the present dataset of 4436 ESTs is derived from 2258 genes based on cluster and comparative genomic analyses of the ESTs. Of the 2258 representative ESTs, 1159 (51.3%) had homologues in the free-living nematode C. elegans, 1174 (51.9%) in parasitic nematodes, 827 (36.6%) in organisms other than nematodes, and 863 (38%) had no significant match to any sequence in the current databases. Of the C. elegans homologues, 569 had observed 'non-wildtype' RNAi phenotypes, including embryonic lethality, maternal sterility, sterility in progeny, larval arrest and slow growth. We could functionally classify 776 (35%) sequences using the Gene Ontologies (GO) and established pathway associations to 696 (31%) sequences in Kyoto Encyclopedia of Genes and Genomes (KEGG). In addition, we predicted 85 secreted proteins which could represent potential candidates for developing novel anthelmintics or vaccines. Conclusion The bioinformatic analyses of ESTs data for D. viviparus has elucidated sets of relatively conserved and potentially novel genes. The genes discovered in this study should assist research toward a better understanding of the basic molecular biology of D. viviparus, which could lead, in the longer term, to novel intervention strategies. The characterization of the D. viviparus transcriptome also provides a foundation for whole genome sequence analysis and future comparative transcriptomic analyses. PMID:17784965

  6. Whole transcriptome analysis of transgenic barley with altered cytokinin homeostasis and increased tolerance to drought stress.

    PubMed

    Vojta, Petr; Kokáš, Filip; Husičková, Alexandra; Grúz, Jiří; Bergougnoux, Veronique; Marchetti, Cintia F; Jiskrová, Eva; Ježilová, Eliška; Mik, Václav; Ikeda, Yoshihisa; Galuszka, Petr

    2016-09-25

    Cytokinin plant hormones have been shown to play an important role in plant response to abiotic stresses. Herein, we expand upon the findings of Pospíšilová et al. [30] regarding preparation of novel transgenic barley lines overexpressing cytokinin dehydrogenase 1 gene from Arabidopsis under the control of mild root-specific promotor of maize β-glycosidase. These lines showed drought-tolerant phenotype mainly due to alteration of root architecture and stronger lignification of root tissue. A detailed transcriptomic analysis of roots of transgenic plants subjected to revitalization after drought stress revealed attenuated response through the HvHK3 cytokinin receptor and up-regulation of two transcription factors implicated in stress responses and abscisic acid sensitivity. Increased expression of several genes involved in the phenylpropanoid pathway as well as of genes encoding arogenate dehydratase/lyase participating in phenylalanine synthesis was found in roots during revitalization. Although more precursors of lignin synthesis were present in roots after drought stress, final lignin accumulation did not change compared to that in plants grown under optimal conditions. Changes in transcriptome indicated a higher auxin turnover in transgenic roots. The same analysis in leaves revealed that genes encoding putative enzymes responsible for production of jasmonates and other volatile compounds were up-regulated. Although transgenic barley leaves showed lower chlorophyll content and down-regulation of genes encoding proteins involved in photosynthesis than did wild-type plants when cultivated under optimal conditions, they did show a tendency to return to initial photochemical activities faster than did wild-type leaves when re-watered after severe drought stress. In contrast to optimal conditions, comparative transcriptomic analysis of revitalized leaves displayed up-regulation of genes encoding enzymes and proteins involved in photosynthesis, and especially

  7. Transcriptomic Analysis of Oenococcus oeni SD-2a Response to Acid Shock by RNA-Seq.

    PubMed

    Liu, Longxiang; Zhao, Hongyu; Peng, Shuai; Wang, Tao; Su, Jing; Liang, Yanying; Li, Hua; Wang, Hua

    2017-01-01

    Oenococcus oeni can be applied to conduct malolactic fermentation (MLF), but also is the main species growing naturally in wine. Due to the high stress tolerance, it is an interesting model for investigating acid response mechanisms. In this study, the changes in the transcriptome of O.oeni SD-2a during the adaptation period have been studied. RNA-seq was introduced for the transcriptomic analysis of O. oeni samples treated with pH 4.8 and pH 3.0 at 0 and 1 h, respectively. Gene ontology (GO) and Kyoto encyclopedia of genes and genome (KEGG) were performed to compare the transcriptome data between different treatments. From GO analysis, the majority of differentially expressed genes (DEGs) (pH 3.0_1 h-VS-pH 4.8_1 h, pH 3.0_1 h-VS-pH 4.8_0 h, and pH 4.8_1 h-VS-pH 4.8_0 h) were found to be involved in the metabolic process, catalytic activity, cellular process, and binding. KEGG analysis reveals that the most functional gene categories affected by acid are membrane transport, amino acid metabolism and carbohydrate metabolism. Some genes, like the heat shock protein Hsp20, malate transporter and malate permease, were also over-expressed in response to acid stress. In addition, a considerable proportion of gene indicate a significantly different expression in this study, are novel, which needs to be investigated further. These results provide a new viewpoint and crucial resource on the acid stress response in O. oeni.

  8. Comparative transcriptome analysis of ginger variety Suprabha from two different agro-climatic zones of Odisha.

    PubMed

    Gaur, Mahendra; Das, Aradhana; Sahoo, Rajesh Kumar; Mohanty, Sujata; Joshi, Raj Kumar; Subudhi, Enketeswara

    2016-09-01

    Ginger (Zingiber officinale Rosc.), a well-known member of family Zingiberaceae, is bestowed with number of medicinal properties which is because of the secondary metabolites, essential oil and oleoresin, it contains in its rhizome. The drug yielding potential is known to depend on agro-climatic conditions prevailing at the place cultivation. Present study deals with comparative transcriptome analysis of two sample of elite ginger variety Suprabha collected from two different agro-climatic zones of Odisha. Transcriptome assembly for both the samples was done using next generation sequencing methodology. The raw data of size 10.8 and 11.8 GB obtained from analysis of two rhizomes S1Z4 and S2Z5 collected from Bhubaneswar and Koraput and are available in NCBI accession number SAMN03761169 and SAMN03761176 respectively. We identified 60,452 and 54,748 transcripts using trinity tool respectively from ginger rhizome of S1Z4 and S2Z5. The transcript length varied from 300 bp to 15,213 bp and 8988 bp and N50 value of 1415 bp and 1334 bp respectively for S1Z4 and S2Z5. To the best of our knowledge, this is the first comparative transcriptome analysis of elite ginger cultivars Suprabha from two different agro-climatic conditions of Odisha, India which will help to understand the effect of agro-climatic conditions on differential expression of secondary metabolites.

  9. Genome-wide methylation and transcriptome analysis in penile carcinoma: uncovering new molecular markers.

    PubMed

    Kuasne, Hellen; Cólus, Ilce Mara de Syllos; Busso, Ariane Fidelis; Hernandez-Vargas, Hector; Barros-Filho, Mateus Camargo; Marchi, Fabio Albuquerque; Scapulatempo-Neto, Cristovam; Faria, Eliney Ferreira; Lopes, Ademar; Guimarães, Gustavo Cardoso; Herceg, Zdenko; Rogatto, Silvia Regina

    2015-01-01

    Despite penile carcinoma (PeCa) being a relatively rare neoplasm, it remains an important public health issue for poor and developing countries. Contrary to most tumors, limited data are available for markers that are capable of assisting in diagnosis, prognosis, and treatment of PeCa. We aimed to identify molecular markers for PeCa by evaluating their epigenomic and transcriptome profiles and comparing them with surrounding non-malignant tissue (SNT) and normal glans (NG). Genome-wide methylation analysis revealed 171 hypermethylated probes in PeCa. Transcriptome profiling presented 2,883 underexpressed and 1,378 overexpressed genes. Integrative analysis revealed a panel of 54 genes with an inverse correlation between methylation and gene expression levels. Distinct methylome and transcriptome patterns were found for human papillomavirus (HPV)-positive (38.6%) and negative tumors. Interestingly, grade 3 tumors showed a distinct methylation profile when compared to grade 1. In addition, univariate analysis revealed that low BDNF methylation was associated with lymph node metastasis and shorter disease-free survival. CpG hypermethylation and gene underexpression were confirmed for a panel of genes, including TWIST1, RSOP2, SOX3, SOX17, PROM1, OTX2, HOXA3, and MEIS1. A unique methylome signature was found for PeCa compared to SNT, with aberrant DNA methylation appearing to modulate the expression of specific genes. This study describes new pathways with the potential to regulate penile carcinogenesis, including stem cell regulatory pathways and markers associated to a worse prognosis. These findings may be instrumental in the discovery and application of new genetic and epigenetic biomarkers in PeCa.

  10. The first report on transcriptome analysis of the venom gland of Iranian scorpion, Hemiscorpius lepturus.

    PubMed

    Kazemi-Lomedasht, Fatemeh; Khalaj, Vahid; Bagheri, Kamran Pooshang; Behdani, Mahdi; Shahbazzadeh, Delavar

    2017-01-01

    Hemiscorpius lepturus scorpion is one of the most venomous members of the Hemiscorpiidae family. H. lepturus is distributed in Iran, Iraq and Yemen. The prevalence and severity of scorpionism is high and health services are not able to control it. Scorpionism in Iran especially in the southern regions (Khuzestan, Sistan and Baluchestan, Hormozgan, Ilam) is one of the main health challenges. Due to the medical and health importance of scorpionism, the focus of various studies has been on the identification of H. lepturus venom components. Nevertheless, until now, only a few percent of H. lepturus venom components have been identified and there is no complete information about the venom components of H. lepturus. The current study reports transcriptome analysis of the venom gland of H. lepturus scorpion. Illumina Next Generation Sequencing results identified venom components of H. lepturus. When compared with other scorpion's venom, the venom of H. lepturus consists of mixtures of peptides, proteins and enzymes such as; phospholipases, metalloproteases, hyaluronidases, potassium channel toxins, calcium channel toxins, antimicrobial peptides (AMPs), venom proteins, venom toxins, allergens, La1-like peptides, proteases and scorpine-like peptides. Comparison of identified components of H. lepturus venom was carried out with venom components of reported scorpions and various identities and similarities between them were observed. With transcriptome analysis of H. lepturus venom unique sequences, coding venom components were investigated. Moreover, our study confirmed transcript expression of previously reported peptides; Hemitoxin, Hemicalcin and Hemilipin. The gene sequences of venom components were investigated employing transcriptome analysis of venom gland of H. lepturus. In summary, new bioactive molecules identified in this study, provide basis for venomics studies of scorpions of Hemiscorpiidae family and promises development of novel biotherapeutics

  11. Genome-scale transcriptomic insights into early-stage fruit development in woodland strawberry Fragaria vesca.

    PubMed

    Kang, Chunying; Darwish, Omar; Geretz, Aviva; Shahan, Rachel; Alkharouf, Nadim; Liu, Zhongchi

    2013-06-01

    Fragaria vesca, a diploid woodland strawberry with a small and sequenced genome, is an excellent model for studying fruit development. The strawberry fruit is unique in that the edible flesh is actually enlarged receptacle tissue. The true fruit are the numerous dry achenes dotting the receptacle's surface. Auxin produced from the achene is essential for the receptacle fruit set, a paradigm for studying crosstalk between hormone signaling and development. To investigate the molecular mechanism underlying strawberry fruit set, next-generation sequencing was employed to profile early-stage fruit development with five fruit tissue types and five developmental stages from floral anthesis to enlarged fruits. This two-dimensional data set provides a systems-level view of molecular events with precise spatial and temporal resolution. The data suggest that the endosperm and seed coat may play a more prominent role than the embryo in auxin and gibberellin biosynthesis for fruit set. A model is proposed to illustrate how hormonal signals produced in the endosperm and seed coat coordinate seed, ovary wall, and receptacle fruit development. The comprehensive fruit transcriptome data set provides a wealth of genomic resources for the strawberry and Rosaceae communities as well as unprecedented molecular insight into fruit set and early stage fruit development.

  12. Transcriptome analysis identifies novel responses and potential regulatory genes involved in seasonal dormancy transitions of leafy spurge (Euphorbia esula L.)

    PubMed Central

    Horvath, David P; Chao, Wun S; Suttle, Jeffrey C; Thimmapuram, Jyothi; Anderson, James V

    2008-01-01

    Background Dormancy of buds is a critical developmental process that allows perennial plants to survive extreme seasonal variations in climate. Dormancy transitions in underground crown buds of the model herbaceous perennial weed leafy spurge were investigated using a 23 K element cDNA microarray. These data represent the first large-scale transcriptome analysis of dormancy in underground buds of an herbaceous perennial species. Crown buds collected monthly from August through December, over a five year period, were used to monitor the changes in the transcriptome during dormancy transitions. Results Nearly 1,000 genes were differentially-expressed through seasonal dormancy transitions. Expected patterns of gene expression were observed for previously characterized genes and physiological processes indicated that resolution in our analysis was sufficient for identifying shifts in global gene expression. Conclusion Gene ontology of differentially-expressed genes suggests dormancy transitions require specific alterations in transport functions (including induction of a series of mitochondrial substrate carriers, and sugar transporters), ethylene, jasmonic acid, auxin, gibberellic acid, and abscisic acid responses, and responses to stress (primarily oxidative and cold/drought). Comparison to other dormancy microarray studies indicated that nearly half of the genes identified in our study were also differentially expressed in at least two other plant species during dormancy transitions. This comparison allowed us to identify a particular MADS-box transcription factor related to the DORMANCY ASSOCIATED MADS-BOX genes from peach and hypothesize that it may play a direct role in dormancy induction and maintenance through regulation of FLOWERING LOCUS T. PMID:19014493

  13. De novo transcriptome analysis in radish (Raphanus sativus L.) and identification of critical genes involved in bolting and flowering.

    PubMed

    Nie, Shanshan; Li, Chao; Xu, Liang; Wang, Yan; Huang, Danqiong; Muleke, Everlyne M; Sun, Xiaochuan; Xie, Yang; Liu, Liwang

    2016-05-23

    The appropriate timing of bolting and flowering is pivotal for reproductive success in Brassicaceae crops including radish (Raphanus sativus L.). Although several flowering regulatory pathways had been described in some plant species, no study on genetic networks of bolting and flowering regulation was performed in radish. In this study, to generate dataset of radish unigene sequences for large-scale gene discovery and functional pathway identification, a cDNA library from mixed radish leaves at different developmental stages was subjected to high-throughput RNA sequencing (RNA-seq). A total of 54.64 million clean reads and 111,167 contigs representing 53,642 unigenes were obtained from the radish leaf transcriptome. Among these, 50,385 unigenes were successfully annotated by BLAST searching against the public protein databases. Functional classification and annotation indicated that 42,903 and 15,382 unique sequences were assigned to 55 GO terms and 25 COG categories, respectively. KEGG pathway analysis revealed that 25,973 unigenes were classified into 128 functional pathways, among which 24 candidate genes related to plant circadian rhythm were identified. Moreover, 142 potential bolting and flowering-related genes involved in various flowering pathways were identified. In addition, seven critical bolting and flowering-related genes were isolated and profiled by T-A cloning and RT-qPCR analysis. Finally, a schematic network model of bolting and flowering regulation and pathways was put forward in radish. This study is the first report on systematic identification of bolting and flowering-related genes based on transcriptome sequencing and assembly in radish. These results could provide a foundation for further investigating bolting and flowering regulatory networks in radish, and facilitate dissecting molecular genetic mechanisms underlying bolting and flowering in Brassicaceae vegetable crops.

  14. De novo transcriptome sequencing of radish (Raphanus sativus L.) and analysis of major genes involved in glucosinolate metabolism.

