Science.gov

Sample records for quantified gene expression

  1. Quantifying the Effect of DNA Packaging on Gene Expression Level

    NASA Astrophysics Data System (ADS)

    Kim, Harold

    2010-10-01

    Gene expression, the process by which the genetic code comes alive in the form of proteins, is one of the most important biological processes in living cells, and begins when transcription factors bind to specific DNA sequences in the promoter region upstream of a gene. The relationship between gene expression output and transcription factor input which is termed the gene regulation function is specific to each promoter, and predicting this gene regulation function from the locations of transcription factor binding sites is one of the challenges in biology. In eukaryotic organisms (for example, animals, plants, fungi etc), DNA is highly compacted into nucleosomes, 147-bp segments of DNA tightly wrapped around histone protein core, and therefore, the accessibility of transcription factor binding sites depends on their locations with respect to nucleosomes - sites inside nucleosomes are less accessible than those outside nucleosomes. To understand how transcription factor binding sites contribute to gene expression in a quantitative manner, we obtain gene regulation functions of promoters with various configurations of transcription factor binding sites by using fluorescent protein reporters to measure transcription factor input and gene expression output in single yeast cells. In this talk, I will show that the affinity of a transcription factor binding site inside and outside the nucleosome controls different aspects of the gene regulation function, and explain this finding based on a mass-action kinetic model that includes competition between nucleosomes and transcription factors.

  2. Ribosomal protein L7 as a suitable reference gene for quantifying gene expression in gastropod Bellamya aeruginosa.

    PubMed

    Liu, Qing; Lei, Kun; Ma, Qingqing; Qiao, Fei; Li, Zi-Cheng; An, Li-Hui

    2016-04-01

    Expression levels of eight candidate reference genes were quantified in tissues of gastropod Bellamya aeruginosa exposed for 10 d to various stressors, including fasting, 17β-estradiol, 17α-methyltestosterone, and Cd(2+). The results showed that 18s rRNA was the most highly expressed of the candidate reference genes, while H2A was the least expressed. There were no significant changes (p>0.05) in the expression of the eight genes in tissues among the different treatments. Using RefFinder to evaluate the expression stabilities of the eight candidate reference genes, ribosomal protein was shown to be the most stable reference gene, and no effects were observed among the different stressor treatments. These results indicate that RPL 7 is the most suitable reference gene for quantifying gene expression in B. aeruginosa under environmental stress, which was verified in B. aeruginosa exposed to high doses of E2 for 24 and 72h. PMID:26991845

  3. A structured population modeling framework for quantifying and predicting gene expression noise in flow cytometry data.

    PubMed

    Flores, Kevin B

    2013-07-01

    We formulated a structured population model with distributed parameters to identify mechanisms that contribute to gene expression noise in time-dependent flow cytometry data. The model was validated using cell population-level gene expression data from two experiments with synthetically engineered eukaryotic cells. Our model captures the qualitative noise features of both experiments and accurately fit the data from the first experiment. Our results suggest that cellular switching between high and low expression states and transcriptional re-initiation are important factors needed to accurately describe gene expression noise with a structured population model.

  4. Shaped singular spectrum analysis for quantifying gene expression, with application to the early Drosophila embryo.

    PubMed

    Shlemov, Alex; Golyandina, Nina; Holloway, David; Spirov, Alexander

    2015-01-01

    In recent years, with the development of automated microscopy technologies, the volume and complexity of image data on gene expression have increased tremendously. The only way to analyze quantitatively and comprehensively such biological data is by developing and applying new sophisticated mathematical approaches. Here, we present extensions of 2D singular spectrum analysis (2D-SSA) for application to 2D and 3D datasets of embryo images. These extensions, circular and shaped 2D-SSA, are applied to gene expression in the nuclear layer just under the surface of the Drosophila (fruit fly) embryo. We consider the commonly used cylindrical projection of the ellipsoidal Drosophila embryo. We demonstrate how circular and shaped versions of 2D-SSA help to decompose expression data into identifiable components (such as trend and noise), as well as separating signals from different genes. Detection and improvement of under- and overcorrection in multichannel imaging is addressed, as well as the extraction and analysis of 3D features in 3D gene expression patterns.

  5. Shaped Singular Spectrum Analysis for Quantifying Gene Expression, with Application to the Early Drosophila Embryo

    PubMed Central

    Holloway, David

    2015-01-01

    In recent years, with the development of automated microscopy technologies, the volume and complexity of image data on gene expression have increased tremendously. The only way to analyze quantitatively and comprehensively such biological data is by developing and applying new sophisticated mathematical approaches. Here, we present extensions of 2D singular spectrum analysis (2D-SSA) for application to 2D and 3D datasets of embryo images. These extensions, circular and shaped 2D-SSA, are applied to gene expression in the nuclear layer just under the surface of the Drosophila (fruit fly) embryo. We consider the commonly used cylindrical projection of the ellipsoidal Drosophila embryo. We demonstrate how circular and shaped versions of 2D-SSA help to decompose expression data into identifiable components (such as trend and noise), as well as separating signals from different genes. Detection and improvement of under- and overcorrection in multichannel imaging is addressed, as well as the extraction and analysis of 3D features in 3D gene expression patterns. PMID:25945341

  6. SuperFly: a comparative database for quantified spatio-temporal gene expression patterns in early dipteran embryos.

    PubMed

    Cicin-Sain, Damjan; Pulido, Antonio Hermoso; Crombach, Anton; Wotton, Karl R; Jiménez-Guri, Eva; Taly, Jean-François; Roma, Guglielmo; Jaeger, Johannes

    2015-01-01

    We present SuperFly (http://superfly.crg.eu), a relational database for quantified spatio-temporal expression data of segmentation genes during early development in different species of dipteran insects (flies, midges and mosquitoes). SuperFly has a special focus on emerging non-drosophilid model systems. The database currently includes data of high spatio-temporal resolution for three species: the vinegar fly Drosophila melanogaster, the scuttle fly Megaselia abdita and the moth midge Clogmia albipunctata. At this point, SuperFly covers up to 9 genes and 16 time points per species, with a total of 1823 individual embryos. It provides an intuitive web interface, enabling the user to query and access original embryo images, quantified expression profiles, extracted positions of expression boundaries and integrated datasets, plus metadata and intermediate processing steps. SuperFly is a valuable new resource for the quantitative comparative study of gene expression patterns across dipteran species. Moreover, it provides an interesting test set for systems biologists interested in fitting mathematical gene network models to data. Both of these aspects are essential ingredients for progress toward a more quantitative and mechanistic understanding of developmental evolution.

  7. SuperFly: a comparative database for quantified spatio-temporal gene expression patterns in early dipteran embryos.

    PubMed

    Cicin-Sain, Damjan; Pulido, Antonio Hermoso; Crombach, Anton; Wotton, Karl R; Jiménez-Guri, Eva; Taly, Jean-François; Roma, Guglielmo; Jaeger, Johannes

    2015-01-01

    We present SuperFly (http://superfly.crg.eu), a relational database for quantified spatio-temporal expression data of segmentation genes during early development in different species of dipteran insects (flies, midges and mosquitoes). SuperFly has a special focus on emerging non-drosophilid model systems. The database currently includes data of high spatio-temporal resolution for three species: the vinegar fly Drosophila melanogaster, the scuttle fly Megaselia abdita and the moth midge Clogmia albipunctata. At this point, SuperFly covers up to 9 genes and 16 time points per species, with a total of 1823 individual embryos. It provides an intuitive web interface, enabling the user to query and access original embryo images, quantified expression profiles, extracted positions of expression boundaries and integrated datasets, plus metadata and intermediate processing steps. SuperFly is a valuable new resource for the quantitative comparative study of gene expression patterns across dipteran species. Moreover, it provides an interesting test set for systems biologists interested in fitting mathematical gene network models to data. Both of these aspects are essential ingredients for progress toward a more quantitative and mechanistic understanding of developmental evolution. PMID:25404137

  8. Sequencing and Validation of Reference Genes to Analyze Endogenous Gene Expression and Quantify Yellow Dwarf Viruses Using RT-qPCR in Viruliferous Rhopalosiphum padi

    PubMed Central

    Wu, Keke; Liu, Wenwen; Mar, Thithi; Liu, Yan; Wu, Yunfeng; Wang, Xifeng

    2014-01-01

    The bird cherry-oat aphid (Rhopalosiphum padi), an important pest of cereal crops, not only directly sucks sap from plants, but also transmits a number of plant viruses, collectively the yellow dwarf viruses (YDVs). For quantifying changes in gene expression in vector aphids, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a touchstone method, but the selection and validation of housekeeping genes (HKGs) as reference genes to normalize the expression level of endogenous genes of the vector and for exogenous genes of the virus in the aphids is critical to obtaining valid results. Such an assessment has not been done, however, for R. padi and YDVs. Here, we tested three algorithms (GeNorm, NormFinder and BestKeeper) to assess the suitability of candidate reference genes (EF-1α, ACT1, GAPDH, 18S rRNA) in 6 combinations of YDV and vector aphid morph. EF-1α and ACT1 together or in combination with GAPDH or with GAPDH and 18S rRNA could confidently be used to normalize virus titre and expression levels of endogenous genes in winged or wingless R. padi infected with Barley yellow dwarf virus isolates (BYDV)-PAV and BYDV-GAV. The use of only one reference gene, whether the most stably expressed (EF-1α) or the least stably expressed (18S rRNA), was not adequate for obtaining valid relative expression data from the RT-qPCR. Because of discrepancies among values for changes in relative expression obtained using 3 regions of the same gene, different regions of an endogenous aphid gene, including each terminus and the middle, should be analyzed at the same time with RT-qPCR. Our results highlight the necessity of choosing the best reference genes to obtain valid experimental data and provide several HKGs for relative quantification of virus titre in YDV-viruliferous aphids. PMID:24810421

  9. Sequencing and validation of reference genes to analyze endogenous gene expression and quantify yellow dwarf viruses using RT-qPCR in viruliferous Rhopalosiphum padi.

    PubMed

    Wu, Keke; Liu, Wenwen; Mar, Thithi; Liu, Yan; Wu, Yunfeng; Wang, Xifeng

    2014-01-01

    The bird cherry-oat aphid (Rhopalosiphum padi), an important pest of cereal crops, not only directly sucks sap from plants, but also transmits a number of plant viruses, collectively the yellow dwarf viruses (YDVs). For quantifying changes in gene expression in vector aphids, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a touchstone method, but the selection and validation of housekeeping genes (HKGs) as reference genes to normalize the expression level of endogenous genes of the vector and for exogenous genes of the virus in the aphids is critical to obtaining valid results. Such an assessment has not been done, however, for R. padi and YDVs. Here, we tested three algorithms (GeNorm, NormFinder and BestKeeper) to assess the suitability of candidate reference genes (EF-1α, ACT1, GAPDH, 18S rRNA) in 6 combinations of YDV and vector aphid morph. EF-1α and ACT1 together or in combination with GAPDH or with GAPDH and 18S rRNA could confidently be used to normalize virus titre and expression levels of endogenous genes in winged or wingless R. padi infected with Barley yellow dwarf virus isolates (BYDV)-PAV and BYDV-GAV. The use of only one reference gene, whether the most stably expressed (EF-1α) or the least stably expressed (18S rRNA), was not adequate for obtaining valid relative expression data from the RT-qPCR. Because of discrepancies among values for changes in relative expression obtained using 3 regions of the same gene, different regions of an endogenous aphid gene, including each terminus and the middle, should be analyzed at the same time with RT-qPCR. Our results highlight the necessity of choosing the best reference genes to obtain valid experimental data and provide several HKGs for relative quantification of virus titre in YDV-viruliferous aphids. PMID:24810421

  10. The use of a real-time luciferase assay to quantify gene expression dynamics in the living yeast cell.

    PubMed

    Rienzo, Alessandro; Pascual-Ahuir, Amparo; Proft, Markus

    2012-06-01

    A destabilized version of firefly luciferase was used in living yeast cells as a real-time reporter for gene expression. This highly sensitive and non-invasive system can be simultaneously used upon many different experimental conditions in small culture aliquots. This allows the dose-response behaviour of gene expression driven by any yeast promoter to be reported and can be used to quantify important parameters, such as the threshold, sensitivity, response time, maximal activity and synthesis rate for a given stimulus. We applied the luciferase assay to the nutrient-regulated GAL1 promoter and the stress-responsive GRE2 promoter. We find that luciferase expression driven by the GAL1 promoter responds dynamically to growing galactose concentrations, with increasing synthesis rates determined by the light increment in the initial linear phase of activation. In the case of the GRE2 promoter, we demonstrate that the very short-lived version of luciferase used here is an excellent tool to quantitatively describe transient transcriptional activation. The luciferase expression controlled by the GRE2 promoter responds dynamically to a gradual increase of osmotic or oxidative stress stimuli, which is mainly based on the progressive increase of the time the promoter remains active. Finally, we determined the dose-response behaviour of a single transcription factor binding site in a synthetic promoter context, using the stress response element (STRE) as an example. Taken together, the luciferase assay described here is an attractive tool to rapidly and precisely determine and compare kinetic parameters of gene expression.

  11. Shaped 3D singular spectrum analysis for quantifying gene expression, with application to the early zebrafish embryo.

    PubMed

    Shlemov, Alex; Golyandina, Nina; Holloway, David; Spirov, Alexander

    2015-01-01

    Recent progress in microscopy technologies, biological markers, and automated processing methods is making possible the development of gene expression atlases at cellular-level resolution over whole embryos. Raw data on gene expression is usually very noisy. This noise comes from both experimental (technical/methodological) and true biological sources (from stochastic biochemical processes). In addition, the cells or nuclei being imaged are irregularly arranged in 3D space. This makes the processing, extraction, and study of expression signals and intrinsic biological noise a serious challenge for 3D data, requiring new computational approaches. Here, we present a new approach for studying gene expression in nuclei located in a thick layer around a spherical surface. The method includes depth equalization on the sphere, flattening, interpolation to a regular grid, pattern extraction by Shaped 3D singular spectrum analysis (SSA), and interpolation back to original nuclear positions. The approach is demonstrated on several examples of gene expression in the zebrafish egg (a model system in vertebrate development). The method is tested on several different data geometries (e.g., nuclear positions) and different forms of gene expression patterns. Fully 3D datasets for developmental gene expression are becoming increasingly available; we discuss the prospects of applying 3D-SSA to data processing and analysis in this growing field.

  12. Real-Time RT-PCR assay to quantify the expression of fum1 and fum19 genes from the Fumonisin-producing Fusarium verticillioides.

    PubMed

    López-Errasquín, Elena; Vázquez, Covadonga; Jiménez, Misericordia; González-Jaén, Maria Teresa

    2007-02-01

    Fumonisins are a group of mycotoxins produced by Fusarium species of the Gibberella fujikuroi species complex that contaminate food and feed products, and represent a risk for human and animal health. In this work, we have developed a specific real-time reverse transcription-PCR (RT-PCR) assay to quantify the level of expression of two genes of the fumonisin biosynthetic cluster in F. verticillioides: fum1 (that encodes a polyketide synthase enzyme) and the ABC transporter encoding gene fum19. The level of expression of both genes was compared with the amount of fumonisin B(1) (FB(1)), measured by HPLC, produced by several strains of F. verticillioides in liquid culture. The results indicated a good correspondence between the levels of fum1 and fum19 expression and the production of fumonisin B(1). The analysis described provides a good approach for the rapid and specific detection and characterization of the potential ability of F. verticillioides strains to produce fumonisins.

  13. Gene Expression Correlates with Process Rates Quantified for Sulfate- and Fe(III)-Reducing Bacteria in U(VI)-Contaminated Sediments

    PubMed Central

    Akob, Denise M.; Lee, Sang Hyon; Sheth, Mili; Küsel, Kirsten; Watson, David B.; Palumbo, Anthony V.; Kostka, Joel E.; Chin, Kuk-Jeong

    2012-01-01

    Though iron- and sulfate-reducing bacteria are well known for mediating uranium(VI) reduction in contaminated subsurface environments, quantifying the in situ activity of the microbial groups responsible remains a challenge. The objective of this study was to demonstrate the use of quantitative molecular tools that target mRNA transcripts of key genes related to Fe(III) and sulfate reduction pathways in order to monitor these processes during in situ U(VI) remediation in the subsurface. Expression of the Geobacteraceae-specific citrate synthase gene (gltA) and the dissimilatory (bi)sulfite reductase gene (dsrA), were correlated with the activity of iron- or sulfate-reducing microorganisms, respectively, under stimulated bioremediation conditions in microcosms of sediments sampled from the U.S. Department of Energy’s Oak Ridge Integrated Field Research Challenge (OR-IFRC) site at Oak Ridge, TN, USA. In addition, Geobacteraceae-specific gltA and dsrA transcript levels were determined in parallel with the predominant electron acceptors present in moderately and highly contaminated subsurface sediments from the OR-IFRC. Phylogenetic analysis of the cDNA generated from dsrA mRNA, sulfate-reducing bacteria-specific 16S rRNA, and gltA mRNA identified activity of specific microbial groups. Active sulfate reducers were members of the Desulfovibrio, Desulfobacterium, and Desulfotomaculum genera. Members of the subsurface Geobacter clade, closely related to uranium-reducing Geobacter uraniireducens and Geobacter daltonii, were the metabolically active iron-reducers in biostimulated microcosms and in situ core samples. Direct correlation of transcripts and process rates demonstrated evidence of competition between the functional guilds in subsurface sediments. We further showed that active populations of Fe(III)-reducing bacteria and sulfate-reducing bacteria are present in OR-IFRC sediments and are good potential targets for in situ bioremediation. PMID:22908009

  14. Quantifying Temporal Autocorrelations for the Expression of Geobacter species mRNA Gene Transcripts at Variable Ammonium Levels during in situ U(VI) Bioremediation

    NASA Astrophysics Data System (ADS)

    Mouser, P. J.

    2010-12-01

    In order to develop decision-making tools for the prediction and optimization of subsurface bioremediation strategies, we must be able to link the molecular-scale activity of microorganisms involved in remediation processes with biogeochemical processes observed at the field-scale. This requires the ability to quantify changes in the in situ metabolic condition of dominant microbes and associate these changes to fluctuations in nutrient levels throughout the bioremediation process. It also necessitates a need to understand the spatiotemporal variability of the molecular-scale information to develop meaningful parameters and constraint ranges in complex bio-physio-chemical models. The expression of three Geobacter species genes (ammonium transporter (amtB), nitrogen fixation (nifD), and a housekeeping gene (recA)) were tracked at two monitoring locations that differed significantly in ammonium (NH4+) concentrations during a field-scale experiment where acetate was injected into the subsurface to simulate Geobacteraceae in a uranium-contaminated aquifer. Analysis of amtB and nifD mRNA transcript levels indicated that NH4+ was the primary form of fixed nitrogen during bioremediation. Overall expression levels of amtB were on average 8-fold higher at NH4+ concentrations of 300 μM or more than at lower NH4+ levels (average 60 μM). The degree of temporal correlation in Geobacter species mRNA expression levels was calculated at both locations using autocorrelation methods that describe the relationship between sample semi-variance and time lag. At the monitoring location with lower NH4+, a temporal correlation lag of 8 days was observed for both amtB and nifD transcript patterns. At the location where higher NH4+ levels were observed, no discernable temporal correlation lag above the sampling frequency (approximately every 2 days) was observed for amtB or nifD transcript fluctuations. Autocorrelation trends in recA expression levels at both locations indicated that

  15. Potential for Quantifying Expression of the Geobacteraceae Citrate Synthase Gene To Assess the Activity of Geobacteraceae in the Subsurface and on Current-Harvesting Electrodes

    PubMed Central

    Holmes, Dawn E.; Nevin, Kelly P.; O'Neil, Regina A.; Ward, Joy E.; Adams, Lorrie A.; Woodard, Trevor L.; Vrionis, Helen A.; Lovley, Derek R.

    2005-01-01

    The Geobacteraceae citrate synthase is phylogenetically distinct from those of other prokaryotes and is a key enzyme in the central metabolism of Geobacteraceae. Therefore, the potential for using levels of citrate synthase mRNA to estimate rates of Geobacter metabolism was evaluated in pure culture studies and in four different Geobacteraceae-dominated environments. Quantitative reverse transcription-PCR studies with mRNA extracted from cultures of Geobacter sulfurreducens grown in chemostats with Fe(III) as the electron acceptor or in batch with electrodes as the electron acceptor indicated that transcript levels of the citrate synthase gene, gltA, increased with increased rates of growth/Fe(III) reduction or current production, whereas the expression of the constitutively expressed housekeeping genes recA, rpoD, and proC remained relatively constant. Analysis of mRNA extracted from groundwater collected from a U(VI)-contaminated site undergoing in situ uranium bioremediation revealed a remarkable correspondence between acetate levels in the groundwater and levels of transcripts of gltA. The expression of gltA was also significantly greater in RNA extracted from groundwater beneath a highway runoff recharge pool that was exposed to calcium magnesium acetate in June, when acetate concentrations were high, than in October, when the levels had significantly decreased. It was also possible to detect gltA transcripts on current-harvesting anodes deployed in freshwater sediments. These results suggest that it is possible to monitor the in situ metabolic rate of Geobacteraceae by tracking the expression of the citrate synthase gene. PMID:16269721

  16. Quantifying facial expression recognition across viewing conditions.

    PubMed

    Goren, Deborah; Wilson, Hugh R

    2006-04-01

    Facial expressions are key to social interactions and to assessment of potential danger in various situations. Therefore, our brains must be able to recognize facial expressions when they are transformed in biologically plausible ways. We used synthetic happy, sad, angry and fearful faces to determine the amount of geometric change required to recognize these emotions during brief presentations. Five-alternative forced choice conditions involving central viewing, peripheral viewing and inversion were used to study recognition among the four emotions. Two-alternative forced choice was used to study affect discrimination when spatial frequency information in the stimulus was modified. The results show an emotion and task-dependent pattern of detection. Facial expressions presented with low peak frequencies are much harder to discriminate from neutral than faces defined by either mid or high peak frequencies. Peripheral presentation of faces also makes recognition much more difficult, except for happy faces. Differences between fearful detection and recognition tasks are probably due to common confusions with sadness when recognizing fear from among other emotions. These findings further support the idea that these emotions are processed separately from each other. PMID:16364393

  17. Epigenetics and gene expression.

    PubMed

    Gibney, E R; Nolan, C M

    2010-07-01

    Transcription, translation and subsequent protein modification represent the transfer of genetic information from the archival copy of DNA to the short-lived messenger RNA, usually with subsequent production of protein. Although all cells in an organism contain essentially the same DNA, cell types and functions differ because of qualitative and quantitative differences in their gene expression. Thus, control of gene expression is at the heart of differentiation and development. Epigenetic processes, including DNA methylation, histone modification and various RNA-mediated processes, are thought to influence gene expression chiefly at the level of transcription; however, other steps in the process (for example, translation) may also be regulated epigenetically. The following paper will outline the role epigenetics is believed to have in influencing gene expression.

  18. A gene expression screen.

    PubMed Central

    Wang, Z; Brown, D D

    1991-01-01

    A gene expression screen identifies mRNAs that differ in abundance between two mRNA mixtures by a subtractive hybridization method. The two mRNA populations are converted to double-stranded cDNAs, fragmented, and ligated to linkers for polymerase chain reaction (PCR) amplification. The multiple cDNA fragments isolated from any given gene can be treated as alleles in a genetic screen. Probability analysis of the frequency with which multiple alleles are found provides an estimation of the total number of up- and down-regulated genes. We have applied this method to genes that are differentially expressed in amphibian tadpole tail tissue in the first 24 hr after thyroid hormone treatment, which ultimately induces tail resorption. We estimate that there are about 30 up-regulated genes; 16 have been isolated. Images PMID:1722336

  19. Gene expression technology

    SciTech Connect

    Goeddel, D.V. )

    1990-01-01

    The articles in this volume were assemble to enable the reader to design effective strategies for the expression of cloned genes and cDNAs. More than a compilation of papers describing the multitude of techniques now available for expressing cloned genes, this volume provides a manual that should prove useful for solving the majority of expression problems one likely to encounter. The four major expression systems commonly available to most investigators are stressed: Escherichia coli, Bacillus subtilis, yeast, and mammalian cells. Each of these system has its advantages and disadvantages, details of which are found in Chapter 1 and the strategic overviews for the four major sections of the volume. The papers in each of these sections provide many suggestions on how to proceed if initial expression levels are not sufficient.