    PubMed

    Wang, Yan; Pan, Yan; Liu, Zhe; Zhu, Xianwen; Zhai, Lulu; Xu, Liang; Yu, Rugang; Gong, Yiqin; Liu, Liwang

    2013-11-27

    Radish (Raphanus sativus L.), is an important root vegetable crop worldwide. Glucosinolates in the fleshy taproot significantly affect the flavor and nutritional quality of radish. However, little is known about the molecular mechanisms underlying glucosinolate metabolism in radish taproots. The limited availability of radish genomic information has greatly hindered functional genomic analysis and molecular breeding in radish. In this study, a high-throughput, large-scale RNA sequencing technology was employed to characterize the de novo transcriptome of radish roots at different stages of development. Approximately 66.11 million paired-end reads representing 73,084 unigenes with a N50 length of 1,095 bp, and a total length of 55.73 Mb were obtained. Comparison with the publicly available protein database indicates that a total of 67,305 (about 92.09% of the assembled unigenes) unigenes exhibit similarity (e -value ≤ 1.0e⁻⁵) to known proteins. The functional annotation and classification including Gene Ontology (GO), Clusters of Orthologous Group (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the main activated genes in radish taproots are predominantly involved in basic physiological and metabolic processes, biosynthesis of secondary metabolite pathways, signal transduction mechanisms and other cellular components and molecular function related terms. The majority of the genes encoding enzymes involved in glucosinolate (GS) metabolism and regulation pathways were identified in the unigene dataset by targeted searches of their annotations. A number of candidate radish genes in the glucosinolate metabolism related pathways were also discovered, from which, eight genes were validated by T-A cloning and sequencing while four were validated by quantitative RT-PCR expression profiling. The ensuing transcriptome dataset provides a comprehensive sequence resource for molecular genetics research in radish. It will serve as an

  15. Global transcriptome analysis of Halolamina sp. to decipher the salt tolerance in extremely halophilic archaea.

    PubMed

    Kurt-Kızıldoğan, Aslıhan; Abanoz, Büşra; Okay, Sezer

    2017-02-15

    Extremely halophilic archaea survive in the hypersaline environments such as salt lakes or salt mines. Therefore, these microorganisms are good sources to investigate the molecular mechanisms underlying the tolerance to high salt concentrations. In this study, a global transcriptome analysis was conducted in an extremely halophilic archaeon, Halolamina sp. YKT1, isolated from a salt mine in Turkey. A comparative RNA-seq analysis was performed using YKT1 isolate grown either at 2.7M NaCl or 5.5M NaCl concentrations. A total of 2149 genes were predicted to be up-regulated and 1638 genes were down-regulated in the presence of 5.5M NaCl. The salt tolerance of Halolamina sp. YKT1 involves the up-regulation of genes related with membrane transporters, CRISPR-Cas systems, osmoprotectant solutes, oxidative stress proteins, and iron metabolism. On the other hand, the genes encoding the proteins involved in DNA replication, transcription, translation, mismatch and nucleotide excision repair were down-regulated. The RNA-seq data were verified for seven up-regulated genes as well as six down-regulated genes via qRT-PCR analysis. This comprehensive transcriptome analysis showed that the halophilic archaeon canalizes its energy towards keeping the intracellular osmotic balance minimizing the production of nucleic acids and peptides.

  16. Genome-wide characterization of transcriptional start sites in humans by integrative transcriptome analysis

    PubMed Central

    Yamashita, Riu; Sathira, Nuankanya P.; Kanai, Akinori; Tanimoto, Kousuke; Arauchi, Takako; Tanaka, Yoshiaki; Hashimoto, Shin-ichi; Sugano, Sumio; Nakai, Kenta; Suzuki, Yutaka

    2011-01-01

    We performed a genome-wide analysis of transcriptional start sites (TSSs) in human genes by multifaceted use of a massively parallel sequencer. By analyzing 800 million sequences that were obtained from various types of transcriptome analyses, we characterized 140 million TSS tags in 12 human cell types. Despite the large number of TSS clusters (TSCs), the number of TSCs was observed to decrease sharply with increasing expression levels. Highly expressed TSCs exhibited several characteristic features: Nucleosome-seq analysis revealed highly ordered nucleosome structures, ChIP-seq analysis detected clear RNA polymerase II binding signals in their surrounding regions, evaluations of previously sequenced and newly shotgun-sequenced complete cDNA sequences showed that they encode preferable transcripts for protein translation, and RNA-seq analysis of polysome-incorporated RNAs yielded direct evidence that those transcripts are actually translated into proteins. We also demonstrate that integrative interpretation of transcriptome data is essential for the selection of putative alternative promoter TSCs, two of which also have protein consequences. Furthermore, discriminative chromatin features that separate TSCs at different expression levels were found for both genic TSCs and intergenic TSCs. The collected integrative information should provide a useful basis for future biological characterization of TSCs. PMID:21372179

  17. Transcriptome analysis of 20 taxonomically related benzylisoquinoline alkaloid-producing plants.

    PubMed

    Hagel, Jillian M; Morris, Jeremy S; Lee, Eun-Jeong; Desgagné-Penix, Isabel; Bross, Crystal D; Chang, Limei; Chen, Xue; Farrow, Scott C; Zhang, Ye; Soh, Jung; Sensen, Christoph W; Facchini, Peter J

    2015-09-18

    Benzylisoquinoline alkaloids (BIAs) represent a diverse class of plant specialized metabolites sharing a common biosynthetic origin beginning with tyrosine. Many BIAs have potent pharmacological activities, and plants accumulating them boast long histories of use in traditional medicine and cultural practices. The decades-long focus on a select number of plant species as model systems has allowed near or full elucidation of major BIA pathways, including those of morphine, sanguinarine and berberine. However, this focus has created a dearth of knowledge surrounding non-model species, which also are known to accumulate a wide-range of BIAs but whose biosynthesis is thus far entirely unexplored. Further, these non-model species represent a rich source of catalyst diversity valuable to plant biochemists and emerging synthetic biology efforts. In order to access the genetic diversity of non-model plants accumulating BIAs, we selected 20 species representing 4 families within the Ranunculales. RNA extracted from each species was processed for analysis by both 1) Roche GS-FLX Titanium and 2) Illumina GA/HiSeq platforms, generating a total of 40 deep-sequencing transcriptome libraries. De novo assembly, annotation and subsequent full-length coding sequence (CDS) predictions indicated greater success for most species using the Illumina-based platform. Assembled data for each transcriptome were deposited into an established web-based BLAST portal ( www.phytometasyn.ca) to allow public access. Homology-based mining of libraries using BIA-biosynthetic enzymes as queries yielded ~850 gene candidates potentially involved in alkaloid biosynthesis. Expression analysis of these candidates was performed using inter-library FPKM normalization methods. These expression data provide a basis for the rational selection of gene candidates, and suggest possible metabolic bottlenecks within BIA metabolism. Phylogenetic analysis was performed for each of 15 different enzyme/protein groupings

  18. Transcriptome profile analysis of floral sex determination in cucumber.

    PubMed

    Wu, Tao; Qin, Zhiwei; Zhou, Xiuyan; Feng, Zhuo; Du, Yalin

    2010-07-15

    Cucumber has been widely studied as a model for floral sex determination. In this investigation, we performed genome-wide transcriptional profiling of apical tissue of a gynoecious mutant (Csg-G) and the monoecious wild-type (Csg-M) of cucumber in an attempt to isolate genes involved in sex determination, using the Solexa technology. The profiling analysis revealed numerous changes in gene expression attributable to the mutation, which resulted in the down-regulation of 600 genes and the up-regulation of 143 genes. The Solexa data were confirmed by reverse transcription polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR (qRT-PCR). Gene ontology (GO) analysis revealed that the differentially expressed genes were mainly involved in biogenesis, transport and organization of cellular component, macromolecular and cellular biosynthesis, localization, establishment of localization, translation and other processes. Furthermore, the expression of some of these genes depended upon the tissue and the developmental stage of the flowers of gynoecious mutant. The results of this study suggest two important concepts, which govern sex determination in cucumber. First, the differential expression of genes involved in plant hormone signaling pathways, such as ACS, Asr1, CsIAA2, CS-AUX1 and TLP, indicate that phytohormones and their crosstalk might play a critical role in the sex determination. Second, the regulation of some transcription factors, including EREBP-9, may also be involved in this developmental process.

  19. RNAseq-based Transcriptome Analysis of Burkholderia glumae Quorum Sensing

    PubMed Central

    Kim, Sunyoung; Park, Jungwook; Kim, Ji Hyeon; Lee, Jongyun; Bang, Bongjun; Hwang, Ingyu; Seo, Young-Su

    2013-01-01

    Burkholderia glumae causes rice grain rot and sheath rot by producing toxoflavin, the expression of which is regulated by quorum sensing (QS). The QS systems of B. glumae rely on N-octanoyl homoserine lactone, synthesized by TofI and its cognate receptor TofR, to activate the genes for toxoflavin biosynthesis and an IclR-type transcriptional regulator gene, qsmR. To understand genome-wide transcriptional profiling of QS signaling, we employed RNAseq of the wild-type B. glumae BGR1 with QS-defective mutant, BGS2 (BGR1 tofI::Ω) and QS-dependent transcriptional regulator mutant, BGS9 (BGR1 qsmR::Ω). A comparison of gene expression profiling among the wild-type BGR1 and the two mutants before and after QS onset as well as gene ontology (GO) enrichment analysis from differential expressed genes (DEGs) revealed that genes involved in motility were highly enriched in TofI-dependent DEGs, whereas genes for transport and DNA polymerase were highly enriched in QsmR-dependent DEGs. Further, a combination of pathways with these DEGs and phenotype analysis of mutants pointed to a couple of metabolic processes, which are dependent on QS in B. glumae, that were directly or indirectly related with bacterial motility. The consistency of observed bacterial phenotypes with GOs or metabolic pathways in QS-regulated genes implied that integration RNAseq with GO enrichment or pathways would be useful to study bacterial physiology and phenotypes. PMID:25288952

  20. Transcriptome analysis of differentiating spermatogonia stimulated with kit ligand.

    PubMed

    Rossi, Pellegrino; Lolicato, Francesca; Grimaldi, Paola; Dolci, Susanna; Di Sauro, Annarita; Filipponi, Doria; Geremia, Raffaele

    2008-01-01

    Kit ligand (KL) is a survival factor and a mitogenic stimulus for differentiating spermatogonia. However, it is not known whether KL also plays a role in the differentiative events that lead to meiotic entry of these cells. We performed a wide genome analysis of difference in gene expression induced by treatment with KL of spermatogonia from 7-day-old mice, using gene chips spanning the whole mouse genome. The analysis revealed that the pattern of RNA expression induced by KL is compatible with the qualitative changes of the cell cycle that occur during the subsequent cell divisions in type A and B spermatogonia, i.e. the progressive lengthening of the S phase and the shortening of the G2/M transition. Moreover, KL up-regulates in differentiating spermatogonia the expression of early meiotic genes (for instance: Lhx8, Nek1, Rnf141, Xrcc3, Tpo1, Tbca, Xrcc2, Mesp1, Phf7, Rtel1), whereas it down-regulates typical spermatogonial markers (for instance: Pole, Ptgs2, Zfpm2, Egr2, Egr3, Gsk3b, Hnrpa1, Fst, Ptch2). Since KL modifies the expression of several genes known to be up-regulated or down-regulated in spermatogonia during the transition from the mitotic to the meiotic cell cycle, these results are consistent with a role of the KL/kit interaction in the induction of their meiotic differentiation.

  1. RNAseq-based Transcriptome Analysis of Burkholderia glumae Quorum Sensing.

    PubMed

    Kim, Sunyoung; Park, Jungwook; Kim, Ji Hyeon; Lee, Jongyun; Bang, Bongjun; Hwang, Ingyu; Seo, Young-Su

    2013-09-01

    Burkholderia glumae causes rice grain rot and sheath rot by producing toxoflavin, the expression of which is regulated by quorum sensing (QS). The QS systems of B. glumae rely on N-octanoyl homoserine lactone, synthesized by TofI and its cognate receptor TofR, to activate the genes for toxoflavin biosynthesis and an IclR-type transcriptional regulator gene, qsmR. To understand genome-wide transcriptional profiling of QS signaling, we employed RNAseq of the wild-type B. glumae BGR1 with QS-defective mutant, BGS2 (BGR1 tofI::Ω) and QS-dependent transcriptional regulator mutant, BGS9 (BGR1 qsmR::Ω). A comparison of gene expression profiling among the wild-type BGR1 and the two mutants before and after QS onset as well as gene ontology (GO) enrichment analysis from differential expressed genes (DEGs) revealed that genes involved in motility were highly enriched in TofI-dependent DEGs, whereas genes for transport and DNA polymerase were highly enriched in QsmR-dependent DEGs. Further, a combination of pathways with these DEGs and phenotype analysis of mutants pointed to a couple of metabolic processes, which are dependent on QS in B. glumae, that were directly or indirectly related with bacterial motility. The consistency of observed bacterial phenotypes with GOs or metabolic pathways in QS-regulated genes implied that integration RNAseq with GO enrichment or pathways would be useful to study bacterial physiology and phenotypes.

  2. Transcriptome Analysis in Domesticated Species: Challenges and Strategies

    PubMed Central

    Hekman, Jessica P.; Johnson, Jennifer L.; Kukekova, Anna V.

    2015-01-01

    Domesticated species occupy a special place in the human world due to their economic and cultural value. In the era of genomic research, domesticated species provide unique advantages for investigation of diseases and complex phenotypes. RNA sequencing, or RNA-seq, has recently emerged as a new approach for studying transcriptional activity of the whole genome, changing the focus from individual genes to gene networks. RNA-seq analysis in domesticated species may complement genome-wide association studies of complex traits with economic importance or direct relevance to biomedical research. However, RNA-seq studies are more challenging in domesticated species than in model organisms. These challenges are at least in part associated with the lack of quality genome assemblies for some domesticated species and the absence of genome assemblies for others. In this review, we discuss strategies for analyzing RNA-seq data, focusing particularly on questions and examples relevant to domesticated species. PMID:26917953

  3. Dynamics in Transcriptomics: Advancements in RNA-seq Time Course and Downstream Analysis

    PubMed Central

    Spies, Daniel; Ciaudo, Constance

    2015-01-01

    Analysis of gene expression has contributed to a plethora of biological and medical research studies. Microarrays have been intensively used for the profiling of gene expression during diverse developmental processes, treatments and diseases. New massively parallel sequencing methods, often named as RNA-sequencing (RNA-seq) are extensively improving our understanding of gene regulation and signaling networks. Computational methods developed originally for microarrays analysis can now be optimized and applied to genome-wide studies in order to have access to a better comprehension of the whole transcriptome. This review addresses current challenges on RNA-seq analysis and specifically focuses on new bioinformatics tools developed for time series experiments. Furthermore, possible improvements in analysis, data integration as well as future applications of differential expression analysis are discussed. PMID:26430493

  4. Transcriptome analysis and molecular signature of human retinal pigment epithelium

    PubMed Central

    Strunnikova, N.V.; Maminishkis, A.; Barb, J.J.; Wang, F.; Zhi, C.; Sergeev, Y.; Chen, W.; Edwards, A.O.; Stambolian, D.; Abecasis, G.; Swaroop, A.; Munson, P.J.; Miller, S.S.