  20. Modifiers, Quantifiers, and the Treatment of Japanese Numeral Expressions.

    ERIC Educational Resources Information Center

    Dubinsky, Stanley

    Analysis of Japanese numeral expressions provides evidence that their two semantically distinct functions, attributive and partitive, can be associated with distinct syntactic positions within the noun phrase (NP). The attributive numeral phrase occupies the position of a modifier and is a complement to N or N'. The quantifier (partitive) numeral…

  1. Gene structure and expression

    SciTech Connect

    Hawkins, J. )

    1990-01-01

    This book describes the structure of genes in molecular terms and summarizes present knowledge about how their activity is regulated. It covers a range of topics, including a review of the structure and replication of DNA, transcription and translation, prokaryotic and eukaryotic gene organization and expression, retroviruses and oncogenes. The book also includes a chapter on the methodology of DNA manipulation including sections on site-directed mutagenesis, the polymerase chain reaction, reporter genes and restriction fragment length polymorphisms. The hemoglobin gene system and the genetics of the proteins of the immune system are presented in the latter half of the book to show the structure and expression of the most well-studied systems in higher eukaryotes. The final chapter reviews the differences between prokaryotic and the eukaryotic genomes.

  2. A novel method to quantify gene set functional association based on gene ontology.

    PubMed

    Lv, Sali; Li, Yan; Wang, Qianghu; Ning, Shangwei; Huang, Teng; Wang, Peng; Sun, Jie; Zheng, Yan; Liu, Weisha; Ai, Jing; Li, Xia

    2012-05-01

    Numerous gene sets have been used as molecular signatures for exploring the genetic basis of complex disorders. These gene sets are distinct but related to each other in many cases; therefore, efforts have been made to compare gene sets for studies such as those evaluating the reproducibility of different experiments. Comparison in terms of biological function has been demonstrated to be helpful to biologists. We improved the measurement of semantic similarity to quantify the functional association between gene sets in the context of gene ontology and developed a web toolkit named Gene Set Functional Similarity (GSFS; http://bioinfo.hrbmu.edu.cn/GSFS). Validation based on protein complexes for which the functional associations are known demonstrated that the GSFS scores tend to be correlated with sequence similarity scores and that complexes with high GSFS scores tend to be involved in the same functional catalogue. Compared with the pairwise method and the annotation method, the GSFS shows better discrimination and more accurately reflects the known functional catalogues shared between complexes. Case studies comparing differentially expressed genes of prostate tumour samples from different microarray platforms and identifying coronary heart disease susceptibility pathways revealed that the method could contribute to future studies exploring the molecular basis of complex disorders.

  3. Gene Express Inc.

    PubMed

    Saccomanno, Colette F

    2006-07-01

    Gene Express, Inc. is a technology-licensing company and provider of Standardized Reverse Transcription Polymerase Chain Reaction (StaRT-PCR) services. Designed by and for clinical researchers involved in pharmaceutical, biomarker and molecular diagnostic product development, StaRT-PCR is a unique quantitative and standardized multigene expression measurement platform. StaRT-PCR meets all of the performance characteristics defined by the US FDA as required to support regulatory submissions [101,102] , and by the Clinical Laboratory Improvement Act of 1988 (CLIA) as necessary to support diagnostic testing [1] . A standardized mixture of internal standards (SMIS), manufactured in bulk, provides integrated quality control wherein each native template target gene is measured relative to a competitive template internal standard. Bulk production enables the compilation of a comprehensive standardized database from across multiple experiments, across collaborating laboratories and across the entire clinical development lifecycle of a given compound or diagnostic product. For the first time, all these data are able to be directly compared. Access to such a database can dramatically shorten the time from investigational new drug (IND) to new drug application (NDA), or save time and money by hastening a substantiated 'no-go' decision. High-throughput StaRT-PCR is conducted at the company's automated Standardized Expression Measurement (SEM) Center. Currently optimized for detection on a microcapillary electrophoretic platform, StaRT-PCR products also may be analyzed on microarray, high-performance liquid chromatography (HPLC), or matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) platforms. SEM Center services deliver standardized genomic data--data that will accelerate the application of pharmacogenomic technology to new drug and diagnostic test development and facilitate personalized medicine.

  4. Modeling gene expression in time and space.

    PubMed

    Rué, Pau; Garcia-Ojalvo, Jordi

    2013-01-01

    Cell populations rarely exhibit gene-expression profiles that are homogeneous in time and space. In the temporal domain, dynamical behaviors such as oscillations and pulses of protein production pervade cell biology, underlying phenomena as diverse as circadian rhythmicity, cell cycle control, stress and damage responses, and stem-cell pluripotency. In multicellular populations, spatial heterogeneities are crucial for decision making and development, among many other functions. Cells need to exquisitely coordinate this temporal and spatial variation to survive. Although the spatiotemporal character of gene expression is challenging to quantify experimentally at the level of individual cells, it is beneficial from the modeling viewpoint, because it provides strong constraints that can be probed by theoretically analyzing mathematical models of candidate gene and protein circuits. Here, we review recent examples of temporal dynamics and spatial patterning in gene expression to show how modeling such phenomenology can help us unravel the molecular mechanisms of cellular function.

  5. 17A-ETHYNYLESTRADIOL-INDUCED VITELLOGENIN GENE TRANSCRIPTION QUANTIFIED IN LIVERS OF ADULT MALES, LARVAE, AND GILLS OF FATHEAD MINNOW (PIMEPHALES PROMELAS)

    EPA Science Inventory

    We have applied a method for quantifying relative levels of messenger RNA (mRNA) transcription to assess chemically-induced gene expression in fathead minnows (Pimephales promelas). Synthetic oligonucleotides designed for the fathead minnow vitellogenin gene transcription (Vg) p...

  6. Nonadditive gene expression in polyploids.

    PubMed

    Yoo, Mi-Jeong; Liu, Xiaoxian; Pires, J Chris; Soltis, Pamela S; Soltis, Douglas E

    2014-01-01

    Allopolyploidy involves hybridization and duplication of divergent parental genomes and provides new avenues for gene expression. The expression levels of duplicated genes in polyploids can show deviation from parental additivity (the arithmetic average of the parental expression levels). Nonadditive expression has been widely observed in diverse polyploids and comprises at least three possible scenarios: (a) The total gene expression level in a polyploid is similar to that of one of its parents (expression-level dominance); (b) total gene expression is lower or higher than in both parents (transgressive expression); and (c) the relative contribution of the parental copies (homeologs) to the total gene expression is unequal (homeolog expression bias). Several factors may result in expression nonadditivity in polyploids, including maternal-paternal influence, gene dosage balance, cis- and/or trans-regulatory networks, and epigenetic regulation. As our understanding of nonadditive gene expression in polyploids remains limited, a new generation of investigators should explore additional phenomena (i.e., alternative splicing) and use other high-throughput "omics" technologies to measure the impact of nonadditive expression on phenotype, proteome, and metabolome. PMID:25421600

  7. Evolution of Gene Expression after Gene Amplification

    PubMed Central

    Garcia, Nelson; Zhang, Wei; Wu, Yongrui; Messing, Joachim

    2015-01-01

    We took a rather unique approach to investigate the conservation of gene expression of prolamin storage protein genes across two different subfamilies of the Poaceae. We took advantage of oat plants carrying single maize chromosomes in different cultivars, called oat–maize addition (OMA) lines, which permitted us to determine whether regulation of gene expression was conserved between the two species. We found that γ-zeins are expressed in OMA7.06, which carries maize chromosome 7 even in the absence of the trans-acting maize prolamin-box-binding factor (PBF), which regulates their expression. This is likely because oat PBF can substitute for the function of maize PBF as shown in our transient expression data, using a γ-zein promoter fused to green fluorescent protein (GFP). Despite this conservation, the younger, recently amplified prolamin genes in maize, absent in oat, are not expressed in the corresponding OMAs. However, maize can express the oldest prolamin gene, the wheat high-molecular weight glutenin Dx5 gene, even when maize Pbf is knocked down (through PbfRNAi), and/or another maize transcription factor, Opaque-2 (O2) is knocked out (in maize o2 mutant). Therefore, older genes are conserved in their regulation, whereas younger ones diverged during evolution and eventually acquired a new repertoire of suitable transcriptional activators. PMID:25912045

  8. Serial analysis of gene expression.

    PubMed

    Velculescu, V E; Zhang, L; Vogelstein, B; Kinzler, K W

    1995-10-20

    The characteristics of an organism are determined by the genes expressed within it. A method was developed, called serial analysis of gene expression (SAGE), that allows the quantitative and simultaneous analysis of a large number of transcripts. To demonstrate this strategy, short diagnostic sequence tags were isolated from pancreas, concatenated, and cloned. Manual sequencing of 1000 tags revealed a gene expression pattern characteristic of pancreatic function. New pancreatic transcripts corresponding to novel tags were identified. SAGE should provide a broadly applicable means for the quantitative cataloging and comparison of expressed genes in a variety of normal, developmental, and disease states. PMID:7570003

  9. Expression and mapping of anthocyanin biosynthesis genes in carrot

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Anthocyanin gene expression has been extensively studied in leaves, fruits and flowers of numerous plants. Little, however, is known about anthocyanin accumulation in roots, or in carrots or other Apiaceae. We quantified expression of six anthocyanin biosynthetic genes (phenylalanine ammonia-lyase (...

  10. Optimal Reference Genes for Gene Expression Normalization in Trichomonas vaginalis.

    PubMed

    dos Santos, Odelta; de Vargas Rigo, Graziela; Frasson, Amanda Piccoli; Macedo, Alexandre José; Tasca, Tiana

    2015-01-01

    Trichomonas vaginalis is the etiologic agent of trichomonosis, the most common non-viral sexually transmitted disease worldwide. This infection is associated with several health consequences, including cervical and prostate cancers and HIV acquisition. Gene expression analysis has been facilitated because of available genome sequences and large-scale transcriptomes in T. vaginalis, particularly using quantitative real-time polymerase chain reaction (qRT-PCR), one of the most used methods for molecular studies. Reference genes for normalization are crucial to ensure the accuracy of this method. However, to the best of our knowledge, a systematic validation of reference genes has not been performed for T. vaginalis. In this study, the transcripts of nine candidate reference genes were quantified using qRT-PCR under different cultivation conditions, and the stability of these genes was compared using the geNorm and NormFinder algorithms. The most stable reference genes were α-tubulin, actin and DNATopII, and, conversely, the widely used T. vaginalis reference genes GAPDH and β-tubulin were less stable. The PFOR gene was used to validate the reliability of the use of these candidate reference genes. As expected, the PFOR gene was upregulated when the trophozoites were cultivated with ferrous ammonium sulfate when the DNATopII, α-tubulin and actin genes were used as normalizing gene. By contrast, the PFOR gene was downregulated when the GAPDH gene was used as an internal control, leading to misinterpretation of the data. These results provide an important starting point for reference gene selection and gene expression analysis with qRT-PCR studies of T. vaginalis.

  11. Optimal Reference Genes for Gene Expression Normalization in Trichomonas vaginalis.

    PubMed

    dos Santos, Odelta; de Vargas Rigo, Graziela; Frasson, Amanda Piccoli; Macedo, Alexandre José; Tasca, Tiana

    2015-01-01

    Trichomonas vaginalis is the etiologic agent of trichomonosis, the most common non-viral sexually transmitted disease worldwide. This infection is associated with several health consequences, including cervical and prostate cancers and HIV acquisition. Gene expression analysis has been facilitated because of available genome sequences and large-scale transcriptomes in T. vaginalis, particularly using quantitative real-time polymerase chain reaction (qRT-PCR), one of the most used methods for molecular studies. Reference genes for normalization are crucial to ensure the accuracy of this method. However, to the best of our knowledge, a systematic validation of reference genes has not been performed for T. vaginalis. In this study, the transcripts of nine candidate reference genes were quantified using qRT-PCR under different cultivation conditions, and the stability of these genes was compared using the geNorm and NormFinder algorithms. The most stable reference genes were α-tubulin, actin and DNATopII, and, conversely, the widely used T. vaginalis reference genes GAPDH and β-tubulin were less stable. The PFOR gene was used to validate the reliability of the use of these candidate reference genes. As expected, the PFOR gene was upregulated when the trophozoites were cultivated with ferrous ammonium sulfate when the DNATopII, α-tubulin and actin genes were used as normalizing gene. By contrast, the PFOR gene was downregulated when the GAPDH gene was used as an internal control, leading to misinterpretation of the data. These results provide an important starting point for reference gene selection and gene expression analysis with qRT-PCR studies of T. vaginalis. PMID:26393928

  12. Optimal Reference Genes for Gene Expression Normalization in Trichomonas vaginalis

    PubMed Central

    dos Santos, Odelta; de Vargas Rigo, Graziela; Frasson, Amanda Piccoli; Macedo, Alexandre José; Tasca, Tiana