    2010-01-01

    Retinal pigment epithelium (RPE) is a polarized cell layer critical for photoreceptor function and survival. The unique physiology and relationship to the photoreceptors make the RPE a critical determinant of human vision. Therefore, we performed a global expression profiling of native and cultured human fetal and adult RPE and determined a set of highly expressed ‘signature’ genes by comparing the observed RPE gene profiles to the Novartis expression database (SymAtlas: http://wombat.gnf.org/index.html) of 78 tissues. Using stringent selection criteria of at least 10-fold higher expression in three distinct preparations, we identified 154 RPE signature genes, which were validated by qRT-PCR analysis in RPE and in an independent set of 11 tissues. Several of the highly expressed signature genes encode proteins involved in visual cycle, melanogenesis and cell adhesion and Gene ontology analysis enabled the assignment of RPE signature genes to epithelial channels and transporters (ClCN4, BEST1, SLCA20) or matrix remodeling (TIMP3, COL8A2). Fifteen RPE signature genes were associated with known ophthalmic diseases, and 25 others were mapped to regions of disease loci. An evaluation of the RPE signature genes in a recently completed AMD genomewide association (GWA) data set revealed that TIMP3, GRAMD3, PITPNA and CHRNA3 signature genes may have potential roles in AMD pathogenesis and deserve further examination. We propose that RPE signature genes are excellent candidates for retinal diseases and for physiological investigations (e.g. dopachrome tautomerase in melanogenesis). The RPE signature gene set should allow the validation of RPE-like cells derived from human embryonic or induced pluripotent stem cells for cell-based therapies of degenerative retinal diseases. PMID:20360305

  5. Elucidating the modes of action for bioactive compounds in a cell-specific manner by large-scale chemically-induced transcriptomics

    PubMed Central

    Iwata, Michio; Sawada, Ryusuke; Iwata, Hiroaki; Kotera, Masaaki; Yamanishi, Yoshihiro

    2017-01-01

    The identification of the modes of action of bioactive compounds is a major challenge in chemical systems biology of diseases. Genome-wide expression profiling of transcriptional responses to compound treatment for human cell lines is a promising unbiased approach for the mode-of-action analysis. Here we developed a novel approach to elucidate the modes of action of bioactive compounds in a cell-specific manner using large-scale chemically-induced transcriptome data acquired from the Library of Integrated Network-based Cellular Signatures (LINCS), and analyzed 16,268 compounds and 68 human cell lines. First, we performed pathway enrichment analyses of regulated genes to reveal active pathways among 163 biological pathways. Next, we explored potential target proteins (including primary targets and off-targets) with cell-specific transcriptional similarity using chemical–protein interactome. Finally, we predicted new therapeutic indications for 461 diseases based on the target proteins. We showed the usefulness of the proposed approach in terms of prediction coverage, interpretation, and large-scale applicability, and validated the new prediction results experimentally by an in vitro cellular assay. The approach has a high potential for advancing drug discovery and repositioning. PMID:28071740

  6. Transcriptome profiling analysis reveals metabolic changes across various growth phases in Bacillus pumilus BA06.

    PubMed

    Han, Lin-Li; Shao, Huan-Huan; Liu, Yong-Cheng; Liu, Gang; Xie, Chao-Ying; Cheng, Xiao-Jie; Wang, Hai-Yan; Tan, Xue-Mei; Feng, Hong

    2017-07-11

    Bacillus pumilus can secret abundant extracellular enzymes, and may be used as a potential host for the industrial production of enzymes. It is necessary to understand the metabolic processes during cellular growth. Here, an RNA-seq based transcriptome analysis was applied to examine B. pumilus BA06 across various growth stages to reveal metabolic changes under two conditions. Based on the gene expression levels, changes to metabolism pathways that were specific to various growth phases were enriched by KEGG analysis. Upon entry into the transition from the exponential growth phase, striking changes were revealed that included down-regulation of the tricarboxylic acid cycle, oxidative phosphorylation, flagellar assembly, and chemotaxis signaling. In contrast, the expression of stress-responding genes was induced when entering the transition phase, suggesting that the cell may suffer from stress during this growth stage. As expected, up-regulation of sporulation-related genes was continuous during the stationary growth phase, which was consistent with the observed sporulation. However, the expression pattern of the various extracellular proteases was different, suggesting that the regulatory mechanism may be distinct for various proteases. In addition, two protein secretion pathways were enriched with genes responsive to the observed protein secretion in B. pumilus. However, the expression of some genes that encode sporulation-related proteins and extracellular proteases was delayed by the addition of gelatin to the minimal medium. The transcriptome data depict global alterations in the genome-wide transcriptome across the various growth phases, which will enable an understanding of the physiology and phenotype of B. pumilus through gene expression.

  7. Transcriptome Analysis in Sheepgrass (Leymus chinensis): A Dominant Perennial Grass of the Eurasian Steppe

    PubMed Central

    Chen, Shuangyan; Huang, Xin; Yan, Xueqing; Liang, Ye; Wang, Yuezhu; Li, Xiaofeng; Peng, Xianjun; Ma, Xingyong; Zhang, Lexin; Cai, Yueyue; Ma, Tian; Cheng, Liqin; Qi, Dongmei; Zheng, Huajun; Yang, Xiaohan; Li, Xiaoxia; Liu, Gongshe

    2013-01-01

    Background Sheepgrass [Leymus chinensis (Trin.) Tzvel.] is an important perennial forage grass across the Eurasian Steppe and is known for its adaptability to various environmental conditions. However, insufficient data resources in public databases for sheepgrass limited our understanding of the mechanism of environmental adaptations, gene discovery and molecular marker development. Results The transcriptome of sheepgrass was sequenced using Roche 454 pyrosequencing technology. We assembled 952,328 high-quality reads into 87,214 unigenes, including 32,416 contigs and 54,798 singletons. There were 15,450 contigs over 500 bp in length. BLAST searches of our database against Swiss-Prot and NCBI non-redundant protein sequences (nr) databases resulted in the annotation of 54,584 (62.6%) of the unigenes. Gene Ontology (GO) analysis assigned 89,129 GO term annotations for 17,463 unigenes. We identified 11,675 core Poaceae-specific and 12,811 putative sheepgrass-specific unigenes by BLAST searches against all plant genome and transcriptome databases. A total of 2,979 specific freezing-responsive unigenes were found from this RNAseq dataset. We identified 3,818 EST-SSRs in 3,597 unigenes, and some SSRs contained unigenes that were also candidates for freezing-response genes. Characterizations of nucleotide repeats and dominant motifs of SSRs in sheepgrass were also performed. Similarity and phylogenetic analysis indicated that sheepgrass is closely related to barley and wheat. Conclusions This research has greatly enriched sheepgrass transcriptome resources. The identified stress-related genes will help us to decipher the genetic basis of the environmental and ecological adaptations of this species and will be used to improve wheat and barley crops through hybridization or genetic transformation. The EST-SSRs reported here will be a valuable resource for future gene-phenotype studies and for the molecular breeding of sheepgrass and other Poaceae species. PMID:23861841

  8. Transcriptome-Wide Analysis of Hepatitis B Virus-Mediated Changes to Normal Hepatocyte Gene Expression

    PubMed Central

    Lamontagne, Jason; Mell, Joshua C.; Bouchard, Michael J.

    2016-01-01

    Globally, a chronic hepatitis B virus (HBV) infection remains the leading cause of primary liver cancer. The mechanisms leading to the development of HBV-associated liver cancer remain incompletely understood. In part, this is because studies have been limited by the lack of effective model systems that are both readily available and mimic the cellular environment of a normal hepatocyte. Additionally, many studies have focused on single, specific factors or pathways that may be affected by HBV, without addressing cell physiology as a whole. Here, we apply RNA-seq technology to investigate transcriptome-wide, HBV-mediated changes in gene expression to identify single factors and pathways as well as networks of genes and pathways that are affected in the context of HBV replication. Importantly, these studies were conducted in an ex vivo model of cultured primary hepatocytes, allowing for the transcriptomic characterization of this model system and an investigation of early HBV-mediated effects in a biologically relevant context. We analyzed differential gene expression within the context of time-mediated gene-expression changes and show that in the context of HBV replication a number of genes and cellular pathways are altered, including those associated with metabolism, cell cycle regulation, and lipid biosynthesis. Multiple analysis pipelines, as well as qRT-PCR and an independent, replicate RNA-seq analysis, were used to identify and confirm differentially expressed genes. HBV-mediated alterations to the transcriptome that we identified likely represent early changes to hepatocytes following an HBV infection, suggesting potential targets for early therapeutic intervention. Overall, these studies have produced a valuable resource that can be used to expand our understanding of the complex network of host-virus interactions and the impact of HBV-mediated changes to normal hepatocyte physiology on viral replication. PMID:26891448

  9. Integrated clinical, whole-genome, and transcriptome analysis of multisampled lethal metastatic prostate cancer

    PubMed Central

    Bova, G. Steven; Kallio, Heini M.L.; Annala, Matti; Kivinummi, Kati; Högnäs, Gunilla; Häyrynen, Sergei; Rantapero, Tommi; Kivinen, Virpi; Isaacs, William B.; Tolonen, Teemu; Nykter, Matti; Visakorpi, Tapio

    2016-01-01

    We report the first combined analysis of whole-genome sequence, detailed clinical history, and transcriptome sequence of multiple prostate cancer metastases in a single patient (A21). Whole-genome and transcriptome sequence was obtained from nine anatomically separate metastases, and targeted DNA sequencing was performed in cancerous and noncancerous foci within the primary tumor specimen removed 5 yr before death. Transcriptome analysis revealed increased expression of androgen receptor (AR)-regulated genes in liver metastases that harbored an AR p.L702H mutation, suggesting a dominant effect by the mutation despite being present in only one of an estimated 16 copies per cell. The metastases harbored several alterations to the PI3K/AKT pathway, including a clonal truncal mutation in PIK3CG and present in all metastatic sites studied. The list of truncal genomic alterations shared by all metastases included homozygous deletion of TP53, hemizygous deletion of RB1 and CHD1, and amplification of FGFR1. If the patient were treated today, given this knowledge, the use of second-generation androgen-directed therapies, cessation of glucocorticoid administration, and therapeutic inhibition of the PI3K/AKT pathway or FGFR1 receptor could provide personalized benefit. Three previously unreported truncal clonal missense mutations (ABCC4 p.R891L, ALDH9A1 p.W89R, and ASNA1 p.P75R) were expressed at the RNA level and assessed as druggable. The truncal status of mutations may be critical for effective actionability and merit further study. Our findings suggest that a large set of deeply analyzed cases could serve as a powerful guide to more effective prostate cancer basic science and personalized cancer medicine clinical trials. PMID:27148588

  10. Transcriptome analysis of the honey bee fungal pathogen, Ascosphaera apis: implications for host pathogenesis

    PubMed Central

    2012-01-01

    Background We present a comprehensive transcriptome analysis of the fungus Ascosphaera apis, an economically important pathogen of the Western honey bee (Apis mellifera) that causes chalkbrood disease. Our goals were to further annotate the A. apis reference genome and to identify genes that are candidates for being differentially expressed during host infection versus axenic culture. Results We compared A. apis transcriptome sequence from mycelia grown on liquid or solid media with that dissected from host-infected tissue. 454 pyrosequencing provided 252 Mb of filtered sequence reads from both culture types that were assembled into 10,087 contigs. Transcript contigs, protein sequences from multiple fungal species, and ab initio gene predictions were included as evidence sources in the Maker gene prediction pipeline, resulting in 6,992 consensus gene models. A phylogeny based on 12 of these protein-coding loci further supported the taxonomic placement of Ascosphaera as sister to the core Onygenales. Several common protein domains were less abundant in A. apis compared with related ascomycete genomes, particularly cytochrome p450 and protein kinase domains. A novel gene family was identified that has expanded in some ascomycete lineages, but not others. We manually annotated genes with homologs in other fungal genomes that have known relevance to fungal virulence and life history. Functional categories of interest included genes involved in mating-type specification, intracellular signal transduction, and stress response. Computational and manual annotations have been made publicly available on the Bee Pests and Pathogens website. Conclusions This comprehensive transcriptome analysis substantially enhances our understanding of the A. apis genome and its expression during infection of honey bee larvae. It also provides resources for future molecular studies of chalkbrood disease and ultimately improved disease management. PMID:22747707

  11. Transcriptome analysis of the honey bee fungal pathogen, Ascosphaera apis: implications for host pathogenesis.

    PubMed

    Cornman, R Scott; Bennett, Anna K; Murray, K Daniel; Evans, Jay D; Elsik, Christine G; Aronstein, Kate

    2012-06-29

    We present a comprehensive transcriptome analysis of the fungus Ascosphaera apis, an economically important pathogen of the Western honey bee (Apis mellifera) that causes chalkbrood disease. Our goals were to further annotate the A. apis reference genome and to identify genes that are candidates for being differentially expressed during host infection versus axenic culture. We compared A. apis transcriptome sequence from mycelia grown on liquid or solid media with that dissected from host-infected tissue. 454 pyrosequencing provided 252 Mb of filtered sequence reads from both culture types that were assembled into 10,087 contigs. Transcript contigs, protein sequences from multiple fungal species, and ab initio gene predictions were included as evidence sources in the Maker gene prediction pipeline, resulting in 6,992 consensus gene models. A phylogeny based on 12 of these protein-coding loci further supported the taxonomic placement of Ascosphaera as sister to the core Onygenales. Several common protein domains were less abundant in A. apis compared with related ascomycete genomes, particularly cytochrome p450 and protein kinase domains. A novel gene family was identified that has expanded in some ascomycete lineages, but not others. We manually annotated genes with homologs in other fungal genomes that have known relevance to fungal virulence and life history. Functional categories of interest included genes involved in mating-type specification, intracellular signal transduction, and stress response. Computational and manual annotations have been made publicly available on the Bee Pests and Pathogens website. This comprehensive transcriptome analysis substantially enhances our understanding of the A. apis genome and its expression during infection of honey bee larvae. It also provides resources for future molecular studies of chalkbrood disease and ultimately improved disease management.

  12. Integrated clinical, whole-genome, and transcriptome analysis of multisampled lethal metastatic prostate cancer.

    PubMed

    Bova, G Steven; Kallio, Heini M L; Annala, Matti; Kivinummi, Kati; Högnäs, Gunilla; Häyrynen, Sergei; Rantapero, Tommi; Kivinen, Virpi; Isaacs, William B; Tolonen, Teemu; Nykter, Matti; Visakorpi, Tapio

    2016-05-01

    We report the first combined analysis of whole-genome sequence, detailed clinical history, and transcriptome sequence of multiple prostate cancer metastases in a single patient (A21). Whole-genome and transcriptome sequence was obtained from nine anatomically separate metastases, and targeted DNA sequencing was performed in cancerous and noncancerous foci within the primary tumor specimen removed 5 yr before death. Transcriptome analysis revealed increased expression of androgen receptor (AR)-regulated genes in liver metastases that harbored an AR p.L702H mutation, suggesting a dominant effect by the mutation despite being present in only one of an estimated 16 copies per cell. The metastases harbored several alterations to the PI3K/AKT pathway, including a clonal truncal mutation in PIK3CG and present in all metastatic sites studied. The list of truncal genomic alterations shared by all metastases included homozygous deletion of TP53, hemizygous deletion of RB1 and CHD1, and amplification of FGFR1. If the patient were treated today, given this knowledge, the use of second-generation androgen-directed therapies, cessation of glucocorticoid administration, and therapeutic inhibition of the PI3K/AKT pathway or FGFR1 receptor could provide personalized benefit. Three previously unreported truncal clonal missense mutations (ABCC4 p.R891L, ALDH9A1 p.W89R, and ASNA1 p.P75R) were expressed at the RNA level and assessed as druggable. The truncal status of mutations may be critical for effective actionability and merit further study. Our findings suggest that a large set of deeply analyzed cases could serve as a powerful guide to more effective prostate cancer basic science and personalized cancer medicine clinical trials.

  13. Transcriptome Analysis in Sheepgrass (Leymus chinensis). A Dominant Perennial Grass of the Eurasian Steppe

    SciTech Connect

    Chen, Shuangyan; Huang, Xin; Yang, Xiaohan; Liu, Gongshe

    2013-07-04

    BACKGROUND: Sheepgrass [Leymus chinensis (Trin.) Tzvel.] is an important perennial forage grass across the Eurasian Steppe and is known for its adaptability to various environmental conditions. However, insufficient data resources in public databases for sheepgrass limited our understanding of the mechanism of environmental adaptations, gene discovery and molecular marker development. RESULTS: The transcriptome of sheepgrass was sequenced using Roche 454 pyrosequencing technology. We assembled 952,328 high-quality reads into 87,214 unigenes, including 32,416 contigs and 54,798 singletons. There were 15,450 contigs over 500 bp in length. BLAST searches of our database against Swiss-Prot and NCBI non-redundant protein sequences (nr) databases resulted in the annotation of 54,584 (62.6%) of the unigenes. Gene Ontology (GO) analysis assigned 89,129 GO term annotations for 17,463 unigenes. We identified 11,675 core Poaceae-specific and 12,811 putative sheepgrass-specific unigenes by BLAST searches against all plant genome and transcriptome databases. A total of 2,979 specific freezing-responsive unigenes were found from this RNAseq dataset. We identified 3,818 EST-SSRs in 3,597 unigenes, and some SSRs contained unigenes that were also candidates for freezing-response genes. Characterizations of nucleotide repeats and dominant motifs of SSRs in sheepgrass were also performed. Similarity and phylogenetic analysis indicated that sheepgrass is closely related to barley and wheat. CONCLUSIONS: This research has greatly enriched sheepgrass transcriptome resources. The identified stress-related genes will help us to decipher the genetic basis of the environmental and ecological adaptations of this species and will be used to improve wheat and barley crops through hybridization or genetic transformation. The EST-SSRs reported here will be a valuable resource for future gene-phenotype studies and for the molecular breeding of sheepgrass and other Poaceae species.

  14. Comparative Analysis of Proteome and Transcriptome Variation in Mouse

    PubMed Central

    Hagopian, Raffi; Mungrue, Imran N.; Farber, Charles R.; Sinsheimer, Janet; Kang, Hyun M.; Furlotte, Nicholas; Park, Christopher C.; Wen, Ping-Zi; Brewer, Heather; Weitz, Karl; Camp, David G.; Pan, Calvin; Yordanova, Roumyana; Neuhaus, Isaac; Tilford, Charles; Siemers, Nathan; Gargalovic, Peter; Eskin, Eleazar; Kirchgessner, Todd; Smith, Desmond J.; Smith, Richard D.; Lusis, Aldons J.

    2011-01-01

    The relationships between the levels of transcripts and the levels of the proteins they encode have not been examined comprehensively in mammals, although previous work in plants and yeast suggest a surprisingly modest correlation. We have examined this issue using a genetic approach in which natural variations were used to perturb both transcript levels and protein levels among inbred strains of mice. We quantified over 5,000 peptides and over 22,000 transcripts in livers of 97 inbred and recombinant inbred strains and focused on the 7,185 most heritable transcripts and 486 most reliable proteins. The transcript levels were quantified by microarray analysis in three replicates and the proteins were quantified by Liquid Chromatography–Mass Spectrometry using O(18)-reference-based isotope labeling approach. We show that the levels of transcripts and proteins correlate significantly for only about half of the genes tested, with an average correlation of 0.27, and the correlations of transcripts and proteins varied depending on the cellular location and biological function of the gene. We examined technical and biological factors that could contribute to the modest correlation. For example, differential splicing clearly affects the analyses for certain genes; but, based on deep sequencing, this does not substantially contribute to the overall estimate of the correlation. We also employed genome-wide association analyses to map loci controlling both transcript and protein levels. Surprisingly, little overlap was observed between the protein- and transcript-mapped loci. We have typed numerous clinically relevant traits among the strains, including adiposity, lipoprotein levels, and tissue parameters. Using correlation analysis, we found that a low number of clinical trait relationships are preserved between the protein and mRNA gene products and that the majority of such relationships are specific to either the protein levels or transcript levels. Surprisingly

  15. Comparative analysis of proteome and transcriptome variation in mouse.

    PubMed

    Ghazalpour, Anatole; Bennett, Brian; Petyuk, Vladislav A; Orozco, Luz; Hagopian, Raffi; Mungrue, Imran N; Farber, Charles R; Sinsheimer, Janet; Kang, Hyun M; Furlotte, Nicholas; Park, Christopher C; Wen, Ping-Zi; Brewer, Heather; Weitz, Karl; Camp, David G; Pan, Calvin; Yordanova, Roumyana; Neuhaus, Isaac; Tilford, Charles; Siemers, Nathan; Gargalovic, Peter; Eskin, Eleazar; Kirchgessner, Todd; Smith, Desmond J; Smith, Richard D; Lusis, Aldons J

    2011-06-01

    The relationships between the levels of transcripts and the levels of the proteins they encode have not been examined comprehensively in mammals, although previous work in plants and yeast suggest a surprisingly modest correlation. We have examined this issue using a genetic approach in which natural variations were used to perturb both transcript levels and protein levels among inbred strains of mice. We quantified over 5,000 peptides and over 22,000 transcripts in livers of 97 inbred and recombinant inbred strains and focused on the 7,185 most heritable transcripts and 486 most reliable proteins. The transcript levels were quantified by microarray analysis in three replicates and the proteins were quantified by Liquid Chromatography-Mass Spectrometry using O(18)-reference-based isotope labeling approach. We show that the levels of transcripts and proteins correlate significantly for only about half of the genes tested, with an average correlation of 0.27, and the correlations of transcripts and proteins varied depending on the cellular location and biological function of the gene. We examined technical and biological factors that could contribute to the modest correlation. For example, differential splicing clearly affects the analyses for certain genes; but, based on deep sequencing, this does not substantially contribute to the overall estimate of the correlation. We also employed genome-wide association analyses to map loci controlling both transcript and protein levels. Surprisingly, little overlap was observed between the protein- and transcript-mapped loci. We have typed numerous clinically relevant traits among the strains, including adiposity, lipoprotein levels, and tissue parameters. Using correlation analysis, we found that a low number of clinical trait relationships are preserved between the protein and mRNA gene products and that the majority of such relationships are specific to either the protein levels or transcript levels. Surprisingly

  16. Deep SAGE analysis of the Caenorhabditis elegans transcriptome.

    PubMed

    Ruzanov, Peter; Riddle, Donald L

    2010-06-01

    We employed the Tag-seq technique to generate global transcription profiles for different strains and life stages of the nematode C. elegans. Tag-seq generates cDNA tags as does Serial Analysis of Gene Expression (SAGE), but the method yields a much larger number of tags, generating much larger data sets than SAGE. We examined differences in the performance of SAGE and Tag-seq by comparing gene expression data for 13 pairs of libraries. We identified genes for which expression was consistently changed in long-lived worms. Additional genes emerged in the deeper Tag-seq profiles, including several 'signature' genes found among those zup-regulated in long-lived dauer larvae (cki-1, aak-2 and daf-16). Fifty to sixty percent of the genes differentially expressed in daf-2(-) versus daf-2(+) adults had fragmentary or no functional annotation, suggesting the involvement of as yet unstudied pathways in aging. We were able to distinguish between changes in gene expression associated with altered genotype or altered growth conditions. We found 62 cases of possible mRNA isoform switching in the 13 Tag-seq libraries, whereas the 13 SAGE libraries allowed detection of only 15 such occurrences. We observed strong expression of anti-sense transcripts for several mitochondrial genes, but nuclear anti-sense transcripts were neither abundant nor consistently expressed among the libraries.

  17. A transcriptome analysis of mitten crab testes (Eriocheir sinensis)

    PubMed Central

    Zhang, Wei; Wan, Haolei; Jiang, Hui; Zhao, Yunlong; Zhang, Xiaowei; Hu, Songnian; Wang, Qun

    2011-01-01

    The identification of expressed genes involved in sexual precocity of the mitten crab (Eriocheir sinensis) is critical for a better understanding of its reproductive development. To this end, we constructed a cDNA library from the rapid developmental stage of testis of E. sinensis and sequenced 3,388 randomly picked clones. After processing, 2,990 high-quality expressed sequence tags (ESTs) were clustered into 2,415 unigenes including 307 contigs and 2,108 singlets, which were then compared to the NCBI non-redundant (nr) protein and nucleotide (nt) database for annotation with Blastx and Blastn, respectively. After further analysis, 922 unigenes were obtained with concrete annotations and 30 unigenes were found to have functions possibly related to the process of reproduction in male crabs – six transcripts relevant to spermatogenesis (especially Cyclin K and RecA homolog DMC1), two transcripts involved in nuclear protein transformation, two heat-shock protein genes, eleven transcription factor genes (a series of zinc-finger proteins), and nine cytoskeleton protein-related genes. Our results, besides providing valuable information related to crustacean reproduction, can also serve as a base for future studies of reproductive and developmental biology. PMID:21637557

  18. Global transcriptome analysis and identification of genes involved in nutrients accumulation during seed development of rice tartary buckwheat (Fagopyrum Tararicum).

    PubMed

    Huang, Juan; Deng, Jiao; Shi, Taoxiong; Chen, Qijiao; Liang, Chenggang; Meng, Ziye; Zhu, Liwei; Wang, Yan; Zhao, Fengli; Yu, Shizhou; Chen, Qingfu

    2017-09-18

    Tartary buckwheat seeds are rich in various nutrients, such as storage proteins, starch, and flavonoids. To get a good knowledge of the transcriptome dynamics and gene regulatory mechanism during the process of seed development and nutrients accumulation, we performed a comprehensive global transcriptome analysis using rice tartary buckwheat seeds at different development stages, namely pre-filling stage, filling stage, and mature stage. 24 819 expressed genes, including 108 specifically expressed genes, and 11 676 differentially expressed genes (DEGs) were identified. qRT-PCR analysis was performed on 34 DEGs to validate the transcriptome data, and a good consistence was obtained. Based on their expression patterns, the identified DEGs were classified to eight clusters, and the enriched GO items in each cluster were analyzed. In addition, 633 DEGs related to plant hormones were identified. Furthermore, genes in the biosynthesis pathway of nutrients accumulation were analyzed, including 10, 20, and 23 DEGs corresponding to the biosynthesis of seed storage proteins, flavonoids, and starch, respectively. This is the first transcriptome analysis during seed development of tartary buckwheat. It would provide us a comprehensive understanding of the complex transcriptome dynamics during seed development and gene regulatory mechanism of nutrients accumulation.

  19. Transcriptome Analysis and Development of SSR Molecular Markers in Glycyrrhiza uralensis Fisch.

    PubMed Central

    Liu, Yaling; Zhang, Pengfei; Song, Meiling; Hou, Junling; Qing, Mei; Wang, Wenquan; Liu, Chunsheng

    2015-01-01

    Licorice is an important traditional Chinese medicine with clinical and industrial applications. Genetic resources of licorice are insufficient for analysis of molecular biology and genetic functions; as such, transcriptome sequencing must be conducted for functional characterization and development of molecular markers. In this study, transcriptome sequencing on the Illumina HiSeq 2500 sequencing platform generated a total of 5.41 Gb clean data. De novo assembly yielded a total of 46,641 unigenes. Comparison analysis using BLAST showed that the annotations of 29,614 unigenes were conserved. Further study revealed 773 genes related to biosynthesis of secondary metabolites of licorice, 40 genes involved in biosynthesis of the terpenoid backbone, and 16 genes associated with biosynthesis of glycyrrhizic acid. Analysis of unigenes larger than 1 Kb with a length of 11,702 nt presented 7,032 simple sequence repeats (SSR). Sixty-four of 69 randomly designed and synthesized SSR pairs were successfully amplified, 33 pairs of primers were polymorphism in in Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat., Glycyrrhiza glabra L. and Glycyrrhiza pallidiflora Maxim. This study not only presents the molecular biology data of licorice but also provides a basis for genetic diversity research and molecular marker-assisted breeding of licorice. PMID:26571372

  20. Transcriptome analysis of creeping bentgrass exposed to drought stress and polyamine treatment

    PubMed Central

    Ma, Yingmei; Shukla, Vijaya; Merewitz, Emily B.

    2017-01-01

    Creeping bentgrass is an important cool-season turfgrass species sensitive to drought. Treatment with polyamines (PAs) has been shown to improve drought tolerance; however, the mechanism is not yet fully understood. Therefore, this study aimed to evaluate transcriptome changes of creeping bentgrass in response to drought and exogenous spermidine (Spd) application using RNA sequencing (RNA-Seq). The high-quality sequences were assembled and 18,682 out of 49,190 (38%) were detected as coding sequences. A total of 22% and 19% of genes were found to be either up- or down-regulated due to drought while 20% and 34% genes were either up- or down- regulated in response to Spd application under drought conditions, respectively. Gene ontology (GO) and enrichment analysis were used to interpret the biological processes of transcripts and relative transcript abundance. Enriched or differentially expressed transcripts due to drought stress and/or Spd application were primarily associated with energy metabolism, transport, antioxidants, photosynthesis, signaling, stress defense, and cellular response to water deprivation. This research is the first to provide transcriptome data for creeping bentgrass under an abiotic stress using RNA-Seq analysis. Differentially expressed transcripts identified here could be further investigated for use as molecular markers or for functional analysis in responses to drought and Spd. PMID:28445484

  1. Single prokaryotic cell isolation and total transcript amplification protocol for transcriptomic analysis

    PubMed Central

    Kang, Yun; McMillan, Ian; Norris, Michael H; Hoang, Tung T.

    2015-01-01

    Until recently, transcriptome analyses of single cells have been confined to eukaryotes. The information obtained from single cell transcripts can provide detailed insight into spatiotemporal gene-expression, and could be even more valuable if expanded to prokaryotic cells. Transcriptome analysis of single prokaryotic cells is a recently developed and powerful tool. Here, we describe a procedure that allows amplification of the total transcript of a single prokaryotic cell for in-depth analysis. This is performed by utilizing a laser capture microdissection instrument for single cell isolation, followed by reverse transcription via Moloney Murine Leukemia virus, degradation of chromosomal DNA with McrBC and DpnI restriction enzymes, ss-cDNA ligation using T4 polynucleotide kinase and CircLigase, and polymerization of ss-cDNA to ds-cDNA by ϕ 29 polymerase. This procedure takes ~5 days, and sufficient amounts of ds-cDNA can be obtained from single cell RNA template for further microarray analysis. PMID:26042386

  2. Comparative transcriptome analysis during early fruit development between three seedy citrus genotypes and their seedless mutants

    PubMed Central

    Zhang, Shujian; Shi, Qingchun; Albrecht, Ute; Shatters, Robert G; Stange, Ric; McCollum, Greg; Zhang, Shuo; Fan, Chengming; Stover, Ed

    2017-01-01

    Identification of genes with differential transcript abundance (GDTA) in seedless mutants may enhance understanding of seedless citrus development. Transcriptome analysis was conducted at three time points during early fruit development (Phase 1) of three seedy citrus genotypes: Fallglo (Bower citrus hybrid (Citrus reticulata×C. reticulata×C. paradisi)×Temple (C. reticulata×C. sinensis)), grapefruit (C. paradisi), Pineapple sweet orange (C. sinensis), and their seedless mutants. Seed abortion in seedless mutants was observed at 26 days post anthesis (Time point 2). Affymetrix transcriptomic analysis revealed 359 to 1077 probe sets with differential transcript abundance in the comparison of seedless versus seedy fruits for each citrus genotypes and time points. The GDTA identified by 18 microarray probe sets were validated by qPCR. Hierarchical clustering analysis revealed a range of GDTA associated with development, hormone and protein metabolism, all of which may reflect genes associated with seedless fruit development. There were 14, 9 and 12 genes found exhibiting similar abundance ratios in all three seedless versus seedy genotype comparisons at time point 1, 2 and 3, respectively. Among those genes were genes coding for an aspartic protease and a cysteine protease, which may play important roles in seedless fruit development. New insights into seedless citrus fruit development may contribute to biotech approaches to create seedless cultivars. PMID:28904803

  3. Single prokaryotic cell isolation and total transcript amplification protocol for transcriptomic analysis.

    PubMed

    Kang, Yun; McMillan, Ian; Norris, Michael H; Hoang, Tung T

    2015-07-01

    Until recently, transcriptome analyses of single cells have been confined to eukaryotes. The information obtained from single-cell transcripts can provide detailed insight into spatiotemporal gene expression, and it could be even more valuable if expanded to prokaryotic cells. Transcriptome analysis of single prokaryotic cells is a recently developed and powerful tool. Here we describe a procedure that allows amplification of the total transcript of a single prokaryotic cell for in-depth analysis. This is performed by using a laser-capture microdissection instrument for single-cell isolation, followed by reverse transcription via Moloney murine leukemia virus, degradation of chromosomal DNA with McrBC and DpnI restriction enzymes, single-stranded cDNA (ss-cDNA) ligation using T4 polynucleotide kinase and CircLigase, and polymerization of ss-cDNA to double-stranded cDNA (ds-cDNA) by Φ29 polymerase. This procedure takes ∼5 d, and sufficient amounts of ds-cDNA can be obtained from single-cell RNA template for further microarray analysis.