    2015-01-01

    Trichomonas vaginalis is the etiologic agent of trichomonosis, the most common non-viral sexually transmitted disease worldwide. This infection is associated with several health consequences, including cervical and prostate cancers and HIV acquisition. Gene expression analysis has been facilitated because of available genome sequences and large-scale transcriptomes in T. vaginalis, particularly using quantitative real-time polymerase chain reaction (qRT-PCR), one of the most used methods for molecular studies. Reference genes for normalization are crucial to ensure the accuracy of this method. However, to the best of our knowledge, a systematic validation of reference genes has not been performed for T. vaginalis. In this study, the transcripts of nine candidate reference genes were quantified using qRT-PCR under different cultivation conditions, and the stability of these genes was compared using the geNorm and NormFinder algorithms. The most stable reference genes were α-tubulin, actin and DNATopII, and, conversely, the widely used T. vaginalis reference genes GAPDH and β-tubulin were less stable. The PFOR gene was used to validate the reliability of the use of these candidate reference genes. As expected, the PFOR gene was upregulated when the trophozoites were cultivated with ferrous ammonium sulfate when the DNATopII, α-tubulin and actin genes were used as normalizing gene. By contrast, the PFOR gene was downregulated when the GAPDH gene was used as an internal control, leading to misinterpretation of the data. These results provide an important starting point for reference gene selection and gene expression analysis with qRT-PCR studies of T. vaginalis. PMID:26393928

  13. Introduction to the Gene Expression Analysis.

    PubMed

    Segundo-Val, Ignacio San; Sanz-Lozano, Catalina S

    2016-01-01

    In 1941, Beadle and Tatum published experiments that would explain the basis of the central dogma of molecular biology, whereby the DNA through an intermediate molecule, called RNA, results proteins that perform the functions in cells. Currently, biomedical research attempts to explain the mechanisms by which develops a particular disease, for this reason, gene expression studies have proven to be a great resource. Strictly, the term "gene expression" comprises from the gene activation until the mature protein is located in its corresponding compartment to perform its function and contribute to the expression of the phenotype of cell.The expression studies are directed to detect and quantify messenger RNA (mRNA) levels of a specific gene. The development of the RNA-based gene expression studies began with the Northern Blot by Alwine et al. in 1977. In 1969, Gall and Pardue and John et al. independently developed the in situ hybridization, but this technique was not employed to detect mRNA until 1986 by Coghlan. Today, many of the techniques for quantification of RNA are deprecated because other new techniques provide more information. Currently the most widely used techniques are qPCR, expression microarrays, and RNAseq for the transcriptome analysis. In this chapter, these techniques will be reviewed. PMID:27300529

  14. Method of controlling gene expression

    DOEpatents

    Peters, Norman K.; Frost, John W.; Long, Sharon R.

    1991-12-03

    A method of controlling expression of a DNA segment under the control of a nod gene promoter which comprises administering to a host containing a nod gene promoter an amount sufficient to control expression of the DNA segment of a compound of the formula: ##STR1## in which each R is independently H or OH, is described.

  15. Gene expression and fractionation resistance

    PubMed Central

    2014-01-01

    Background Previous work on whole genome doubling in plants established the importance of gene functional category in provoking or suppressing duplicate gene loss, or fractionation. Other studies, particularly in Paramecium have correlated levels of gene expression with vulnerability or resistance to duplicate loss. Results Here we analyze the simultaneous effect of function category and expression in two plant data sets, rosids and asterids. Conclusion We demonstrate function category and expression level have independent effects, though expression does not play the dominant role it does in Paramecium. PMID:25573431

  16. Analysis of neuronal gene expression with laser capture microdissection.

    PubMed

    Vincent, Valerie A M; DeVoss, Jason J; Ryan, Heather S; Murphy, Greer M

    2002-09-01

    The brain is a heterogeneous tissue in which the numbers of neurons, glia, and other cell types vary among anatomic regions. Gene expression studies performed on brain homogenates yield results reflecting mRNA abundance in a mixture of cell types. Therefore, a method for quantifying gene expression in individual cell populations would be useful. Laser capture microdissection (LCM) is a new technique for obtaining pure populations of cells from heterogeneous tissues. Most studies thus far have used LCM to detect DNA sequences. We developed a method to quantify gene expression in hippocampal neurons from mouse brain using LCM and real-time reverse transcriptase-polymerase chain reaction (RT-PCR). This method was optimized to permit histochemical or immunocytochemical visualization of nerve cells during LCM while minimizing RNA degradation. As an example, gene expression was quantified in hippocampal neurons from the Tg2576 mouse model for Alzheimer's disease.

  17. Validation of reference genes for gene expression studies in soybean aphid, Aphis glycines Matsumura

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Quantitative real-time PCR (qRT-PCR) is a common tool for quantifying mRNA transcripts. To normalize results, a reference gene is mandatory. Aphis glycines is a significant soybean pest, yet gene expression and functional genomics studies are hindered by a lack of stable reference genes. We evalu...

  18. Regulation of Neuronal Gene Expression

    NASA Astrophysics Data System (ADS)

    Thiel, Gerald; Lietz, Michael; Leichter, Michael

    Humans as multicellular organisms contain a variety of different cell types where each cell population must fulfill a distinct function in the interest of the whole organism. The molecular basis for the variations in morphology, biochemistry, molecular biology, and function of the various cell types is the cell-type specific expression of genes. These genes encode proteins necessary for executing the specialized functions of each cell type within an organism. We describe here a regulatory mechanism for the expression of neuronal genes. The zinc finger protein REST binds to the regulatory region of many neuronal genes and represses neuronal gene expression in nonneuronal tissues. A negative regulatory mechanism, involving a transcriptional repressor, seems to play an important role in establishing the neuronal phenotype.

  19. Regulation of methane genes and genome expression

    SciTech Connect

    John N. Reeve

    2009-09-09

    At the start of this project, it was known that methanogens were Archaeabacteria (now Archaea) and were therefore predicted to have gene expression and regulatory systems different from Bacteria, but few of the molecular biology details were established. The goals were then to establish the structures and organizations of genes in methanogens, and to develop the genetic technologies needed to investigate and dissect methanogen gene expression and regulation in vivo. By cloning and sequencing, we established the gene and operon structures of all of the “methane” genes that encode the enzymes that catalyze methane biosynthesis from carbon dioxide and hydrogen. This work identified unique sequences in the methane gene that we designated mcrA, that encodes the largest subunit of methyl-coenzyme M reductase, that could be used to identify methanogen DNA and establish methanogen phylogenetic relationships. McrA sequences are now the accepted standard and used extensively as hybridization probes to identify and quantify methanogens in environmental research. With the methane genes in hand, we used northern blot and then later whole-genome microarray hybridization analyses to establish how growth phase and substrate availability regulated methane gene expression in Methanobacterium thermautotrophicus ΔH (now Methanothermobacter thermautotrophicus). Isoenzymes or pairs of functionally equivalent enzymes catalyze several steps in the hydrogen-dependent reduction of carbon dioxide to methane. We established that hydrogen availability determine which of these pairs of methane genes is expressed and therefore which of the alternative enzymes is employed to catalyze methane biosynthesis under different environmental conditions. As were unable to establish a reliable genetic system for M. thermautotrophicus, we developed in vitro transcription as an alternative system to investigate methanogen gene expression and regulation. This led to the discovery that an archaeal protein

  20. Stochastic models of gene expression and post-transcriptional regulation

    NASA Astrophysics Data System (ADS)

    Pendar, Hodjat; Kulkarni, Rahul; Jia, Tao

    2011-10-01

    The intrinsic stochasticity of gene expression can give rise to phenotypic heterogeneity in a population of genetically identical cells. Correspondingly, there is considerable interest in understanding how different molecular mechanisms impact the 'noise' in gene expression. Of particular interest are post-transcriptional regulatory mechanisms involving genes called small RNAs, which control important processes such as development and cancer. We propose and analyze general stochastic models of gene expression and derive exact analytical expressions quantifying the noise in protein distributions [1]. Focusing on specific regulatory mechanisms, we analyze a general model for post-transcriptional regulation of stochastic gene expression [2]. The results obtained provide new insights into the role of post-transcriptional regulation in controlling the noise in gene expression. [4pt] [1] T. Jia and R. V. Kulkarni, Phys. Rev. Lett.,106, 058102 (2011) [0pt] [2] T. Jia and R. V. Kulkarni, Phys. Rev. Lett., 105, 018101 (2010)

  1. Nutritional regulation of gene expression.

    PubMed

    Cousins, R J

    1999-01-25

    Genes are regulated by complex arrays of response elements that influence the rate of transcription. Nutrients and hormones either act directly to influence these rates or act indirectly through specialized signaling pathways. Metabolites of vitamins A and D, fatty acids, some sterols, and zinc are among the nutrients that influence transcription directly. Components of dietary fiber may influence gene expression indirectly through changes in hormonal signaling, mechanical stimuli, and metabolites produced by the intestinal microflora. In addition, consumption of water-soluble fibers may lead to changes in gene expression mediated through indirect mechanisms that influence transcription rates. In the large intestine, short-chain fatty acids, including butyric acid, are produced by microflora. Butyric acid can indirectly influence gene expression. Some sources of fiber limit nutrient absorption, particularly of trace elements. This could have direct or indirect effects on gene expression. Identification of genes in colonic epithelial cells that are differentially regulated by dietary fiber will be an important step toward understanding the role of dietary factors in colorectal cancer progression.

  2. GLAST: gene expression regulation by phorbol esters.

    PubMed

    Espinoza-Rojo, M; López-Bayghen, E; Ortega, A

    2000-08-21

    The gene expression regulation of the Na+-dependent high affinity glutamate/aspartate transporter GLAST expressed in cultured Bergmann glia cells from chick cerebellum was studied. A 679 bp fragment of the chick GLAST cDNA was cloned and sequenced. Specific PCR primers were used to quantify chick GLAST mRNA levels. Treatment of the cells with the Ca2+/diacylglycerol dependent protein kinase C (PKC) activator, phorbol 12-tetradecanoyl-13-acetate (TPA) produced a decrease in transporter mRNA levels, without an effect in its mRNA half life, suggesting a transcriptional down regulation. Activation of the cAMP pathway results in a transient decrease in GLAST mRNA levels, in contrast with the TPA effect. These findings suggest that GLAST expression is under control of distinct signaling pathways.

  3. Transgenic Arabidopsis Gene Expression System

    NASA Technical Reports Server (NTRS)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  4. Zipf's Law in Gene Expression

    NASA Astrophysics Data System (ADS)

    Furusawa, Chikara; Kaneko, Kunihiko

    2003-02-01

    Using data from gene expression databases on various organisms and tissues, including yeast, nematodes, human normal and cancer tissues, and embryonic stem cells, we found that the abundances of expressed genes exhibit a power-law distribution with an exponent close to -1; i.e., they obey Zipf’s law. Furthermore, by simulations of a simple model with an intracellular reaction network, we found that Zipf’s law of chemical abundance is a universal feature of cells where such a network optimizes the efficiency and faithfulness of self-reproduction. These findings provide novel insights into the nature of the organization of reaction dynamics in living cells.

  5. Neighboring Genes Show Correlated Evolution in Gene Expression.

    PubMed

    Ghanbarian, Avazeh T; Hurst, Laurence D

    2015-07-01

    When considering the evolution of a gene's expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (<100 kb) but extends much further. Sex-specific expression change is also genomically clustered. As genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking.

  6. Neighboring Genes Show Correlated Evolution in Gene Expression

    PubMed Central

    Ghanbarian, Avazeh T.; Hurst, Laurence D.

    2015-01-01

    When considering the evolution of a gene’s expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (<100 kb) but extends much further. Sex-specific expression change is also genomically clustered. As genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking. PMID:25743543

  7. Real-Time PCR for Gene Expression Quantification in Asthma.

    PubMed

    Segundo-Val, Ignacio San; García-Solaesa, Virginia; García-Sánchez, Asunción

    2016-01-01

    The quantitative real-time PCR (qPCR) has become the reference technique for studying gene expression in recent years. The application of qPCR to the study of asthma provides very useful information regarding the gene expression mechanisms. The quantification of RNA from cDNA can be performed by using fluorescent dyes or specific sequence probes. Here, we describe the protocol to quantify gene expression levels using SYBR Green as fluorescent dye. The protocol starts with the RNA extraction, followed by reverse transcription to obtain cDNA, quantification and finally data analysis.

  8. Real-Time PCR for Gene Expression Quantification in Asthma.

    PubMed

    Segundo-Val, Ignacio San; García-Solaesa, Virginia; García-Sánchez, Asunción

    2016-01-01

    The quantitative real-time PCR (qPCR) has become the reference technique for studying gene expression in recent years. The application of qPCR to the study of asthma provides very useful information regarding the gene expression mechanisms. The quantification of RNA from cDNA can be performed by using fluorescent dyes or specific sequence probes. Here, we describe the protocol to quantify gene expression levels using SYBR Green as fluorescent dye. The protocol starts with the RNA extraction, followed by reverse transcription to obtain cDNA, quantification and finally data analysis. PMID:27300530

  9. General method for quantifying base adducts in specific mammalian genes.

    PubMed Central

    Thomas, D C; Morton, A G; Bohr, V A; Sancar, A

    1988-01-01

    A general method has been developed to measure the formation and removal of DNA adducts in defined sequences of mammalian genomes. Adducted genomic DNA is digested with an appropriate restriction enzyme, treated with Escherichia coli UvrABC excision nuclease (ABC excinuclease), subjected to alkaline gel electrophoresis, and probed for specific sequences by Southern hybridization. The ABC excinuclease incises DNA containing bulky adducts and thus reduces the intensity of the full-length fragments in Southern hybridization in proportion to the number of adducts present in the probed sequence. This method is similar to that developed by Bohr et al. [Bohr, V. A., Smith, C. A., Okumoto, D. S. & Hanawalt, P. C. (1985) Cell 40, 359-369] for quantifying pyrimidine dimers by using T4 endonuclease V. Because of the wide substrate range of ABC exinuclease, however, our method can be used to quantify a large variety of DNA adducts in specific genomic sequences. Images PMID:2836856

  10. Modulation of R-gene expression across environments

    PubMed Central

    MacQueen, Alice; Bergelson, Joy

    2016-01-01

    Some environments are more conducive to pathogen growth than others, and, as a consequence, plants might be expected to invest more in resistance when pathogen growth is favored. Resistance (R-) genes in Arabidopsis thaliana have unusually extensive variation in basal expression when comparing the same R-gene among accessions collected from different environments. R-gene expression variation was characterized to explore whether R-gene expression is up-regulated in environments favoring pathogen proliferation and down-regulated when risks of infection are low; down-regulation would follow if costs of R-gene expression negatively impact plant fitness in the absence of disease. Quantitative reverse transcription–PCR was used to quantify the expression of 13 R-gene loci in plants grown in eight environmental conditions for each of 12 A. thaliana accessions, and large effects of the environment on R-gene expression were found. Surprisingly, almost every change in the environment—be it a change in biotic or abiotic conditions—led to an increase in R-gene expression, a response that was distinct from the average transcriptome response and from that of other stress response genes. These changes in expression are functional in that environmental change prior to infection affected levels of specific disease resistance to isolates of Pseudomonas syringae. In addition, there are strong latitudinal clines in basal R-gene expression and clines in R-gene expression plasticity correlated with drought and high temperatures. These results suggest that variation in R-gene expression across environments may be shaped by natural selection to reduce fitness costs of R-gene expression in permissive or predictable environments. PMID:26983577

  11. Modulation of R-gene expression across environments.

    PubMed

    MacQueen, Alice; Bergelson, Joy

    2016-03-01

    Some environments are more conducive to pathogen growth than others, and, as a consequence, plants might be expected to invest more in resistance when pathogen growth is favored. Resistance (R-) genes in Arabidopsis thaliana have unusually extensive variation in basal expression when comparing the same R-gene among accessions collected from different environments. R-gene expression variation was characterized to explore whether R-gene expression is up-regulated in environments favoring pathogen proliferation and down-regulated when risks of infection are low; down-regulation would follow if costs of R-gene expression negatively impact plant fitness in the absence of disease. Quantitative reverse transcription-PCR was used to quantify the expression of 13 R-gene loci in plants grown in eight environmental conditions for each of 12 A. thaliana accessions, and large effects of the environment on R-gene expression were found. Surprisingly, almost every change in the environment--be it a change in biotic or abiotic conditions--led to an increase in R-gene expression, a response that was distinct from the average transcriptome response and from that of other stress response genes. These changes in expression are functional in that environmental change prior to infection affected levels of specific disease resistance to isolates of Pseudomonas syringae. In addition, there are strong latitudinal clines in basal R-gene expression and clines in R-gene expression plasticity correlated with drought and high temperatures. These results suggest that variation in R-gene expression across environments may be shaped by natural selection to reduce fitness costs of R-gene expression in permissive or predictable environments. PMID:26983577

  12. Vascular gene expression: a hypothesis

    PubMed Central

    Martínez-Navarro, Angélica C.; Galván-Gordillo, Santiago V.; Xoconostle-Cázares, Beatriz; Ruiz-Medrano, Roberto

    2013-01-01

    The phloem is the conduit through which photoassimilates are distributed from autotrophic to heterotrophic tissues and is involved in the distribution of signaling molecules that coordinate plant growth and responses to the environment. Phloem function depends on the coordinate expression of a large array of genes. We have previously identified conserved motifs in upstream regions of the Arabidopsis genes, encoding the homologs of pumpkin phloem sap mRNAs, displaying expression in vascular tissues. This tissue-specific expression in Arabidopsis is predicted by the overrepresentation of GA/CT-rich motifs in gene promoters. In this work we have searched for common motifs in upstream regions of the homologous genes from plants considered to possess a “primitive” vascular tissue (a lycophyte), as well as from others that lack a true vascular tissue (a bryophyte), and finally from chlorophytes. Both lycophyte and bryophyte display motifs similar to those found in Arabidopsis with a significantly low E-value, while the chlorophytes showed either a different conserved motif or no conserved motif at all. These results suggest that these same genes are expressed coordinately in non-vascular plants; this coordinate expression may have been one of the prerequisites for the development of conducting tissues in plants. We have also analyzed the phylogeny of conserved proteins that may be involved in phloem function and development. The presence of CmPP16, APL, FT, and YDA in chlorophytes suggests the recruitment of ancient regulatory networks for the development of the vascular tissue during evolution while OPS is a novel protein specific to vascular plants. PMID:23882276

  13. Gene expression profile of pulpitis

    PubMed Central

    Galicia, Johnah C.; Henson, Brett R.; Parker, Joel S.; Khan, Asma A.

    2016-01-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the Significance Analysis of Microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (≥30mm on VAS) compared to those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology. PMID:27052691

  14. Gene expression profile of pulpitis.

    PubMed

    Galicia, J C; Henson, B R; Parker, J S; Khan, A A

    2016-06-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the significance analysis of microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (⩾30 mm on VAS) compared with those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology. PMID:27052691

  15. Duplicate genes increase gene expression diversity within and between species.

    PubMed

    Gu, Zhenglong; Rifkin, Scott A; White, Kevin P; Li, Wen-Hsiung

    2004-06-01

    Using microarray gene expression data from several Drosophila species and strains, we show that duplicated genes, compared with single-copy genes, significantly increase gene expression diversity during development. We show further that duplicate genes tend to cause expression divergences between Drosophila species (or strains) to evolve faster than do single-copy genes. This conclusion is also supported by data from different yeast strains.

  16. Systems Biophysics of Gene Expression

    PubMed Central

    Vilar, Jose M.G.; Saiz, Leonor

    2013-01-01

    Gene expression is a process central to any form of life. It involves multiple temporal and functional scales that extend from specific protein-DNA interactions to the coordinated regulation of multiple genes in response to intracellular and extracellular changes. This diversity in scales poses fundamental challenges to the use of traditional approaches to fully understand even the simplest gene expression systems. Recent advances in computational systems biophysics have provided promising avenues to reliably integrate the molecular detail of biophysical process into the system behavior. Here, we review recent advances in the description of gene regulation as a system of biophysical processes that extend from specific protein-DNA interactions to the combinatorial assembly of nucleoprotein complexes. There is now basic mechanistic understanding on how promoters controlled by multiple, local and distal, DNA binding sites for transcription factors can actively control transcriptional noise, cell-to-cell variability, and other properties of gene regulation, including precision and flexibility of the transcriptional responses. PMID:23790365

  17. An atlas of gene expression and gene co-regulation in the human retina

    PubMed Central

    Pinelli, Michele; Carissimo, Annamaria; Cutillo, Luisa; Lai, Ching-Hung; Mutarelli, Margherita; Moretti, Maria Nicoletta; Singh, Marwah Veer; Karali, Marianthi; Carrella, Diego; Pizzo, Mariateresa; Russo, Francesco; Ferrari, Stefano; Ponzin, Diego; Angelini, Claudia; Banfi, Sandro; di Bernardo, Diego

    2016-01-01

    The human retina is a specialized tissue involved in light stimulus transduction. Despite its unique biology, an accurate reference transcriptome is still missing. Here, we performed gene expression analysis (RNA-seq) of 50 retinal samples from non-visually impaired post-mortem donors. We identified novel transcripts with high confidence (Observed Transcriptome (ObsT)) and quantified the expression level of known transcripts (Reference Transcriptome (RefT)). The ObsT included 77 623 transcripts (23 960 genes) covering 137 Mb (35 Mb new transcribed genome). Most of the transcripts (92%) were multi-exonic: 81% with known isoforms, 16% with new isoforms and 3% belonging to new genes. The RefT included 13 792 genes across 94 521 known transcripts. Mitochondrial genes were among the most highly expressed, accounting for about 10% of the reads. Of all the protein-coding genes in Gencode, 65% are expressed in the retina. We exploited inter-individual variability in gene expression to infer a gene co-expression network and to identify genes specifically expressed in photoreceptor cells. We experimentally validated the photoreceptors localization of three genes in human retina that had not been previously reported. RNA-seq data and the gene co-expression network are available online (http://retina.tigem.it). PMID:27235414

  18. The Gene Expression Omnibus database

    PubMed Central

    Clough, Emily; Barrett, Tanya

    2016-01-01

    The Gene Expression Omnibus (GEO) database is an international public repository that archives and freely distributes high-throughput gene expression and other functional genomics data sets. Created in 2000 as a worldwide resource for gene expression studies, GEO has evolved with rapidly changing technologies and now accepts high-throughput data for many other data applications, including those that examine genome methylation, chromatin structure, and genome–protein interactions. GEO supports community-derived reporting standards that specify provision of several critical study elements including raw data, processed data, and descriptive metadata. The database not only provides access to data for tens of thousands of studies, but also offers various Web-based tools and strategies that enable users to locate data relevant to their specific interests, as well as to visualize and analyze the data. This chapter includes detailed descriptions of methods to query and download GEO data and use the analysis and visualization tools. The GEO homepage is at http://www.ncbi.nlm.nih.gov/geo/. PMID:27008011

  19. The Gene Expression Omnibus Database.

    PubMed

    Clough, Emily; Barrett, Tanya

    2016-01-01

    The Gene Expression Omnibus (GEO) database is an international public repository that archives and freely distributes high-throughput gene expression and other functional genomics data sets. Created in 2000 as a worldwide resource for gene expression studies, GEO has evolved with rapidly changing technologies and now accepts high-throughput data for many other data applications, including those that examine genome methylation, chromatin structure, and genome-protein interactions. GEO supports community-derived reporting standards that specify provision of several critical study elements including raw data, processed data, and descriptive metadata. The database not only provides access to data for tens of thousands of studies, but also offers various Web-based tools and strategies that enable users to locate data relevant to their specific interests, as well as to visualize and analyze the data. This chapter includes detailed descriptions of methods to query and download GEO data and use the analysis and visualization tools. The GEO homepage is at http://www.ncbi.nlm.nih.gov/geo/. PMID:27008011

  20. Gene Expression Studies in Mosquitoes

    PubMed Central

    Chen, Xlao-Guang; Mathur, Geetika; James, Anthony A.