  4. Transcriptome analysis of Phoenix canariensis Chabaud in response to Rhynchophorus ferrugineus Olivier attacks

    PubMed Central

    Giovino, Antonio; Bertolini, Edoardo; Fileccia, Veronica; Al Hassan, Mohamad; Labra, Massimo; Martinelli, Federico

    2015-01-01

    Red Palm Weevil (RPW, Rhynchophorus ferrugineus Olivier) threatens most palm species worldwide. Until now, no studies have analyzed the gene regulatory networks of Phoenix canariensis (Chabaud) in response to RPW attacks. The aim of this study was to fill this knowledge gap. Providing this basic knowledge is very important to improve its management. Results: A deep transcriptome analysis was performed on fully expanded leaves of healthy non-infested trees and attacked trees at two symptom stages (middle and late infestation). A total of 54 genes were significantly regulated during middle stage. Pathway enrichment analysis showed that phenylpropanoid-related pathways were induced at this stage. More than 3300 genes were affected during late stage of attacks. Higher transcript abundances were observed for lipid fatty acid metabolism (fatty acid and glycerolipids), tryptophan metabolism, phenylpropanoid metabolism. Key RPW-modulated genes involved in innate response mediated by hormone crosstalk were observed belonging to auxin, jasmonate and salicylic acid (SA) pathways. Among transcription factors, some WRKYs were clearly induced. qRT-PCR validation confirmed the upregulation of key genes chosen as validation of transcriptomic analysis. Conclusion: A subset of these genes may be further analyzed in future studies to confirm their specificity to be induced by RPW infestations. PMID:26528297

  5. Transcriptome Analysis and Development of SSR Molecular Markers in Glycyrrhiza uralensis Fisch.

    PubMed

    Liu, Yaling; Zhang, Pengfei; Song, Meiling; Hou, Junling; Qing, Mei; Wang, Wenquan; Liu, Chunsheng

    2015-01-01

    Licorice is an important traditional Chinese medicine with clinical and industrial applications. Genetic resources of licorice are insufficient for analysis of molecular biology and genetic functions; as such, transcriptome sequencing must be conducted for functional characterization and development of molecular markers. In this study, transcriptome sequencing on the Illumina HiSeq 2500 sequencing platform generated a total of 5.41 Gb clean data. De novo assembly yielded a total of 46,641 unigenes. Comparison analysis using BLAST showed that the annotations of 29,614 unigenes were conserved. Further study revealed 773 genes related to biosynthesis of secondary metabolites of licorice, 40 genes involved in biosynthesis of the terpenoid backbone, and 16 genes associated with biosynthesis of glycyrrhizic acid. Analysis of unigenes larger than 1 Kb with a length of 11,702 nt presented 7,032 simple sequence repeats (SSR). Sixty-four of 69 randomly designed and synthesized SSR pairs were successfully amplified, 33 pairs of primers were polymorphism in in Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat., Glycyrrhiza glabra L. and Glycyrrhiza pallidiflora Maxim. This study not only presents the molecular biology data of licorice but also provides a basis for genetic diversity research and molecular marker-assisted breeding of licorice.

  6. Gut transcriptome analysis on females of Ornithodoros mimon (Acari: Argasidae) and phylogenetic inference of ticks.

    PubMed

    Landulfo, Gabriel Alves; Patané, José Salvatore Leister; Silva, Dalton Giovanni Nogueira da; Junqueira-de-Azevedo, Inácio Loiola Meirelles; Mendonca, Ronaldo Zucatelli; Simons, Simone Michaela; Carvalho, Eneas de; Barros-Battesti, Darci Moraes

    2017-01-01

    Ornithodoros mimon is an argasid tick that parasitizes bats, birds and opossums and is also harmful to humans. Knowledge of the transcripts present in the tick gut helps in understanding the role of vital molecules in the digestion process and parasite-host relationship, while also providing information about the evolution of arthropod hematophagy. Thus, the present study aimed to know and ascertain the main molecules expressed in the gut of argasid after their blood meal, through analysis on the gut transcriptome of engorged females of O. mimon using 454-based RNA sequencing. The gut transcriptome analysis reveals several transcripts associated with hemoglobin digestion, such as serine, cysteine, aspartic proteases and metalloenzymes. The phylogenetic analysis on the peptidases confirmed that most of them are clustered with other tick genes. We recorded the presence a cathepsin O peptidase-coding transcript in ticks. The topology of the phylogenetic inferences, based on transcripts of inferred families of homologues, was similar to that of previous reports based on mitochondrial genome and nuclear rRNA sequences. We deposited 2,213 sequence of O. mimon to the public databases. Our findings may help towards better understanding of important argasid metabolic processes, such as digestion, nutrition and immunity.

  7. Transcriptome Analysis of Differentially Expressed Genes Provides Insight into Stolon Formation in Tulipa edulis.

    PubMed

    Miao, Yuanyuan; Zhu, Zaibiao; Guo, Qiaosheng; Zhu, Yunhao; Yang, Xiaohua; Sun, Yuan

    2016-01-01

    Tulipa edulis (Miq.) Baker is an important medicinal plant with a variety of anti-cancer properties. The stolon is one of the main asexual reproductive organs of T. edulis and possesses a unique morphology. To explore the molecular mechanism of stolon formation, we performed an RNA-seq analysis of the transcriptomes of stolons at three developmental stages. In the present study, 15.49 Gb of raw data were generated and assembled into 74,006 unigenes, and a total of 2,811 simple sequence repeats were detected in T. edulis. Among the three libraries of stolons at different developmental stages, there were 5,119 differentially expressed genes (DEGs). A functional annotation analysis based on sequence similarity queries of the GO, COG, KEGG databases showed that these DEGs were mainly involved in many physiological and biochemical processes, such as material and energy metabolism, hormone signaling, cell growth, and transcription regulation. In addition, quantitative real-time PCR analysis revealed that the expression patterns of the DEGs were consistent with the transcriptome data, which further supported a role for the DEGs in stolon formation. This study provides novel resources for future genetic and molecular studies in T. edulis.

  8. Transcriptome Analysis of Differentially Expressed Genes Provides Insight into Stolon Formation in Tulipa edulis

    PubMed Central

    Miao, Yuanyuan; Zhu, Zaibiao; Guo, Qiaosheng; Zhu, Yunhao; Yang, Xiaohua; Sun, Yuan

    2016-01-01

    Tulipa edulis (Miq.) Baker is an important medicinal plant with a variety of anti-cancer properties. The stolon is one of the main asexual reproductive organs of T. edulis and possesses a unique morphology. To explore the molecular mechanism of stolon formation, we performed an RNA-seq analysis of the transcriptomes of stolons at three developmental stages. In the present study, 15.49 Gb of raw data were generated and assembled into 74,006 unigenes, and a total of 2,811 simple sequence repeats were detected in T. edulis. Among the three libraries of stolons at different developmental stages, there were 5,119 differentially expressed genes (DEGs). A functional annotation analysis based on sequence similarity queries of the GO, COG, KEGG databases showed that these DEGs were mainly involved in many physiological and biochemical processes, such as material and energy metabolism, hormone signaling, cell growth, and transcription regulation. In addition, quantitative real-time PCR analysis revealed that the expression patterns of the DEGs were consistent with the transcriptome data, which further supported a role for the DEGs in stolon formation. This study provides novel resources for future genetic and molecular studies in T. edulis. PMID:27064558

  9. De novo transcriptomic analysis of cowpea (Vigna unguiculata L. Walp.) for genic SSR marker development.

    PubMed

    Chen, Honglin; Wang, Lixia; Liu, Xiaoyan; Hu, Liangliang; Wang, Suhua; Cheng, Xuzhen

    2017-07-11

    Cowpea [Vigna unguiculata (L.) Walp.] is one of the most important legumes in tropical and semi-arid regions. However, there is relatively little genomic information available for genetic research on and breeding of cowpea. The objectives of this study were to analyse the cowpea transcriptome and develop genic molecular markers for future genetic studies of this genus. Approximately 54 million high-quality cDNA sequence reads were obtained from cowpea based on Illumina paired-end sequencing technology and were de novo assembled to generate 47,899 unigenes with an N50 length of 1534 bp. Sequence similarity analysis revealed 36,289 unigenes (75.8%) with significant similarity to known proteins in the non-redundant (Nr) protein database, 23,471 unigenes (49.0%) with BLAST hits in the Swiss-Prot database, and 20,654 unigenes (43.1%) with high similarity in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Further analysis identified 5560 simple sequence repeats (SSRs) as potential genic molecular markers. Validating a random set of 500 SSR markers yielded 54 polymorphic markers among 32 cowpea accessions. This transcriptomic analysis of cowpea provided a valuable set of genomic data for characterizing genes with important agronomic traits in Vigna unguiculata and a new set of genic SSR markers for further genetic studies and breeding in cowpea and related Vigna species.

  10. Transcriptome Analysis of Manganese-deficient Chlamydomonas reinhardtii Provides Insight on the Chlorophyll Biosynthesis Pathway

    SciTech Connect

    Lockhart, Ainsley; Zvenigorodsky, Natasha; Pedraza, Mary Ann; Lindquist, Erika

    2011-08-11

    The biosynthesis of chlorophyll and other tetrapyrroles is a vital but poorly understood process. Recent genomic advances with the unicellular green algae Chlamydomonas reinhardtii have created opportunity to more closely examine the mechanisms of the chlorophyll biosynthesis pathway via transcriptome analysis. Manganese is a nutrient of interest for complex reactions because of its multiple stable oxidation states and role in molecular oxygen coordination. C. reinhardtii was cultured in Manganese-deplete Tris-acetate-phosphate (TAP) media for 24 hours and used to create cDNA libraries for sequencing using Illumina TruSeq technology. Transcriptome analysis provided intriguing insight on possible regulatory mechanisms in the pathway. Evidence supports similarities of GTR (Glutamyl-tRNA synthase) to its Chlorella vulgaris homolog in terms of Mn requirements. Data was also suggestive of Mn-related compensatory up-regulation for pathway proteins CHLH1 (Manganese Chelatase), GUN4 (Magnesium chelatase activating protein), and POR1 (Light-dependent protochlorophyllide reductase). Intriguingly, data suggests possible reciprocal expression of oxygen dependent CPX1 (coproporphyrinogen III oxidase) and oxygen independent CPX2. Further analysis using RT-PCR could provide compelling evidence for several novel regulatory mechanisms in the chlorophyll biosynthesis pathway.

  11. Transcriptome analysis reveals novel patterning and pigmentation genes underlying Heliconius butterfly wing pattern variation

    PubMed Central

    2012-01-01

    Background Heliconius butterfly wing pattern diversity offers a unique opportunity to investigate how natural genetic variation can drive the evolution of complex adaptive phenotypes. Positional cloning and candidate gene studies have identified a handful of regulatory and pigmentation genes implicated in Heliconius wing pattern variation, but little is known about the greater developmental networks within which these genes interact to pattern a wing. Here we took a large-scale transcriptomic approach to identify the network of genes involved in Heliconius wing pattern development and variation. This included applying over 140 transcriptome microarrays to assay gene expression in dissected wing pattern elements across a range of developmental stages and wing pattern morphs of Heliconius erato. Results We identified a number of putative early prepattern genes with color-pattern related expression domains. We also identified 51 genes differentially expressed in association with natural color pattern variation. Of these, the previously identified color pattern “switch gene” optix was recovered as the first transcript to show color-specific differential expression. Most differentially expressed genes were transcribed late in pupal development and have roles in cuticle formation or pigment synthesis. These include previously undescribed transporter genes associated with ommochrome pigmentation. Furthermore, we observed upregulation of melanin-repressing genes such as ebony and Dat1 in non-melanic patterns. Conclusions This study identifies many new genes implicated in butterfly wing pattern development and provides a glimpse into the number and types of genes affected by variation in genes that drive color pattern evolution. PMID:22747837

  12. Transcriptomic and Genetic Analysis of Direct Interspecies Electron Transfer

    PubMed Central

    Rotaru, Amelia-Elena; Summers, Zarath M.; Shrestha, Minita; Liu, Fanghua; Lovley, Derek R.

    2013-01-01

    The possibility that metatranscriptomic analysis could distinguish between direct interspecies electron transfer (DIET) and H2 interspecies transfer (HIT) in anaerobic communities was investigated by comparing gene transcript abundance in cocultures in which Geobacter sulfurreducens was the electron-accepting partner for either Geobacter metallireducens, which performs DIET, or Pelobacter carbinolicus, which relies on HIT. Transcript abundance for G. sulfurreducens uptake hydrogenase genes was 7-fold lower in cocultures with G. metallireducens than in cocultures with P. carbinolicus, consistent with DIET and HIT, respectively, in the two cocultures. Transcript abundance for the pilus-associated cytochrome OmcS, which is essential for DIET but not for HIT, was 240-fold higher in the cocultures with G. metallireducens than in cocultures with P. carbinolicus. The pilin gene pilA was moderately expressed despite a mutation that might be expected to repress pilA expression. Lower transcript abundance for G. sulfurreducens genes associated with acetate metabolism in the cocultures with P. carbinolicus was consistent with the repression of these genes by H2 during HIT. Genes for the biogenesis of pili and flagella and several c-type cytochrome genes were among the most highly expressed in G. metallireducens. Mutant strains that lacked the ability to produce pili, flagella, or the outer surface c-type cytochrome encoded by Gmet_2896 were not able to form cocultures with G. sulfurreducens. These results demonstrate that there are unique gene expression patterns that distinguish DIET from HIT and suggest that metatranscriptomics may be a promising route to investigate interspecies electron transfer pathways in more-complex environments. PMID:23377933

  13. Global transcriptome analysis of the heat shock response ofshewanella oneidensis

    SciTech Connect

    Gao, Haichun; Wang, Sarah; Liu, Xueduan; Yan, Tinfeng; Wu, Liyou; Alm, Eric; Arkin, Adam P.; Thompson, Dorothea K.; Zhou, Jizhong

    2004-04-30

    Shewanella oneidensis is an important model organism for bioremediation studies because of its diverse respiratory capabilities. However, the genetic basis and regulatory mechanisms underlying the ability of S. oneidensis to survive and adapt to various environmentally relevant stresses is poorly understood. To define this organism's molecular response to elevated growth temperatures, temporal gene expression profiles were examined in cells subjected to heat stress using whole-genome DNA microarrays for S. oneidensis MR-1. Approximately 15 percent (711) of the predicted S. oneidensis genes represented on the microarray were significantly up- or down-regulated (P < 0.05) over a 25-min period following shift to the heat shock temperature (42 C). As expected, the majority of S. oneidensis genes exhibiting homology to known chaperones and heat shock proteins (Hsps) were highly and transiently induced. In addition, a number of predicted genes encoding enzymes in glycolys is and the pentose cycle, [NiFe] dehydrogenase, serine proteases, transcriptional regulators (MerR, LysR, and TetR families), histidine kinases, and hypothetical proteins were induced in response to heat stress. Genes encoding membrane proteins were differentially expressed, suggesting that cells possibly alter their membrane composition or structure in response to variations in growth temperature. A substantial number of the genes encoding ribosomal proteins displayed down-regulated co-expression patterns in response to heat stress, as did genes encoding prophage and flagellar proteins. Finally, based on computational comparative analysis of the upstream promoter regions of S.oneidensis heat-inducible genes, a putative regulatory motif, showing high conservation to the Escherichia coli sigma 32-binding consensus sequence, was identified.