    2009-01-01

    Research on gene expression in mosquitoes is motivated by both basic and applied interests. Studies of genes involved in hematophagy, reproduction, olfaction, and immune responses reveal an exquisite confluence of biological adaptations that result in these highly-successful life forms. The requirement of female mosquitoes for a bloodmeal for propagation has been exploited by a wide diversity of viral, protozoan and metazoan pathogens as part of their life cycles. Identifying genes involved in host-seeking, blood feeding and digestion, reproduction, insecticide resistance and susceptibility/refractoriness to pathogen development is expected to provide the bases for the development of novel methods to control mosquito-borne diseases. Advances in mosquito transgenesis technologies, the availability of whole genome sequence information, mass sequencing and analyses of transcriptomes and RNAi techniques will assist development of these tools as well as deepen the understanding of the underlying genetic components for biological phenomena characteristic of these insect species. PMID:19161831

  1. Relating Perturbation Magnitude to Temporal Gene Expression in Biological Systems

    SciTech Connect

    Callister, Stephen J.; Parnell, John J.; Pfrender, Michael E.; Hashsham, Syed

    2009-03-19

    A method to quantitatively relate stress to response at the level of gene expression is described using Saccharomyces cerevisiae as a model organism. Stress was defined as the magnitude of perturbation and strain was defined as the magnitude of cumulative response in terms of gene expression. Expression patterns of sixty genes previously reported to be significantly impacted by osmotic shock or belonging to the high-osmotic glycerol, glycerolipid metabolism, and glycolysis pathways were determined following perturbations of increasing sodium chloride concentrations (0, 0.5, 0.7, 1.0, 1.5, and 1.4 M). Expression of these genes was quantified temporally using reverse transcriptase real time polymerase chain reaction. The magnitude of cumulative response was obtained by calculating the total moment of area of the temporal response envelope for all the 60 genes, either together or for the set of genes related to each pathway. A non-linear relationship between stress and response was observed for the range of stress studied. This study examines a quantitative approach to quantify the strain at the level of gene expression to relate stress to strain in biological systems. The approach should be generally applicable to quantitatively evaluate the response of organisms to environmental change.

  2. Expression profile of genes associated with mastitis in dairy cattle

    PubMed Central

    2009-01-01

    In order to characterize the expression of genes associated with immune response mechanisms to mastitis, we quantified the relative expression of the IL-2, IL-4, IL-6, IL-8, IL-10, IFN-γ and TNF- α genes in milk cells of healthy cows and cows with clinical mastitis. Total RNA was extracted from milk cells of six Black and White Holstein (BW) cows and six Gyr cows, including three animals with and three without mastitis per breed. Gene expression was analyzed by real-time PCR. IL-10 gene expression was higher in the group of BW and Gyr cows with mastitis compared to animals free of infection from both breeds (p < 0.05). It was also higher in BW Holstein animals with clinical mastitis (p < 0.001), but it was not significant when Gyr cows with and without mastitis were compared (0.05 < p < 0.10). Among healthy cows, BW Holstein animals tended to present a higher expression of all genes studied, with a significant difference for the IL-2 and IFN- γ genes (p < 0.001). For animals with mastitis no significant difference in gene expression was observed between the two breeds. These findings suggest that animals with mastitis develop a preferentially cell-mediated immune response. Further studies including larger samples are necessary to better characterize the gene expression profile in cows with mastitis. PMID:21637453

  3. Identification of four soybean reference genes for gene expression normalization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gene expression analysis requires the use of reference genes stably expressed independently of specific tissues or environmental conditions. Housekeeping genes (e.g., actin, tubulin, ribosomal, polyubiquitin and elongation factor 1-alpha) are commonly used as reference genes with the assumption tha...

  4. Mitochondrial RNA granules: Compartmentalizing mitochondrial gene expression

    PubMed Central

    Jourdain, Alexis A.; Boehm, Erik; Maundrell, Kinsey

    2016-01-01

    In mitochondria, DNA replication, gene expression, and RNA degradation machineries coexist within a common nondelimited space, raising the question of how functional compartmentalization of gene expression is achieved. Here, we discuss the recently characterized “mitochondrial RNA granules,” mitochondrial subdomains with an emerging role in the regulation of gene expression. PMID:26953349

  5. Gravity-Induced Gene Expression in Plants.

    NASA Astrophysics Data System (ADS)

    Sederoff, Heike; Heber, Steffen; Howard, Brian; Myburg-Nichols, Henrietta; Hammond, Rebecca; Salinas-Mondragon, Raul; Brown, Christopher S.

    Plants sense changes in their orientation towards the vector of gravity and respond with directional growth. Several metabolites in the signal transduction cascade have been identified. However, very little is known about the interaction between these sensing and signal transduction events and even less is known about their role in the differential growth response. Gravity induced changes in transcript abundance have been identified in Arabidopsis whole seedlings and root apices (Moseyko et al. 2002; Kimbrough et al. 2004). Gravity induced transcript abundance changes can be observed within less than 1 min after stimulation (Salinas-Mondragon et al. 2005). Gene expression however requires not only transcription but also translation of the mRNA. Translation can only occur when mRNA is associated with ribosomes, even though not all mRNA associated with ribosomes is actively translated. To approximate translational capacity we quantified whole genome transcript abundances in corn stem pulvini during the first hour after gravity stimulation in total and poly-ribosomal fractions. As in Arabidopsis root apices, transcript abundances of several clusters of genes responded to gravity stimulation. The vast majority of these transcripts were also found to associate with polyribosomes in the same temporal and quantitative pattern. These genes are transcriptionally regulated by gravity stimulation, but do not exhibit translational regulation. However, a small group of genes showed increased transcriptional regulation after gravity stimulation, but no association with polysomes. These transcripts likely are translationally repressed. The mechanism of translational repression for these transcripts is unknown. Based on the hypothesis that the genes essential for gravitropic responses should be expressed in most or all species, we compared the temporal gravity induced expression pattern of all orthologs identified between maize and Arabidopsis. A small group of genes showed high

  6. Identification of housekeeping genes suitable for gene expression analysis in Jian carp (Cyprinus carpio var. jian).

    PubMed

    Tang, Yong-kai; Yu, Ju-hua; Xu, Pao; Li, Jian-lin; Li, Hong-xia; Ren, Hong-tao

    2012-10-01

    Jian carp (Cyprinus carpio var. jian) is an important economic fish species cultured in China. In this report, we performed a systematic analysis to identify an appropriate housekeeping (HK) gene for the study of gene expression in Jian carp. For this purpose, partial DNA sequences of four potential candidate genes (elongation factor 1 alpha (EF-1α), glyceraldehyde-3-phosphate (GAPDH), beta-actin (ACTB), and 18S ribosomal RNA (18S rRNA) were isolated, and their expression levels were studied using RNA extracted from nine tissues (forebrain, hypothalamus, liver, fore-intestine, hind-intestine, ovary, muscle, heart, kidney) in juvenile and adult Jian carp. Gene expression levels were quantified by quantitative real time RT-PCR (qRT-PCR), and expression stability was evaluated by comparing the coefficients of variation (CV) of the Ct values. The results showed that EF-1α was the most suitable HK gene in all tissues of juvenile and adult Jian carp. However, at distinct juvenile and adult developmental stages, there was not a single optimal gene for normalization of expression levels in all tissues. EF-1α was the most stable gene only in forebrain, hypothalamus, liver, heart, and kidney. These results provide data that can be expected to aid gene expression analysis in Jian carp research, but underline the importance of identifying the optimal HK gene for each new experimental paradigm. PMID:22789712

  7. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2012-05-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  8. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2008-06-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  9. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2013-10-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  10. Microspatial gene expression patterns in the Amazon River Plume.

    PubMed

    Satinsky, Brandon M; Crump, Byron C; Smith, Christa B; Sharma, Shalabh; Zielinski, Brian L; Doherty, Mary; Meng, Jun; Sun, Shulei; Medeiros, Patricia M; Paul, John H; Coles, Victoria J; Yager, Patricia L; Moran, Mary Ann

    2014-07-29

    We investigated expression of genes mediating elemental cycling at the microspatial scale in the ocean's largest river plume using, to our knowledge, the first fully quantitative inventory of genes and transcripts. The bacterial and archaeal communities associated with a phytoplankton bloom in Amazon River Plume waters at the outer continental shelf in June 2010 harbored ∼ 1.0 × 10(13) genes and 4.7 × 10(11) transcripts per liter that mapped to several thousand microbial genomes. Genomes from free-living cells were more abundant than those from particle-associated cells, and they generated more transcripts per liter for carbon fixation, heterotrophy, nitrogen and phosphorus uptake, and iron acquisition, although they had lower expression ratios (transcripts ⋅ gene(-1)) overall. Genomes from particle-associated cells contributed more transcripts for sulfur cycling, aromatic compound degradation, and the synthesis of biologically essential vitamins, with an overall twofold up-regulation of expression compared with free-living cells. Quantitatively, gene regulation differences were more important than genome abundance differences in explaining why microenvironment transcriptomes differed. Taxa contributing genomes to both free-living and particle-associated communities had up to 65% of their expressed genes regulated differently between the two, quantifying the extent of transcriptional plasticity in marine microbes in situ. In response to patchiness in carbon, nutrients, and light at the micrometer scale, Amazon Plume microbes regulated the expression of genes relevant to biogeochemical processes at the ecosystem scale.

  11. Influence of gene copy number on self-regulated gene expression.

    PubMed

    Jędrak, Jakub; Ochab-Marcinek, Anna

    2016-11-01

    Using an analytically solvable stochastic model, we study the properties of a simple genetic circuit consisting of multiple copies of a self-regulating gene. We analyse how the variation in gene copy number and the mutations changing the auto-regulation strength affect the steady-state distribution of protein concentration. We predict that one-reporter assay, an experimental method where the extrinsic noise level is inferred from the comparison of expression variance of a single and duplicated reporter gene, may give an incorrect estimation of the extrinsic noise contribution when applied to self-regulating genes. We also show that an imperfect duplication of an auto-activated gene, changing the regulation strength of one of the copies, may lead to a hybrid, binary+graded response of these genes to external signal. The analysis of relative changes in mean gene expression before and after duplication suggests that evolutionary accumulation of gene duplications may, at a given mean burst size, non-trivially depend on the inherent noisiness of a given gene, quantified by the inverse of the maximal mean frequency of bursts. Moreover, we find that the dependence of gene expression noise on gene copy number and auto-regulation strength may qualitatively differ, e.g. in monotonicity, depending on whether the noise is measured by Fano factor or coefficient of variation. Thus, experimentally-based hypotheses linking gene expression noise and evolutionary optimisation in the context of gene copy number variation may be ambiguous as they are dependent on the particular function chosen to quantify noise. PMID:27528448

  12. Gene-expression profile comparisons distinguish seven organs of maize

    PubMed Central

    Cho, Yangrae; Fernandes, John; Kim, Soo-Hwan; Walbot, Virginia

    2002-01-01

    Background A maize array was fabricated with 5,376 unique expressed sequence tag (EST) clones sequenced from 4-day-old roots, immature ears and adult organ cDNA libraries. To elucidate organ relationships, relative mRNA levels were quantified by hybridization with embryos, three maize vegetative organs (leaf blades, leaf sheaths and roots) from multiple developmental stages, husk leaves and two types of floral organs (immature ears and silks). Results Clustering analyses of the hybridization data suggest that maize utilizes both the PEPCK and NADP-ME C4 photosynthetic routes as genes in these pathways are co-regulated. Husk RNA has a gene-expression profile more similar to floral organs than to vegetative leaves. Only 7% of the genes were highly organ specific, showing over a fourfold difference in at least one of 12 comparisons and 37% showed a two- to fourfold difference. The majority of genes were expressed in diverse organs with little difference in transcript levels. Cross-hybridization among closely related genes within multigene families could obscure tissue specificity. As a first step in elucidating individual gene-expression patterns, we show that 45-nucleotide oligo probes produce signal intensities and signal ratios comparable to PCR probes on the same matrix. Conclusions Gene-expression profile studies with cDNA microarrays provide a new molecular tool for defining plant organs and their relationships and for discovering new biological processes in silico. cDNA microarrays are insufficient for differentiating recently duplicated genes. Gene-specific oligo probes printed along with cDNA probes can query individual gene-expression profiles and gene families simultaneously. PMID:12225584

  13. Quantifying the sequence–function relation in gene silencing by bacterial small RNAs

    PubMed Central

    Hao, Yue; Zhang, Zhongge J.; Erickson, David W.; Huang, Min; Huang, Yingwu; Li, Junbai; Hwa, Terence; Shi, Hualin

    2011-01-01

    Sequence–function relations for small RNA (sRNA)-mediated gene silencing were quantified for the sRNA RyhB and some of its mRNA targets in Escherichia coli. Numerous mutants of RyhB and its targets were generated and their in vivo functions characterized at various levels of target and RyhB expression. Although a core complementary region is required for repression by RyhB, variations in the complementary sequences of the core region gave rise to a continuum of repression strengths, correlated exponentially with the computed free energy of RyhB-target duplex formation. Moreover, sequence variations in the linker region known to interact with the RNA chaperone Hfq also gave rise to a continuum of repression strengths, correlated exponentially with the computed energy cost of keeping the linker region open. These results support the applicability of the thermodynamic model in predicting sRNA–mRNA interaction and suggest that sequences at these locations may be used to fine-tune the degree of repression. Surprisingly, a truncated RyhB without the Hfq-binding region is found to repress multiple targets of the wild-type RyhB effectively, both in the presence and absence of Hfq, even though the former is required for the activity of wild-type RyhB itself. These findings challenge the commonly accepted model concerning the function of Hfq in gene silencing—both in providing stability to the sRNAs and in catalyzing the target mRNAs to take on active conformations—and raise the intriguing question of why many endogenous sRNAs subject their functions to Hfq-dependences. PMID:21742981