  14. 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

  15. Transcriptomic analysis of Clostridium thermocellum ATCC 27405 cellulose fermentation

    SciTech Connect

    McKeown, Catherine K; Brown, Steven D

    2011-01-01

    The ability of Clostridium thermocellum ATCC 27405 wild-type strain to hydrolyze cellulose and ferment the degradation products directly to ethanol and other metabolic byproducts makes it an attractive candidate for consolidated bioprocessing of cellulosic biomass to biofuels. In this study, whole-genome microarrays were used to investigate the expression of C. thermocellum mRNA during growth on crystalline cellulose in controlled replicate batch fermentations. A time-series analysis of gene expression revealed changes in transcript levels of {approx}40% of genes ({approx}1300 out of 3198 ORFs encoded in the genome) during transition from early-exponential to late-stationary phase. K-means clustering of genes with statistically significant changes in transcript levels identified six distinct clusters of temporal expression. Broadly, genes involved in energy production, translation, glycolysis and amino acid, nucleotide and coenzyme metabolism displayed a decreasing trend in gene expression as cells entered stationary phase. In comparison, genes involved in cell structure and motility, chemotaxis, signal transduction and transcription showed an increasing trend in gene expression. Hierarchical clustering of cellulosome-related genes highlighted temporal changes in composition of this multi-enzyme complex during batch growth on crystalline cellulose, with increased expression of several genes encoding hydrolytic enzymes involved in degradation of non-cellulosic substrates in stationary phase. Overall, the results suggest that under low substrate availability, growth slows due to decreased metabolic potential and C. thermocellum alters its gene expression to (i) modulate the composition of cellulosomes that are released into the environment with an increased proportion of enzymes than can efficiently degrade plant polysaccharides other than cellulose, (ii) enhance signal transduction and chemotaxis mechanisms perhaps to sense the oligosaccharide hydrolysis products

  16. Combined Large-Scale Phenotyping and Transcriptomics in Maize Reveals a Robust Growth Regulatory Network1[OPEN

    PubMed Central

    Herman, Dorota; Slabbinck, Bram; Pè, Mario Enrico

    2016-01-01

    Leaves are vital organs for biomass and seed production because of their role in the generation of metabolic energy and organic compounds. A better understanding of the molecular networks underlying leaf development is crucial to sustain global requirements for food and renewable energy. Here, we combined transcriptome profiling of proliferative leaf tissue with in-depth phenotyping of the fourth leaf at later stages of development in 197 recombinant inbred lines of two different maize (Zea mays) populations. Previously, correlation analysis in a classical biparental mapping population identified 1,740 genes correlated with at least one of 14 traits. Here, we extended these results with data from a multiparent advanced generation intercross population. As expected, the phenotypic variability was found to be larger in the latter population than in the biparental population, although general conclusions on the correlations among the traits are comparable. Data integration from the two diverse populations allowed us to identify a set of 226 genes that are robustly associated with diverse leaf traits. This set of genes is enriched for transcriptional regulators and genes involved in protein synthesis and cell wall metabolism. In order to investigate the molecular network context of the candidate gene set, we integrated our data with publicly available functional genomics data and identified a growth regulatory network of 185 genes. Our results illustrate the power of combining in-depth phenotyping with transcriptomics in mapping populations to dissect the genetic control of complex traits and present a set of candidate genes for use in biomass improvement. PMID:26754667

  17. Transcriptome analysis of sex-related genes in the blood clam Tegillarca granosa.

    PubMed

    Chen, Heng; Xiao, Guoqiang; Chai, Xueliang; Lin, Xingguan; Fang, Jun; Teng, Shuangshuang

    2017-01-01

    polymorphisms (SNPs) were identified from the RNA-seq results. This study provided the first complete gonadal transcriptome data for the blood clam and allowed us to search many aspects of gene sequence information, not limited to gender. This data will improve our understanding of the transcriptomics and reproductive biology of the blood clam. Furthermore, molecular markers such as SSRs and SNPs will be useful in the analysis of genetic evolution, bulked segregant analysis (BSA) and genome-wide association studies (GWAS). Our transcriptome data will therefore provide important genetic information for the breeding and conservation of germplasm.

  18. Transcriptome Analysis of Leaves, Flowers and Fruits Perisperm of Coffea arabica L. Reveals the Differential Expression of Genes Involved in Raffinose Biosynthesis

    PubMed Central

    dos Santos, Tiago Benedito; de Oliveira, Fernanda Freitas; Pot, David; Leroy, Thierry; Vieira, Luiz Gonzaga Esteves; Carazzolle, Marcelo Falsarella; Pereira, Gonçalo Amarante Guimarães

    2017-01-01

    Coffea arabica L. is an important crop in several developing countries. Despite its economic importance, minimal transcriptome data are available for fruit tissues, especially during fruit development where several compounds related to coffee quality are produced. To understand the molecular aspects related to coffee fruit and grain development, we report a large-scale transcriptome analysis of leaf, flower and perisperm fruit tissue development. Illumina sequencing yielded 41,881,572 high-quality filtered reads. De novo assembly generated 65,364 unigenes with an average length of 1,264 bp. A total of 24,548 unigenes were annotated as protein coding genes, including 12,560 full-length sequences. In the annotation process, we identified nine candidate genes related to the biosynthesis of raffinose family oligossacarides (RFOs). These sugars confer osmoprotection and are accumulated during initial fruit development. Four genes from this pathway had their transcriptional pattern validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Furthermore, we identified ~24,000 putative target sites for microRNAs (miRNAs) and 134 putative transcriptionally active transposable elements (TE) sequences in our dataset. This C. arabica transcriptomic atlas provides an important step for identifying candidate genes related to several coffee metabolic pathways, especially those related to fruit chemical composition and therefore beverage quality. Our results are the starting point for enhancing our knowledge about the coffee genes that are transcribed during the flowering and initial fruit development stages. PMID:28068432

  19. Transcriptome Analysis of Leaves, Flowers and Fruits Perisperm of Coffea arabica L. Reveals the Differential Expression of Genes Involved in Raffinose Biosynthesis.

    PubMed

    Ivamoto, Suzana Tiemi; Reis, Osvaldo; Domingues, Douglas Silva; Dos Santos, Tiago Benedito; de Oliveira, Fernanda Freitas; Pot, David; Leroy, Thierry; Vieira, Luiz Gonzaga Esteves; Carazzolle, Marcelo Falsarella; Pereira, Gonçalo Amarante Guimarães; Pereira, Luiz Filipe Protasio

    2017-01-01

    Coffea arabica L. is an important crop in several developing countries. Despite its economic importance, minimal transcriptome data are available for fruit tissues, especially during fruit development where several compounds related to coffee quality are produced. To understand the molecular aspects related to coffee fruit and grain development, we report a large-scale transcriptome analysis of leaf, flower and perisperm fruit tissue development. Illumina sequencing yielded 41,881,572 high-quality filtered reads. De novo assembly generated 65,364 unigenes with an average length of 1,264 bp. A total of 24,548 unigenes were annotated as protein coding genes, including 12,560 full-length sequences. In the annotation process, we identified nine candidate genes related to the biosynthesis of raffinose family oligossacarides (RFOs). These sugars confer osmoprotection and are accumulated during initial fruit development. Four genes from this pathway had their transcriptional pattern validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Furthermore, we identified ~24,000 putative target sites for microRNAs (miRNAs) and 134 putative transcriptionally active transposable elements (TE) sequences in our dataset. This C. arabica transcriptomic atlas provides an important step for identifying candidate genes related to several coffee metabolic pathways, especially those related to fruit chemical composition and therefore beverage quality. Our results are the starting point for enhancing our knowledge about the coffee genes that are transcribed during the flowering and initial fruit development stages.

  20. Efficient assembly and annotation of the transcriptome of catfish by RNA-Seq analysis of a doubled haploid homozygote

    PubMed Central

    2012-01-01

    assembly of the full-length transcripts. Conclusions The large set of transcripts assembled in this study is the most comprehensive set of genome resources ever developed from catfish, which will provide the much needed resources for functional genome research in catfish, serving as a reference transcriptome for genome annotation, analysis of gene duplication, gene family structures, and digital gene expression analysis. The putative set of duplicated genes provide a starting point for genome scale analysis of gene duplication in the catfish genome, and should be a valuable resource for comparative genome analysis, genome evolution, and genome function studies. PMID:23127152

  1. 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

  2. Transcriptomic Analysis Reveals Mechanisms of Sterile and Fertile Flower Differentiation and Development in Viburnum macrocephalum f. keteleeri

    PubMed Central

    Lu, Zhaogeng; Xu, Jing; Li, Weixing; Zhang, Li; Cui, Jiawen; He, Qingsong; Wang, Li; Jin, Biao

    2017-01-01

    Sterile and fertile flowers are an important evolutionary developmental (evo-devo) phenotype in angiosperm flowers, playing important roles in pollinator attraction and sexual reproductive success. However, the gene regulatory mechanisms underlying fertile and sterile flower differentiation and development remain largely unknown. Viburnum macrocephalum f. keteleeri, which possesses fertile and sterile flowers in a single inflorescence, is a useful candidate species for investigating the regulatory networks in differentiation and development. We developed a de novo-assembled flower reference transcriptome. Using RNA sequencing (RNA-seq), we compared the expression patterns of fertile and sterile flowers isolated from the same inflorescence over its rapid developmental stages. The flower reference transcriptome consisted of 105,683 non-redundant transcripts, of which 5,675 transcripts showed significant differential expression between fertile and sterile flowers. Combined with morphological and cytological changes between fertile and sterile flowers, we identified expression changes of many genes potentially involved in reproductive processes, phytohormone signaling, and cell proliferation and expansion using RNA-seq and qRT-PCR. In particular, many transcription factors (TFs), including MADS-box family members and ABCDE-class genes, were identified, and expression changes in TFs involved in multiple functions were analyzed and highlighted to determine their roles in regulating fertile and sterile flower differentiation and development. Our large-scale transcriptional analysis of fertile and sterile flowers revealed the dynamics of transcriptional networks and potentially key components in regulating differentiation and development of fertile and sterile flowers in Viburnum macrocephalum f. keteleeri. Our data provide a useful resource for Viburnum transcriptional research and offer insights into gene regulation of differentiation of diverse evo-devo processes in

  3. Exploration of rice pistil responses during early post-pollination through a combined proteomic and transcriptomic analysis.

    PubMed

    Li, Ming; Wang, Kun; Li, Shaoqing; Yang, Pingfang

    2016-01-10

    Pollen-stigma interaction is a multi-step and complex physiological process which contains different signaling and biochemical pathways. However, little is known about the molecular mechanism underlying this process in rice (Oryza sativa). In this study, the changes of gene expression were investigated through a combination study of transcriptome and proteome profiles in rice pistil during pollination. Totally, 1117 differentially expressed genes were identified, among which 962 and 167 were detected at transcriptional and protein level respectively. Functional categorization analysis showed that the genes involved in central metabolism were up-regulated, which can lead to the enhancement of these metabolisms. The reactive oxygen species (ROS) were over-accumulated in the stigma. In response to this, the proteins or transcripts involved in redox homeostasis regulation were differentially expressed. Furthermore, significant changes of protein ubiquitination and its related genes or proteins, especially some E3 ligases encoding genes, indicate that protein ubiquitination might play important roles in cell signal transduction during the pollination process. Our study sheds some lights on gene and protein expression profiles of rice pistil pollination process, and gives us a comprehensive understanding of the basic molecular mechanisms controlling pollination in rice. Using RNA-seq, 2-DE and iTRAQ assays, we have generated the large-scale transcriptomic and proteomic data containing abundant information on genes involved in pollen and pistil interaction. Our results showed that ROS were significantly accumulated in stigma after pollination, and the abundance of genes involve in redox homeostasis system were changed variously. We also show that, changes of some E3 ligases encoding genes might indicate that protein ubiquitination play important roles in cell signal transduction during the pollination process. Data in this study might be helpful to deeply understand the

  4. Transcriptomic Analysis Reveals Mechanisms of Sterile and Fertile Flower Differentiation and Development in Viburnum macrocephalum f. keteleeri.

    PubMed

    Lu, Zhaogeng; Xu, Jing; Li, Weixing; Zhang, Li; Cui, Jiawen; He, Qingsong; Wang, Li; Jin, Biao

    2017-01-01

    Sterile and fertile flowers are an important evolutionary developmental (evo-devo) phenotype in angiosperm flowers, playing important roles in pollinator attraction and sexual reproductive success. However, the gene regulatory mechanisms underlying fertile and sterile flower differentiation and development remain largely unknown. Viburnum macrocephalum f. keteleeri, which possesses fertile and sterile flowers in a single inflorescence, is a useful candidate species for investigating the regulatory networks in differentiation and development. We developed a de novo-assembled flower reference transcriptome. Using RNA sequencing (RNA-seq), we compared the expression patterns of fertile and sterile flowers isolated from the same inflorescence over its rapid developmental stages. The flower reference transcriptome consisted of 105,683 non-redundant transcripts, of which 5,675 transcripts showed significant differential expression between fertile and sterile flowers. Combined with morphological and cytological changes between fertile and sterile flowers, we identified expression changes of many genes potentially involved in reproductive processes, phytohormone signaling, and cell proliferation and expansion using RNA-seq and qRT-PCR. In particular, many transcription factors (TFs), including MADS-box family members and ABCDE-class genes, were identified, and expression changes in TFs involved in multiple functions were analyzed and highlighted to determine their roles in regulating fertile and sterile flower differentiation and development. Our large-scale transcriptional analysis of fertile and sterile flowers revealed the dynamics of transcriptional networks and potentially key components in regulating differentiation and development of fertile and sterile flowers in Viburnum macrocephalum f. keteleeri. Our data provide a useful resource for Viburnum transcriptional research and offer insights into gene regulation of differentiation of diverse evo-devo processes in

  5. Comprehensive transcriptome analysis of the highly complex Pisum sativum genome using next generation sequencing

    PubMed Central

    2011-01-01

    Background The garden pea, Pisum sativum, is among the best-investigated legume plants and of significant agro-commercial relevance. Pisum sativum has a large and complex genome and accordingly few comprehensive genomic resources exist. Results We analyzed the pea transcriptome at the highest possible amount of accuracy by current technology. We used next generation sequencing with the Roche/454 platform and evaluated and compared a variety of approaches, including diverse tissue libraries, normalization, alternative sequencing technologies, saturation estimation and diverse assembly strategies. We generated libraries from flowers, leaves, cotyledons, epi- and hypocotyl, and etiolated and light treated etiolated seedlings, comprising a total of 450 megabases. Libraries were assembled into 324,428 unigenes in a first pass assembly. A second pass assembly reduced the amount to 81,449 unigenes but caused a significant number of chimeras. Analyses of the assemblies identified the assembly step as a major possibility for improvement. By recording frequencies of Arabidopsis orthologs hit by randomly drawn reads and fitting parameters of the saturation curve we concluded that sequencing was exhaustive. For leaf libraries we found normalization allows partial recovery of expression strength aside the desired effect of increased coverage. Based on theoretical and biological considerations we concluded that the sequence reads in the database tagged the vast majority of transcripts in the aerial tissues. A pathway representation analysis showed the merits of sampling multiple aerial tissues to increase the number of tagged genes. All results have been made available as a fully annotated database in fasta format. Conclusions We conclude that the approach taken resulted in a high quality - dataset which serves well as a first comprehensive reference set for the model legume pea. We suggest future deep sequencing transcriptome projects of species lacking a genomics backbone will

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

    PubMed Central

    2012-01-01

    Background 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. Results 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. Conclusions 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

  7. Transcriptome-wide analysis supports environmental adaptations of two Pinus pinaster populations from contrasting habitats.

    PubMed

    Cañas, Rafael A; Feito, Isabel; Fuente-Maqueda, José Francisco; Ávila, Concepción; Majada, Juan; Cánovas, Francisco M

    2015-11-06

    Maritime pine (Pinus pinaster Aiton) grows in a range of different climates in the southwestern Mediterranean region and the existence of a variety of latitudinal ecotypes or provenances is well established. In this study, we have conducted a deep analysis of the transcriptome in needles from two P. pinaster provenances, Leiria (Portugal) and Tamrabta (Morocco), which were grown in northern Spain under the same conditions. An oligonucleotide microarray (PINARRAY3) and RNA-Seq were used for whole-transcriptome analyses, and we found that 90.95% of the data were concordant between the two platforms. Furthermore, the two methods identified very similar percentages of differentially expressed genes with values of 5.5% for PINARRAY3 and 5.7% for RNA-Seq. In total, 6,023 transcripts were shared and 88 differentially expressed genes overlapped in the two platforms. Among the differentially expressed genes, all transport related genes except aquaporins were expressed at higher levels in Tamrabta than in Leiria. In contrast, genes involved in secondary metabolism were expressed at higher levels in Tamrabta, and photosynthesis-related genes were expressed more highly in Leiria. The genes involved in light sensing in plants were well represented in the differentially expressed groups of genes. In addition, increased levels of hormones such as abscisic acid, gibberellins, jasmonic and salicylic acid were observed in Leiria. Both transcriptome platforms have proven to be useful resources, showing complementary and reliable results. The results presented here highlight the different abilities of the two maritime pine populations to sense environmental conditions and reveal one type of regulation that can be ascribed to different genetic and epigenetic backgrounds.

  8. Transcriptome Analysis of Syringa oblata Lindl. Inflorescence Identifies Genes Associated with Pigment Biosynthesis and Scent Metabolism

    PubMed Central

    Zheng, Jian; Hu, Zenghui; Guan, Xuelian; Dou, Dequan; Bai, Guo; Wang, Yu; Guo, Yingtian; Li, Wei; Leng, Pingsheng

    2015-01-01

    Syringa oblata Lindl. is a woody ornamental plant with high economic value and characteristics that include early flowering, multiple flower colors, and strong fragrance. Despite a long history of cultivation, the genetics and molecular biology of S. oblata are poorly understood. Transcriptome and expression profiling data are needed to identify genes and to better understand the biological mechanisms of floral pigments and scents in this species. Nine cDNA libraries were obtained from three replicates of three developmental stages: inflorescence with enlarged flower buds not protruded, inflorescence with corolla lobes not displayed, and inflorescence with flowers fully opened and emitting strong fragrance. Using the Illumina RNA-Seq technique, 319,425,972 clean reads were obtained and were assembled into 104,691 final unigenes (average length of 853 bp), 41.75% of which were annotated in the NCBI non-redundant protein database. Among the annotated unigenes, 36,967 were assigned to gene ontology categories and 19,956 were assigned to eukaryoticorthologous groups. Using the Kyoto Encyclopedia of Genes and Genomes pathway database, 12,388 unigenes were sorted into 286 pathways. Based on these transcriptomic data, we obtained a large number of candidate genes that were differentially expressed at different flower stages and that were related to floral pigment biosynthesis and fragrance metabolism. This comprehensive transcriptomic analysis provides fundamental information on the genes and pathways involved in flower secondary metabolism and development in S. oblata, providing a useful database for further research on S. oblata and other plants of genus Syringa. PMID:26587670

  9. Transcriptome Analysis of Syringa oblata Lindl. Inflorescence Identifies Genes Associated with Pigment Biosynthesis and Scent Metabolism.

    PubMed

    Zheng, Jian; Hu, Zenghui; Guan, Xuelian; Dou, Dequan; Bai, Guo; Wang, Yu; Guo, Yingtian; Li, Wei; Leng, Pingsheng

    2015-01-01

    Syringa oblata Lindl. is a woody ornamental plant with high economic value and characteristics that include early flowering, multiple flower colors, and strong fragrance. Despite a long history of cultivation, the genetics and molecular biology of S. oblata are poorly understood. Transcriptome and expression profiling data are needed to identify genes and to better understand the biological mechanisms of floral pigments and scents in this species. Nine cDNA libraries were obtained from three replicates of three developmental stages: inflorescence with enlarged flower buds not protruded, inflorescence with corolla lobes not displayed, and inflorescence with flowers fully opened and emitting strong fragrance. Using the Illumina RNA-Seq technique, 319,425,972 clean reads were obtained and were assembled into 104,691 final unigenes (average length of 853 bp), 41.75% of which were annotated in the NCBI non-redundant protein database. Among the annotated unigenes, 36,967 were assigned to gene ontology categories and 19,956 were assigned to eukaryoticorthologous groups. Using the Kyoto Encyclopedia of Genes and Genomes pathway database, 12,388 unigenes were sorted into 286 pathways. Based on these transcriptomic data, we obtained a large number of candidate genes that were differentially expressed at different flower stages and that were related to floral pigment biosynthesis and fragrance metabolism. This comprehensive transcriptomic analysis provides fundamental information on the genes and pathways involved in flower secondary metabolism and development in S. oblata, providing a useful database for further research on S. oblata and other plants of genus Syringa.

  10. Analysis of Litopenaeus vannamei Transcriptome Using the Next-Generation DNA Sequencing Technique

    PubMed Central

    Li, Chaozheng; Weng, Shaoping; Chen, Yonggui; Yu, Xiaoqiang; Lü, Ling; Zhang, Haiqing; He, Jianguo; Xu, Xiaopeng

    2012-01-01

    Background Pacific white shrimp (Litopenaeus vannamei), the major species of farmed shrimps in the world, has been attracting extensive studies, which require more and more genome background knowledge. The now available transcriptome data of L. vannamei are insufficient for research requirements, and have not been adequately assembled and annotated. Methodology/Principal Findings This is the first study that used a next-generation high-throughput DNA sequencing technique, the Solexa/Illumina GA II method, to analyze the transcriptome from whole bodies of L. vannamei larvae. More than 2.4 Gb of raw data were generated, and 109,169 unigenes with a mean length of 396 bp were assembled using the SOAP denovo software. 73,505 unigenes (>200 bp) with good quality sequences were selected and subjected to annotation analysis, among which 37.80% can be matched in NCBI Nr database, 37.3% matched in Swissprot, and 44.1% matched in TrEMBL. Using BLAST and BLAST2Go softwares, 11,153 unigenes were classified into 25 Clusters of Orthologous Groups of proteins (COG) categories, 8171 unigenes were assigned into 51 Gene ontology (GO) functional groups, and 18,154 unigenes were divided into 220 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. To primarily verify part of the results of assembly and annotations, 12 assembled unigenes that are homologous to many embryo development-related genes were chosen and subjected to RT-PCR for electrophoresis and Sanger sequencing analyses, and to real-time PCR for expression profile analyses during embryo development. Conclusions/Significance The L. vannamei transcriptome analyzed using the next-generation sequencing technique enriches the information of L. vannamei genes, which will facilitate our understanding of the genome background of crustaceans, and promote the studies on L. vannamei. PMID:23071809

  11. Deep Insight into the Ganoderma lucidum by Comprehensive Analysis of Its Transcriptome

    PubMed Central

    Yu, Guo-Jun; Wang, Man; Huang, Jie; Yin, Ya-Lin; Chen, Yi-Jie; Jiang, Shuai; Jin, Yan-Xia; Lan, Xian-Qing; Wong, Barry Hon Cheung; Liang, Yi; Sun, Hui

    2012-01-01

    Background Ganoderma lucidum is a basidiomycete white rot fungus and is of medicinal importance in China, Japan and other countries in the Asiatic region. To date, much research has been performed in identifying the medicinal ingredients in Ganoderma lucidum. Despite its important therapeutic effects in disease, little is known about Ganoderma lucidum at the genomic level. In order to gain a molecular understanding of this fungus, we utilized Illumina high-throughput technology to sequence and analyze the transcriptome of Ganoderma lucidum. Methodology/Principal Findings We obtained 6,439,690 and 6,416,670 high-quality reads from the mycelium and fruiting body of Ganoderma lucidum, and these were assembled to form 18,892 and 27,408 unigenes, respectively. A similarity search was performed against the NCBI non-redundant nucleotide database and a customized database composed of five fungal genomes. 11,098 and 8, 775 unigenes were matched to the NCBI non-redundant nucleotide database and our customized database, respectively. All unigenes were subjected to annotation by Gene Ontology, Eukaryotic Orthologous Group terms and Kyoto Encyclopedia of Genes and Genomes. Differentially expressed genes from the Ganoderma lucidum mycelium and fruiting body stage were analyzed, resulting in the identification of 13 unigenes which are involved in the terpenoid backbone biosynthesis pathway. Quantitative real-time PCR was used to confirm the expression levels of these unigenes. Ganoderma lucidum was also studied for wood degrading activity and a total of 22 putative FOLymes (fungal oxidative lignin enzymes) and 120 CAZymes (carbohydrate-active enzymes) were predicted from our Ganoderma lucidum transcriptome. Conclusions Our study provides comprehensive gene expression information on Ganoderma lucidum at the transcriptional level, which will form the foundation for functional genomics studies in this fungus. The use of Illumina sequencing technology has made de novo transcriptome

  12. Transcriptome analysis and identification of induced genes in the response of Harmonia axyridis to cold hardiness.

    PubMed

    Tang, Bin; Liu, Xiao-Jun; Shi, Zuo-Kun; Shen, Qi-Da; Xu, Yan-Xia; Wang, Su; Zhang, Fan; Wang, Shi-Gui

    2017-06-01

    Harmonia axyridis is an important predatory lady beetle that is a natural enemy of agricultural and forestry pests. In this research, the cold hardiness induced genes and their expression changes in H. axyridis were screened and detected by the way of the transcriptome and qualitative real-time PCR under normal and low temperatures, using high-throughput transcriptome and digital gene-expression-tag technologies. We obtained a 10Gb transcriptome and an 8Mb gene expression tag pool using Illumina deep sequencing technology and RNA-Seq analysis (accession number SRX540102). Of the 46,980 non-redundant unigenes identified, 28,037 (59.7%) were matched to known genes in GenBank, 21,604 (46.0%) in Swiss-Prot, 19,482 (41.5%) in Kyoto Encyclopedia of Genes and Genomes and 13,193 (28.1%) in Gene Ontology databases. Seventy-five percent of the unigene sequences had top matches with gene sequences from Tribolium castaneum. Results indicated that 60 genes regulated the entire cold-acclimation response, and, of these, seven genes were always up-regulated and five genes always down-regulated. Further screening revealed that six cold-resistant genes, E3 ubiquitin-protein ligase, transketolase, trehalase, serine/arginine repetitive matrix protein 2, glycerol kinase and sugar transporter SWEET1-like, play key roles in the response. Expression from a number of the differentially expressed genes was confirmed with quantitative real-time PCR (HaCS_Trans). The paper attempted to identify cold-resistance response genes, and study the potential mechanism by which cold acclimation enhances the insect's cold endurance. Information on these cold-resistance response genes will improve the development of low-temperature storage technology of natural enemy insects for future use in biological control. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Transcriptomic and Proteomic Analysis of Oenococcus oeni Adaptation to Wine Stress Conditions

    PubMed Central

    Margalef-Català, Mar; Araque, Isabel; Bordons, Albert; Reguant, Cristina; Bautista-Gallego, Joaquín

    2016-01-01

    Oenococcus oeni, the main lactic acid bacteria responsible for malolactic fermentation in wine, has to adapt to stressful conditions, such as low pH and high ethanol content. In this study, the changes in the transcriptome and the proteome of O. oeni PSU-1 during the adaptation period before MLF start have been studied. DNA microarrays were used for the transcriptomic analysis and two complementary proteomic techniques, 2-D DIGE and iTRAQ labeling were used to analyze the proteomic response. One of the most influenced functions in PSU-1 due to inoculation into wine-like medium (WLM) was translation, showing the over-expression of certain ribosomal genes and the corresponding proteins. Amino acid metabolism and transport was also altered and several peptidases were up regulated both at gene and protein level. Certain proteins involved in glutamine and glutamate metabolism showed an increased abundance revealing the key role of nitrogen uptake under stressful conditions. A strong transcriptional inhibition of carbohydrate metabolism related genes was observed. On the other hand, the transcriptional up-regulation of malate transport and citrate consumption was indicative of the use of L-malate and citrate associated to stress response and as an alternative energy source to sugar metabolism. Regarding the stress mechanisms, our results support the relevance of the thioredoxin and glutathione systems in the adaptation of O. oeni to wine related stress. Genes and proteins related to cell wall showed also significant changes indicating the relevance of the cell envelop as protective barrier to environmental stress. The differences found between transcriptomic and proteomic data suggested the relevance of post-transcriptional mechanisms and the complexity of the stress response in O. oeni adaptation. Further research should deepen into the metabolisms mostly altered due to wine conditions to elucidate the role of each mechanism in the O. oeni ability to develop MLF. PMID

  14. 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

  15. 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

  16. Transcriptome and Functional Analysis of Mating in the Basidiomycete Schizophyllum commune

    PubMed Central

    Erdmann, Susann; Freihorst, Daniela; Raudaskoski, Marjatta; Schmidt-Heck, Wolfgang; Jung, Elke-Martina; Senftleben, Dominik

    2012-01-01

    In this study, we undertook a functional characterization and transcriptome analysis that enabled a comprehensive study of the mating type loci of the mushroom Schizophyllum commune. Induced expression of both the bar2 receptor and the bap2(2) pheromone gene within 6 to 12 h after mates' contact was demonstrated by quantitative real-time PCR. Similar temporal expression patterns were confirmed for the allelic bbr1 receptor and bbp1 pheromone-encoding genes by Northern hybridization. Interestingly, the fusion of clamp connections to the subterminal cell was delayed in mating interactions in which one of the compatible partners expressed the bar2 receptor with a truncated C terminus. This developmental delay allowed the visualization of a green fluorescent protein (Gfp)-labeled truncated receptor at the cell periphery, consistent with a localization in the plasma membrane of unfused pseudoclamps. This finding does not support hypotheses envisioning a receptor localization to the nuclear membrane facilitating recognition between the two different nuclei present in each dikaryotic cell. Rather, Gfp fluorescence observed in such pseudoclamps indicated a role of receptor-pheromone interaction in clamp fusion. Transcriptome changes associated with mating interactions were analyzed in order to identify a role for pheromone-receptor interactions. We detected a total of 89 genes that were transcriptionally regulated in a mating type locus A-dependent manner, employing a cutoff of 5-fold changes in transcript abundance. Upregulation in cell cycle-related genes and downregulation of genes involved in metabolism were seen with this set of experiments. In contrast, mating type locus B-dependent transcriptome changes were observed in 208 genes, with a specific impact on genes related to cell wall and membrane metabolism, stress response, and the redox status of the cell. PMID:22210832

  17. Comparative transcriptome analysis of three color variants of the sea cucumber Apostichopus japonicus.

    PubMed

    Jo, Jihoon; Park, Jongsun; Lee, Hyun-Gwan; Kern, Elizabeth M A; Cheon, Seongmin; Jin, Soyeong; Park, Joong-Ki; Cho, Sung-Jin; Park, Chungoo

    2016-08-01

    The sea cucumber Apostichopus japonicus Selenka 1867 represents an important resource in biomedical research, traditional medicine, and the seafood industry. Much of the commercial value of A. japonicus is determined by dorsal/ventral color variation (red, green, and black), yet the taxonomic relationships between these color variants are not clearly understood. We performed the first comparative analysis of de novo assembled transcriptome data from three color variants of A. japonicus. Using the Illumina platform, we sequenced nearly 177,596,774 clean reads representing a total of 18.2Gbp of sea cucumber transcriptome. A comparison of over 0.3 million transcript scaffolds against the Uniprot/Swiss-Prot database yielded 8513, 8602, and 8588 positive matches for green, red, and black body color transcriptomes, respectively. Using the Panther gene classification system, we assessed an extensive and diverse set of expressed genes in three color variants and found that (1) among the three color variants of A. japonicus, genes associated with RNA binding protein, oxidoreductase, nucleic acid binding, transferase, and KRAB box transcription factor were most commonly expressed; and (2) the main protein functional classes are differently regulated in all three color variants (extracellular matrix protein and phosphatase for green color, transporter and potassium channel for red color, and G-protein modulator and enzyme modulator for black color). This work will assist in the discovery and annotation of novel genes that play significant morphological and physiological roles in color variants of A. japonicus, and these sequence data will provide a useful set of resources for the rapidly growing sea cucumber aquaculture industry.

  18. RNA-seq analysis and de novo transcriptome assembly of Jerusalem artichoke (Helianthus tuberosus Linne).

    PubMed

    Jung, Won Yong; Lee, Sang Sook; Kim, Chul Wook; Kim, Hyun-Soon; Min, Sung Ran; Moon, Jae Sun; Kwon, Suk-Yoon; Jeon, Jae-Heung; Cho, Hye Sun

    2014-01-01

    Jerusalem artichoke (Helianthus tuberosus L.) has long been cultivated as a vegetable and as a source of fructans (inulin) for pharmaceutical applications in diabetes and obesity prevention. However, transcriptomic and genomic data for Jerusalem artichoke remain scarce. In this study, Illumina RNA sequencing (RNA-Seq) was performed on samples from Jerusalem artichoke leaves, roots, stems and two different tuber tissues (early and late tuber development). Data were used for de novo assembly and characterization of the transcriptome. In total 206,215,632 paired-end reads were generated. These were assembled into 66,322 loci with 272,548 transcripts. Loci were annotated by querying against the NCBI non-redundant, Phytozome and UniProt databases, and 40,215 loci were homologous to existing database sequences. Gene Ontology terms were assigned to 19,848 loci, 15,434 loci were matched to 25 Clusters of Eukaryotic Orthologous Groups classifications, and 11,844 loci were classified into 142 Kyoto Encyclopedia of Genes and Genomes pathways. The assembled loci also contained 10,778 potential simple sequence repeats. The newly assembled transcriptome was used to identify loci with tissue-specific differential expression patterns. In total, 670 loci exhibited tissue-specific expression, and a subset of these were confirmed using RT-PCR and qRT-PCR. Gene expression related to inulin biosynthesis in tuber tissue was also investigated. Exsiting genetic and genomic data for H. tuberosus are scarce. The sequence resources developed in this study will enable the analysis of thousands of transcripts and will thus accelerate marker-assisted breeding studies and studies of inulin biosynthesis in Jerusalem artichoke.