  14. Profiling Gene Expression in Germinating Brassica Roots.

    PubMed

    Park, Myoung Ryoul; Wang, Yi-Hong; Hasenstein, Karl H

    2014-01-01

    Based on previously developed solid-phase gene extraction (SPGE) we examined the mRNA profile in primary roots of Brassica rapa seedlings for highly expressed genes like ACT7 (actin7), TUB (tubulin1), UBQ (ubiquitin), and low expressed GLK (glucokinase) during the first day post-germination. The assessment was based on the mRNA load of the SPGE probe of about 2.1 ng. The number of copies of the investigated genes changed spatially along the length of primary roots. The expression level of all genes differed significantly at each sample position. Among the examined genes ACT7 expression was most even along the root. UBQ was highest at the tip and root-shoot junction (RS). TUB and GLK showed a basipetal gradient. The temporal expression of UBQ was highest in the MZ 9 h after primary root emergence and higher than at any other sample position. Expressions of GLK in EZ and RS increased gradually over time. SPGE extraction is the result of oligo-dT and oligo-dA hybridization and the results illustrate that SPGE can be used for gene expression profiling at high spatial and temporal resolution. SPGE needles can be used within two weeks when stored at 4 °C. Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

  15. Antisense transcription as a tool to tune gene expression.

    PubMed

    Brophy, Jennifer A N; Voigt, Christopher A

    2016-01-14

    A surprise that has emerged from transcriptomics is the prevalence of genomic antisense transcription, which occurs counter to gene orientation. While frequent, the roles of antisense transcription in regulation are poorly understood. We built a synthetic system in Escherichia coli to study how antisense transcription can change the expression of a gene and tune the response characteristics of a regulatory circuit. We developed a new genetic part that consists of a unidirectional terminator followed by a constitutive antisense promoter and demonstrate that this part represses gene expression proportionally to the antisense promoter strength. Chip-based oligo synthesis was applied to build a large library of 5,668 terminator-promoter combinations that was used to control the expression of three repressors (PhlF, SrpR, and TarA) in a simple genetic circuit (NOT gate). Using the library, we demonstrate that antisense promoters can be used to tune the threshold of a regulatory circuit without impacting other properties of its response function. Finally, we determined the relative contributions of antisense RNA and transcriptional interference to repressing gene expression and introduce a biophysical model to capture the impact of RNA polymerase collisions on gene repression. This work quantifies the role of antisense transcription in regulatory networks and introduces a new mode to control gene expression that has been previously overlooked in genetic engineering.

  16. In vitro maturation alters gene expression in bovine oocytes.

    PubMed

    Adona, Paulo R; Leal, Cláudia L V; Biase, Fernando H; De Bem, Tiago H; Mesquita, Lígia G; Meirelles, Flávio V; Ferraz, André L; Furlan, Luiz R; Monzani, Paulo S; Guemra, Samuel

    2016-08-01

    Gene expression profiling of in vivo- and in vitro-matured bovine oocytes can identify transcripts related to the developmental potential of oocytes. Nonetheless, the effects of in vitro culturing oocytes are yet to be fully understood. We tested the effects of in vitro maturation on the transcript profile of oocytes collected from Bos taurus indicus cows. We quantified the expression of 1488 genes in in vivo- and in vitro-matured oocytes. Of these, 51 genes were up-regulated, whereas 56 were down-regulated (≥2-fold) in in vivo-matured oocytes in comparison with in vitro-matured oocytes. Quantitative real-time polymerase chain reaction (PCR) of nine genes confirmed the microarray results of differential expression between in vivo- and in vitro-matured oocytes (EZR, EPN1, PSEN2, FST, IGFBP3, RBBP4, STAT3, FDPS and IRS1). We interrogated the results for enrichment of Gene Ontology categories and overlap with protein-protein interactions. The results revealed that the genes altered by in vitro maturation are mostly related to the regulation of oocyte metabolism. Additionally, analysis of protein-protein interactions uncovered two regulatory networks affected by the in vitro culture system. We propose that the differentially expressed genes are candidates for biomarkers of oocyte competence. In vitro oocyte maturation can affect the abundance of specific transcripts and are likely to deplete the developmental competence.

  17. Gene Expression Patterns in Ovarian Carcinomas

    PubMed Central

    Schaner, Marci E.; Ross, Douglas T.; Ciaravino, Giuseppe; Sørlie, Therese; Troyanskaya, Olga; Diehn, Maximilian; Wang, Yan C.; Duran, George E.; Sikic, Thomas L.; Caldeira, Sandra; Skomedal, Hanne; Tu, I-Ping; Hernandez-Boussard, Tina; Johnson, Steven W.; O'Dwyer, Peter J.; Fero, Michael J.; Kristensen, Gunnar B.; Børresen-Dale, Anne-Lise; Hastie, Trevor; Tibshirani, Robert; van de Rijn, Matt; Teng, Nelson N.; Longacre, Teri A.; Botstein, David; Brown, Patrick O.; Sikic, Branimir I.

    2003-01-01

    We used DNA microarrays to characterize the global gene expression patterns in surface epithelial cancers of the ovary. We identified groups of genes that distinguished the clear cell subtype from other ovarian carcinomas, grade I and II from grade III serous papillary carcinomas, and ovarian from breast carcinomas. Six clear cell carcinomas were distinguished from 36 other ovarian carcinomas (predominantly serous papillary) based on their gene expression patterns. The differences may yield insights into the worse prognosis and therapeutic resistance associated with clear cell carcinomas. A comparison of the gene expression patterns in the ovarian cancers to published data of gene expression in breast cancers revealed a large number of differentially expressed genes. We identified a group of 62 genes that correctly classified all 125 breast and ovarian cancer specimens. Among the best discriminators more highly expressed in the ovarian carcinomas were PAX8 (paired box gene 8), mesothelin, and ephrin-B1 (EFNB1). Although estrogen receptor was expressed in both the ovarian and breast cancers, genes that are coregulated with the estrogen receptor in breast cancers, including GATA-3, LIV-1, and X-box binding protein 1, did not show a similar pattern of coexpression in the ovarian cancers. PMID:12960427

  18. Arabidopsis gene expression patterns during spaceflight

    NASA Astrophysics Data System (ADS)

    Paul, A.-L.; Ferl, R. J.

    The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments resulted in the differential expression of hundreds of genes. A 5 day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β -Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on two fronts. First, expression patterns visualized with the Adh/GUS transgene were used to address specifically the possibility that spaceflight induces a hypoxic stress response, and to assess whether any spaceflight response was similar to control terrestrial hypoxia-induced gene expression patterns. (Paul et al., Plant Physiol. 2001, 126:613). Second, genome-wide patterns of native gene expression were evaluated utilizing the Affymetrix ATH1 GeneChip? array of 8,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes identified with the arrays was further characterized with quantitative Real-Time RT PCR (ABI - TaqmanTM). Comparison of the patterns of expression for arrays of hybridized with RNA isolated from plants exposed to spaceflight compared to the control arrays revealed hundreds of genes that were differentially expressed in response to spaceflight, yet most genes that are hallmarks of hypoxic stress were unaffected. These results will be discussed in light of current models for plant responses to the spaceflight environment, and with regard to potential future flight opportunities.

  19. Gearbox gene expression and growth rate.

    PubMed

    Aldea, M; Garrido, T; Tormo, A

    1993-07-01

    Regulation of gene expression in prokaryotic cells usually takes place at the level of transcription initiation. Different forms of RNA polymerase recognizing specific promoters are engaged in the control of many prokaryotic regulons. This also seems to be the case for some Escherichia coli genes that are induced at low growth rates and by nutrient starvation. Their gene products are synthesized at levels inversely proportional to growth rate, and this mode of regulation has been termed gearbox gene expression. This kind of growth-rate modulation is exerted by specific transcriptional initiation signals, the gearbox promoters, and some of them depend on a putative new σ factor (RpoS). Gearbox promoters drive expression of morphogenetic and cell division genes at constant levels per cell and cycle to meet the demands of cell division and septum formation. A mechanism is proposed that could sense the growth rate of the cell to alter gene expression by the action of specific σ factors.

  20. The gene expression signatures of melanoma progression

    PubMed Central

    Haqq, Christopher; Nosrati, Mehdi; Sudilovsky, Daniel; Crothers, Julia; Khodabakhsh, Daniel; Pulliam, Brian L.; Federman, Scot; Miller, James R.; Allen, Robert E.; Singer, Mark I.; Leong, Stanley P. L.; Ljung, Britt-Marie; Sagebiel, Richard W.; Kashani-Sabet, Mohammed

    2005-01-01

    Because of the paucity of available tissue, little information has previously been available regarding the gene expression profiles of primary melanomas. To understand the molecular basis of melanoma progression, we compared the gene expression profiles of a series of nevi, primary melanomas, and melanoma metastases. We found that metastatic melanomas exhibit two dichotomous patterns of gene expression, which unexpectedly reflect gene expression differences already apparent in comparing laser-capture microdissected radial and vertical phases of a large primary melanoma. Unsupervised hierarchical clustering accurately separated nevi and primary melanomas. Multiclass significance analysis of microarrays comparing normal skin, nevi, primary melanomas, and the two types of metastatic melanoma identified 2,602 transcripts that significantly correlated with sample class. These results suggest that melanoma pathogenesis can be understood as a series of distinct molecular events. The gene expression signatures identified here provide the basis for developing new diagnostics and targeting therapies for patients with malignant melanoma. PMID:15833814

  1. QUANTIFYING ATYPICALITY IN AFFECTIVE FACIAL EXPRESSIONS OF CHILDREN WITH AUTISM SPECTRUM DISORDERS.

    PubMed

    Metallinou, Angeliki; Grossman, Ruth B; Narayanan, Shrikanth

    2013-01-01

    We focus on the analysis, quantification and visualization of atypicality in affective facial expressions of children with High Functioning Autism (HFA). We examine facial Motion Capture data from typically developing (TD) children and children with HFA, using various statistical methods, including Functional Data Analysis, in order to quantify atypical expression characteristics and uncover patterns of expression evolution in the two populations. Our results show that children with HFA display higher asynchrony of motion between facial regions, more rough facial and head motion, and a larger range of facial region motion. Overall, subjects with HFA consistently display a wider variability in the expressive facial gestures that they employ. Our analysis demonstrates the utility of computational approaches for understanding behavioral data and brings new insights into the autism domain regarding the atypicality that is often associated with facial expressions of subjects with HFA.

  2. The Mouse Gene Expression Database (GXD)

    PubMed Central

    Ringwald, Martin; Eppig, Janan T.; Begley, Dale A.; Corradi, John P.; McCright, Ingeborg J.; Hayamizu, Terry F.; Hill, David P.; Kadin, James A.; Richardson, Joel E.

    2001-01-01

    The Gene Expression Database (GXD) is a community resource of gene expression information for the laboratory mouse. By combining the different types of expression data, GXD aims to provide increasingly complete information about the expression profiles of genes in different mouse strains and mutants, thus enabling valuable insights into the molecular networks that underlie normal development and disease. GXD is integrated with the Mouse Genome Database (MGD). Extensive interconnections with sequence databases and with databases from other species, and the development and use of shared controlled vocabularies extend GXD’s utility for the analysis of gene expression information. GXD is accessible through the Mouse Genome Informatics web site at http://www.informatic s.jax.org/ or directly at http://www.informatics.jax.org/me nus/expression_menu.shtml. PMID:11125060

  3. Photosynthetic gene expression in higher plants.

    PubMed

    Berry, James O; Yerramsetty, Pradeep; Zielinski, Amy M; Mure, Christopher M

    2013-11-01

    Within the chloroplasts of higher plants and algae, photosynthesis converts light into biological energy, fueling the assimilation of atmospheric carbon dioxide into biologically useful molecules. Two major steps, photosynthetic electron transport and the Calvin-Benson cycle, require many gene products encoded from chloroplast as well as nuclear genomes. The expression of genes in both cellular compartments is highly dynamic and influenced by a diverse range of factors. Light is the primary environmental determinant of photosynthetic gene expression. Working through photoreceptors such as phytochrome, light regulates photosynthetic genes at transcriptional and posttranscriptional levels. Other processes that affect photosynthetic gene expression include photosynthetic activity, development, and biotic and abiotic stress. Anterograde (from nucleus to chloroplast) and retrograde (from chloroplast to nucleus) signaling insures the highly coordinated expression of the many photosynthetic genes between these different compartments. Anterograde signaling incorporates nuclear-encoded transcriptional and posttranscriptional regulators, such as sigma factors and RNA-binding proteins, respectively. Retrograde signaling utilizes photosynthetic processes such as photosynthetic electron transport and redox signaling to influence the expression of photosynthetic genes in the nucleus. The basic C3 photosynthetic pathway serves as the default form used by most of the plant species on earth. High temperature and water stress associated with arid environments have led to the development of specialized C4 and CAM photosynthesis, which evolved as modifications of the basic default expression program. The goal of this article is to explain and summarize the many gene expression and regulatory processes that work together to support photosynthetic function in plants.