  19. Transcriptome Profiling of Beach Morning Glory (Ipomoea imperati) under Salinity and Its Comparative Analysis with Sweetpotato

    PubMed Central

    Solis, Julio; Baisakh, Niranjan; Brandt, Steven R.; Villordon, Arthur; La Bonte, Don

    2016-01-01

    The response and adaption to salt remains poorly understood for beach morning glory [Ipomoea imperati (Vahl) Griseb], one of a few relatives of sweetpotato, known to thrive under salty and extreme drought conditions. In order to understand the genetic mechanisms underlying salt tolerance of a Convolvulaceae member, a genome-wide transcriptome study was carried out in beach morning glory by 454 pyrosequencing. A total of 286,584 filtered reads from both salt stressed and unstressed (control) root and shoot tissues were assembled into 95,790 unigenes with an average length of 667 base pairs (bp) and N50 of 706 bp. Putative differentially expressed genes (DEGs) were identified as transcripts overrepresented under salt stressed tissues compared to the control, and were placed into metabolic pathways. Most of these DEGs were involved in stress response, membrane transport, signal transduction, transcription activity and other cellular and molecular processes. We further analyzed the gene expression of 14 candidate genes of interest for salt tolerance through quantitative reverse transcription PCR (qRT-PCR) and confirmed their differential expression under salt stress in both beach morning glory and sweetpotato. The results comparing transcripts of I. imperati against the transcriptome of other Ipomoea species, including sweetpotato are also presented in this study. In addition, 6,233 SSR markers were identified, and an in silico analysis predicted that 434 primer pairs out of 4,897 target an identifiable homologous sequence in other Ipomoea transcriptomes, including sweetpotato. The data generated in this study will help in understanding the basics of salt tolerance of beach morning glory and the SSR resources generated will be useful for comparative genomics studies and further enhance the path to the marker-assisted breeding of sweetpotato for salt tolerance. PMID:26848754

  20. Insights into Vibrio parahaemolyticus CHN25 Response to Artificial Gastric Fluid Stress by Transcriptomic Analysis

    PubMed Central

    Sun, Xuejiao; Liu, Taigang; Peng, Xu; Chen, Lanming

    2014-01-01

    Vibrio parahaemolyticus is the causative agent of food-borne gastroenteritis disease. Once consumed, human acid gastric fluid is perhaps one of the most important environmental stresses imposed on the bacterium. Herein, for the first time, we investigated Vibrio parahaemolyticus CHN25 response to artificial gastric fluid (AGF) stress by transcriptomic analysis. The bacterium at logarithmic growth phase (LGP) displayed lower survival rates than that at stationary growth phase (SGP) under a sub-lethal acid condition (pH 4.9). Transcriptome data revealed that 11.6% of the expressed genes in Vibrio parahaemolyticus CHN25 was up-regulated in LGP cells after exposed to AGF (pH 4.9) for 30 min, including those involved in sugar transport, nitrogen metabolism, energy production and protein biosynthesis, whereas 14.0% of the genes was down-regulated, such as ATP-binding cassette (ABC) transporter and flagellar biosynthesis genes. In contrast, the AGF stress only elicited 3.4% of the genes from SGP cells, the majority of which were attenuated in expression. Moreover, the number of expressed regulator genes was also substantially reduced in SGP cells. Comparison of transcriptome profiles further revealed forty-one growth-phase independent genes in the AGF stress, however, half of which displayed distinct expression features between the two growth phases. Vibrio parahaemolyticus seemed to have evolved a number of molecular strategies for coping with the acid stress. The data here will facilitate future studies for environmental stresses and pathogenicity of the leading seafood-borne pathogen worldwide. PMID:25490137

  1. De Novo Transcriptome Sequencing and Analysis of the Cereal Cyst Nematode, Heterodera avenae

    PubMed Central

    Kumar, Mukesh; Gantasala, Nagavara Prasad; Roychowdhury, Tanmoy; Thakur, Prasoon Kumar; Banakar, Prakash; Shukla, Rohit N.; Jones, Michael G. K.; Rao, Uma

    2014-01-01

    The cereal cyst nematode (CCN, Heterodera avenae) is a major pest of wheat (Triticum spp) that reduces crop yields in many countries. Cyst nematodes are obligate sedentary endoparasites that reproduce by amphimixis. Here, we report the first transcriptome analysis of two stages of H. avenae. After sequencing extracted RNA from pre parasitic infective juvenile and adult stages of the life cycle, 131 million Illumina high quality paired end reads were obtained which generated 27,765 contigs with N50 of 1,028 base pairs, of which 10,452 were annotated. Comparative analyses were undertaken to evaluate H. avenae sequences with those of other plant, animal and free living nematodes to identify differences in expressed genes. There were 4,431 transcripts common to H. avenae and the free living nematode Caenorhabditis elegans, and 9,462 in common with more closely related potato cyst nematode, Globodera pallida. Annotation of H. avenae carbohydrate active enzymes (CAZy) revealed fewer glycoside hydrolases (GHs) but more glycosyl transferases (GTs) and carbohydrate esterases (CEs) when compared to M. incognita. 1,280 transcripts were found to have secretory signature, presence of signal peptide and absence of transmembrane. In a comparison of genes expressed in the pre-parasitic juvenile and feeding female stages, expression levels of 30 genes with high RPKM (reads per base per kilo million) value, were analysed by qRT-PCR which confirmed the observed differences in their levels of expression levels. In addition, we have also developed a user-friendly resource, Heterodera transcriptome database (HATdb) for public access of the data generated in this study. The new data provided on the transcriptome of H. avenae adds to the genetic resources available to study plant parasitic nematodes and provides an opportunity to seek new effectors that are specifically involved in the H. avenae-cereal host interaction. PMID:24802510

  2. Deep analysis of wild Vitis flower transcriptome reveals unexplored genome regions associated with sex specification.

    PubMed

    Ramos, Miguel Jesus Nunes; Coito, João Lucas; Fino, Joana; Cunha, Jorge; Silva, Helena; de Almeida, Patrícia Gomes; Costa, Maria Manuela Ribeiro; Amâncio, Sara; Paulo, Octávio S; Rocheta, Margarida

    2017-01-01

    RNA-seq of Vitis during early stages of bud development, in male, female and hermaphrodite flowers, identified new loci outside of annotated gene models, suggesting their involvement in sex establishment. The molecular mechanisms responsible for flower sex specification remain unclear for most plant species. In the case of V. vinifera ssp. vinifera, it is not fully understood what determines hermaphroditism in the domesticated subspecies and male or female flowers in wild dioecious relatives (Vitis vinifera ssp. sylvestris). Here, we describe a de novo assembly of the transcriptome of three flower developmental stages from the three Vitis vinifera flower types. The validation of de novo assembly showed a correlation of 0.825. The main goals of this work were the identification of V. v. sylvestris exclusive transcripts and the characterization of differential gene expression during flower development. RNA from several flower developmental stages was used previously to generate Illumina sequence reads. Through a sequential de novo assembly strategy one comprehensive transcriptome comprising 95,516 non-redundant transcripts was assembled. From this dataset 81,064 transcripts were annotated to V. v. vinifera reference transcriptome and 11,084 were annotated against V. v. vinifera reference genome. Moreover, we found 3368 transcripts that could not be mapped to Vitis reference genome. From all the non-redundant transcripts that were assembled, bioinformatics analysis identified 133 specific of V. v. sylvestris and 516 transcripts differentially expressed among the three flower types. The detection of transcription from areas of the genome not currently annotated suggests active transcription of previously unannotated genomic loci during early stages of bud development.

  3. Transcriptome analysis of an endoparasitoid wasp Cotesia chilonis (Hymenoptera: Braconidae) reveals genes involved in successful parasitism.

    PubMed

    Qi, Yixiang; Teng, Ziwen; Gao, Lingfeng; Wu, Shunfan; Huang, Jia; Ye, Gongyin; Fang, Qi

    2015-04-01

    For successful parasitization, parasitiods usually depend on the chemosensory cues for the selection of hosts, as well as a variety of virulence factors introduced into their hosts to overcome host immunity and prevent rejection of progeny development. In bracovirus-carrying wasps, the symbiotic polydnaviruses act in manipulating development and immunity of hosts. The endoparasitoid Cotesia chilonis carrying bracovirus as a key host immunosuppressive factor is a superior endoparasitoid of rice stem borer, Chilo suppressalis. So far, genomic information for C. chilonis is not available and transcriptomic data may provide valuable resources for global studying on physiological processes of C. chilonis, including chemosensation and parasitism at molecular level. Here, we performed RNA-seq to characterize the transcriptome of C. chilonis adults. We obtained 27,717,892 reads, assembled into 38,318 unigenes with a mean size of 690 bp. Approximately, 62.1% of the unigenes were annotated using NCBI databases. A large number of chemoreception-related genes encoding proteins including odorant receptors, gustatory receptors, odorant-binding proteins, chemosensory proteins, transient receptor potential ion channels, and sensory neuron membrane proteins were identified in silico. Totally, 72 transcripts possessing high identities with the bracovirus-related genes were identified. We investigated the mRNA expression levels of several transcripts at different developmental stages (including egg, larva, pupae, and adult) by quantitative real-time PCR analysis. The results revealed that some genes had adult-specific expression, indicating their potential significance for mating and parasitism. Overall, these results provide comprehensive insights into transcriptomic data of a polydnavirus-carrying parasitoid of a rice pest. © 2014 Wiley Periodicals, Inc.

  4. Elucidating fish oil-induced milk fat depression in dairy sheep: Milk somatic cell transcriptome analysis.

    PubMed

    Suárez-Vega, Aroa; Toral, Pablo G; Gutiérrez-Gil, Beatriz; Hervás, Gonzalo; Arranz, Juan José; Frutos, Pilar

    2017-04-05

    In this study, RNA sequencing was used to obtain a comprehensive profile of the transcriptomic changes occurring in the mammary gland of lactating sheep suffering from fish oil-induced milk fat depression (FO-MFD). The milk somatic cell transcriptome analysis of four control and four FO-MFD ewes generated an average of 42 million paired-end reads per sample. In both conditions, less than 220 genes constitute approximately 89% of the total counts. These genes, which are considered as core genes, were mainly involved in cytoplasmic ribosomal proteins and electron transport chain pathways. In total, 117 genes were upregulated, and 96 genes were downregulated in FO-MFD samples. Functional analysis of the latter indicated a downregulation of genes involved in the SREBP signaling pathway (e.g., ACACA, ACSL, and ACSS) and Gene Ontology terms related to lipid metabolism and lipid biosynthetic processes. Integrated interpretation of upregulated genes indicated enrichment in genes encoding plasma membrane proteins and proteins regulating protein kinase activity. Overall, our results indicate that FO-MFD is associated with the downregulation of key genes involved in the mammary lipogenesis process. In addition, the results also suggest that this syndrome may be related to upregulation of other genes implicated in signal transduction and codification of transcription factors.

  5. 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.

  6. Comparative transcriptome and proteome analysis to reveal the biosynthesis of gold nanoparticles in Arabidopsis

    PubMed Central

    Tiwari, Manish; Krishnamurthy, Sneha; Shukla, Devesh; Kiiskila, Jeffrey; Jain, Ajay; Datta, Rupali; Sharma, Nilesh; Sahi, Shivendra V.

    2016-01-01

    A large number of plants have been tested and exploited in search of a green chemistry approach for the fabrication of gold or other precious metal nanomaterials. Despite the potential of plant based methods, very little is known about the underlying biochemical reactions and genes involved in the biotransformation mechanism of AuCl4 into gold nanoparticles (AuNPs). In this research, we thus focused on studying the effect of Au on growth and nanoparticles formation by analyses of transcriptome, proteome and ionome shift in Arabidopsis. Au exposure favored the growth of Arabidopsis seedling and induced formation of nanoparticles in root and shoot, as indicated by optical and hyperspectral imaging. Root transcriptome analysis demonstrated the differential expression of the members of WRKY, MYB and BHLH gene families, which are involved in the Fe and other essential metals homeostasis. The proteome analysis revealed that Glutathione S-transferases were induced in the shoot and suggested its potential role in the biosynthesis AuNPs. This study also demonstrated the role of plant hormone auxin in determining the Au induced root system architecture. This is the first study using an integrated approach to understand the in planta biotransformation of KAuCl4 into AuNPs. PMID:26902325

  7. Elucidating fish oil-induced milk fat depression in dairy sheep: Milk somatic cell transcriptome analysis

    PubMed Central

    Suárez-Vega, Aroa; Toral, Pablo G.; Gutiérrez-Gil, Beatriz; Hervás, Gonzalo; Arranz, Juan José; Frutos, Pilar

    2017-01-01

    In this study, RNA sequencing was used to obtain a comprehensive profile of the transcriptomic changes occurring in the mammary gland of lactating sheep suffering from fish oil-induced milk fat depression (FO-MFD). The milk somatic cell transcriptome analysis of four control and four FO-MFD ewes generated an average of 42 million paired-end reads per sample. In both conditions, less than 220 genes constitute approximately 89% of the total counts. These genes, which are considered as core genes, were mainly involved in cytoplasmic ribosomal proteins and electron transport chain pathways. In total, 117 genes were upregulated, and 96 genes were downregulated in FO-MFD samples. Functional analysis of the latter indicated a downregulation of genes involved in the SREBP signaling pathway (e.g., ACACA, ACSL, and ACSS) and Gene Ontology terms related to lipid metabolism and lipid biosynthetic processes. Integrated interpretation of upregulated genes indicated enrichment in genes encoding plasma membrane proteins and proteins regulating protein kinase activity. Overall, our results indicate that FO-MFD is associated with the downregulation of key genes involved in the mammary lipogenesis process. In addition, the results also suggest that this syndrome may be related to upregulation of other genes implicated in signal transduction and codification of transcription factors. PMID:28378756

  8. Interactome-transcriptome analysis discovers signatures complementary to GWAS Loci of Type 2 Diabetes

    PubMed Central

    Li, Jing-Woei; Lee, Heung-Man; Wang, Ying; Tong, Amy Hin-Yan; Yip, Kevin Y.; Tsui, Stephen Kwok-Wing; Lok, Si; Ozaki, Risa; Luk, Andrea O; Kong, Alice P. S.; So, Wing-Yee; Ma, Ronald C. W.; Chan, Juliana C. N.; Chan, Ting-Fung

    2016-01-01

    Protein interactions play significant roles in complex diseases. We analyzed peripheral blood mononuclear cells (PBMC) transcriptome using a multi-method strategy. We constructed a tissue-specific interactome (T2Di) and identified 420 molecular signatures associated with T2D-related comorbidity and symptoms, mainly implicated in inflammation, adipogenesis, protein phosphorylation and hormonal secretion. Apart from explaining the residual associations within the DIAbetes Genetics Replication And Meta-analysis (DIAGRAM) study, the T2Di signatures were enriched in pathogenic cell type-specific regulatory elements related to fetal development, immunity and expression quantitative trait loci (eQTL). The T2Di revealed a novel locus near a well-established GWAS loci AChE, in which SRRT interacts with JAZF1, a T2D-GWAS gene implicated in pancreatic function. The T2Di also included known anti-diabetic drug targets (e.g. PPARD, MAOB) and identified possible druggable targets (e.g. NCOR2, PDGFR). These T2Di signatures were validated by an independent computational method, and by expression data of pancreatic islet, muscle and liver with some of the signatures (CEBPB, SREBF1, MLST8, SRF, SRRT and SLC12A9) confirmed in PBMC from an independent cohort of 66 T2D and 66 control subjects. By combining prior knowledge and transcriptome analysis, we have constructed an interactome to explain the multi-layered regulatory pathways in T2D. PMID:27752041

  9. Transcriptomic and proteomic analysis of Oenococcus oeni PSU-1 response to ethanol shock.

    PubMed

    Olguín, Nair; Champomier-Vergès, Marie; Anglade, Patricia; Baraige, Fabienne; Cordero-Otero, Ricardo; Bordons, Albert; Zagorec, Monique; Reguant, Cristina

    2015-10-01

    The correct development of malolactic fermentation depends on the capacity of Oenococcus oeni to survive under harsh wine conditions. The presence of ethanol is one of the most stressful factors affecting O. oeni performance. In this study, the effect of ethanol addition (12% vol/vol) on O. oeni PSU-1 has been evaluated using a transcriptomic and proteomic approach. Transcriptomic analysis revealed that the main functional categories of the genes affected