  4. TFRC and ACTB as the best reference genes to quantify Urokinase Plasminogen Activator in breast cancer

    PubMed Central

    2011-01-01

    Background Biomedical researchers have long looked for ways to diagnose and treat cancer patients at the early stages through biomarkers. Although conventional techniques are routinely applied in the detection of biomarkers, attitudes towards using Real-Time PCR techniques in detection of many biomarkers are increasing. Normalization of quantitative Real-Time PCR is necessary to validate non-biological alteration occurring during the steps of RNA quantification. Selection of variably expressed housekeeping genes (HKs) will affect the validity of the data. The aim of the present study was to identify uniformly expressed housekeeping genes in order to use in the breast cancer gene expression studies. Urokinase Plasminogen Activator was used as a gene of interest. Findings The expression of six HKs (TFRC, GUSB, GAPDH, ACTB, HPRT1 and RPLP0) was investigated using geNorm and NormFinder softwares in forty breast tumor, four normal and eight adjacent tissues. RPLP0 and GAPDH revealed maximum M value, while TFRC demonstrated lowest M value. Conclusions In the present study the most and the least stable genes were TFRC and RPLP0 respectively. TFRC and ACTB were verified as the best combination of two genes for breast cancer quantification. The result of this study shows that in each gene expression analysis HKs selection should be done based on experiment conditions. PMID:21702980

  5. Gene Expression Noise, Fitness Landscapes, and Evolution

    NASA Astrophysics Data System (ADS)

    Charlebois, Daniel

    The stochastic (or noisy) process of gene expression can have fitness consequences for living organisms. For example, gene expression noise facilitates the development of drug resistance by increasing the time scale at which beneficial phenotypic states can be maintained. The present work investigates the relationship between gene expression noise and the fitness landscape. By incorporating the costs and benefits of gene expression, we track how the fluctuation magnitude and timescale of expression noise evolve in simulations of cell populations under stress. We find that properties of expression noise evolve to maximize fitness on the fitness landscape, and that low levels of expression noise emerge when the fitness benefits of gene expression exceed the fitness costs (and that high levels of noise emerge when the costs of expression exceed the benefits). The findings from our theoretical/computational work offer new hypotheses on the development of drug resistance, some of which are now being investigated in evolution experiments in our laboratory using well-characterized synthetic gene regulatory networks in budding yeast. Nserc Postdoctoral Fellowship (Grant No. PDF-453977-2014).

  6. Identification of Cancer Related Genes Using a Comprehensive Map of Human Gene Expression

    PubMed Central

    Lukk, Margus; Xue, Vincent; Parkinson, Helen; Rung, Johan; Brazma, Alvis

    2016-01-01

    Rapid accumulation and availability of gene expression datasets in public repositories have enabled large-scale meta-analyses of combined data. The richness of cross-experiment data has provided new biological insights, including identification of new cancer genes. In this study, we compiled a human gene expression dataset from ∼40,000 publicly available Affymetrix HG-U133Plus2 arrays. After strict quality control and data normalisation the data was quantified in an expression matrix of ∼20,000 genes and ∼28,000 samples. To enable different ways of sample grouping, existing annotations where subjected to systematic ontology assisted categorisation and manual curation. Groups like normal tissues, neoplasmic tissues, cell lines, homoeotic cells and incompletely differentiated cells were created. Unsupervised analysis of the data confirmed global structure of expression consistent with earlier analysis but with more details revealed due to increased resolution. A suitable mixed-effects linear model was used to further investigate gene expression in solid tissue tumours, and to compare these with the respective healthy solid tissues. The analysis identified 1,285 genes with systematic expression change in cancer. The list is significantly enriched with known cancer genes from large, public, peer-reviewed databases, whereas the remaining ones are proposed as new cancer gene candidates. The compiled dataset is publicly available in the ArrayExpress Archive. It contains the most diverse collection of biological samples, making it the largest systematically annotated gene expression dataset of its kind in the public domain. PMID:27322383

  7. Gene expression correlates of unexplained fatigue.

    PubMed

    Whistler, Toni; Taylor, Renee; Craddock, R Cameron; Broderick, Gordon; Klimas, Nancy; Unger, Elizabeth R

    2006-04-01

    Quantitative trait analysis (QTA) can be used to test whether the expression of a particular gene significantly correlates with some ordinal variable. To limit the number of false discoveries in the gene list, a multivariate permutation test can also be performed. The purpose of this study is to identify peripheral blood gene expression correlates of fatigue using quantitative trait analysis on gene expression data from 20,000 genes and fatigue traits measured using the multidimensional fatigue inventory (MFI). A total of 839 genes were statistically associated with fatigue measures. These mapped to biological pathways such as oxidative phosphorylation, gluconeogenesis, lipid metabolism, and several signal transduction pathways. However, more than 50% are not functionally annotated or associated with identified pathways. There is some overlap with genes implicated in other studies using differential gene expression. However, QTA allows detection of alterations that may not reach statistical significance in class comparison analyses, but which could contribute to disease pathophysiology. This study supports the use of phenotypic measures of chronic fatigue syndrome (CFS) and QTA as important for additional studies of this complex illness. Gene expression correlates of other phenotypic measures in the CFS Computational Challenge (C3) data set could be useful. Future studies of CFS should include as many precise measures of disease phenotype as is practical.

  8. Noise Minimisation in Gene Expression Switches

    PubMed Central

    Monteoliva, Diana; McCarthy, Christina B.; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators. PMID:24376783

  9. Noise minimisation in gene expression switches.

    PubMed

    Monteoliva, Diana; McCarthy, Christina B; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators.

  10. Nucleosome repositioning underlies dynamic gene expression

    PubMed Central

    Nocetti, Nicolas; Whitehouse, Iestyn

    2016-01-01

    Nucleosome repositioning at gene promoters is a fundamental aspect of the regulation of gene expression. However, the extent to which nucleosome repositioning is used within eukaryotic genomes is poorly understood. Here we report a comprehensive analysis of nucleosome positions as budding yeast transit through an ultradian cycle in which expression of >50% of all genes is highly synchronized. We present evidence of extensive nucleosome repositioning at thousands of gene promoters as genes are activated and repressed. During activation, nucleosomes are relocated to allow sites of general transcription factor binding and transcription initiation to become accessible. The extent of nucleosome shifting is closely related to the dynamic range of gene transcription and generally related to DNA sequence properties and use of the coactivators TFIID or SAGA. However, dynamic gene expression is not limited to SAGA-regulated promoters and is an inherent feature of most genes. While nucleosome repositioning occurs pervasively, we found that a class of genes required for growth experience acute nucleosome shifting as cells enter the cell cycle. Significantly, our data identify that the ATP-dependent chromatin-remodeling enzyme Snf2 plays a fundamental role in nucleosome repositioning and the expression of growth genes. We also reveal that nucleosome organization changes extensively in concert with phases of the cell cycle, with large, regularly spaced nucleosome arrays being established in mitosis. Collectively, our data and analysis provide a framework for understanding nucleosome dynamics in relation to fundamental DNA-dependent transactions. PMID:26966245

  11. Nucleosome repositioning underlies dynamic gene expression.

    PubMed

    Nocetti, Nicolas; Whitehouse, Iestyn

    2016-03-15

    Nucleosome repositioning at gene promoters is a fundamental aspect of the regulation of gene expression. However, the extent to which nucleosome repositioning is used within eukaryotic genomes is poorly understood. Here we report a comprehensive analysis of nucleosome positions as budding yeast transit through an ultradian cycle in which expression of >50% of all genes is highly synchronized. We present evidence of extensive nucleosome repositioning at thousands of gene promoters as genes are activated and repressed. During activation, nucleosomes are relocated to allow sites of general transcription factor binding and transcription initiation to become accessible. The extent of nucleosome shifting is closely related to the dynamic range of gene transcription and generally related to DNA sequence properties and use of the coactivators TFIID or SAGA. However, dynamic gene expression is not limited to SAGA-regulated promoters and is an inherent feature of most genes. While nucleosome repositioning occurs pervasively, we found that a class of genes required for growth experience acute nucleosome shifting as cells enter the cell cycle. Significantly, our data identify that the ATP-dependent chromatin-remodeling enzyme Snf2 plays a fundamental role in nucleosome repositioning and the expression of growth genes. We also reveal that nucleosome organization changes extensively in concert with phases of the cell cycle, with large, regularly spaced nucleosome arrays being established in mitosis. Collectively, our data and analysis provide a framework for understanding nucleosome dynamics in relation to fundamental DNA-dependent transactions.

  12. Regulation of Flagellar Gene Expression in Bacteria.

    PubMed

    Osterman, I A; Dikhtyar, Yu Yu; Bogdanov, A A; Dontsova, O A; Sergiev, P V

    2015-11-01

    The flagellum of a bacterium is a supramolecular structure of extreme complexity comprising simultaneously both a unique system of protein transport and a molecular machine that enables the bacterial cell movement. The cascade of expression of genes encoding flagellar components is closely coordinated with the steps of molecular machine assembly, constituting an amazing regulatory system. Data on structure, assembly, and regulation of flagellar gene expression are summarized in this review. The regulatory mechanisms and correlation of the process of regulation of gene expression and flagellum assembly known from the literature are described. PMID:26615435

  13. Candidate reference genes for gene expression studies in water lily.

    PubMed

    Luo, Huolin; Chen, Sumei; Wan, Hongjian; Chen, Fadi; Gu, Chunsun; Liu, Zhaolei

    2010-09-01

    The selection of an appropriate reference gene(s) is a prerequisite for the proper interpretation of quantitative Real-Time polymerase chain reaction data. We report the evaluation of eight candidate reference genes across various tissues and treatments in the water lily by the two software packages geNorm and NormFinder. Across all samples, clathrin adaptor complexes medium subunit (AP47) and actin 11 (ACT11) emerged as the most suitable reference genes. Across different tissues, ACT11 and elongation factor 1-alpha (EF1alpha) exhibited a stable expression pattern. ACT11 and AP47 also stably expressed in roots subjected to various treatments, but in the leaves of the same plants the most stably expressed genes were ubiquitin-conjugating enzyme 16 (UBC16) and ACT11. PMID:20452325

  14. Expression of Polarity Genes in Human Cancer

    PubMed Central

    Lin, Wan-Hsin; Asmann, Yan W; Anastasiadis, Panos Z

    2015-01-01

    Polarity protein complexes are crucial for epithelial apical–basal polarity and directed cell migration. Since alterations of these processes are common in cancer, polarity proteins have been proposed to function as tumor suppressors or oncogenic promoters. Here, we review the current understanding of polarity protein functions in epithelial homeostasis, as well as tumor formation and progression. As most previous studies focused on the function of single polarity proteins in simplified model systems, we used a genomics approach to systematically examine and identify the expression profiles of polarity genes in human cancer. The expression profiles of polarity genes were distinct in different human tissues and classified cancer types. Additionally, polarity expression profiles correlated with disease progression and aggressiveness, as well as with identified cancer types, where specific polarity genes were commonly altered. In the case of Scribble, gene expression analysis indicated its common amplification and upregulation in human cancer, suggesting a tumor promoting function. PMID:25991909

  15. Optogenetic Control of Gene Expression in Drosophila

    PubMed Central

    Chan, Yick-Bun; Alekseyenko, Olga V.; Kravitz, Edward A.

    2015-01-01

    To study the molecular mechanism of complex biological systems, it is important to be able to artificially manipulate gene expression in desired target sites with high precision. Based on the light dependent binding of cryptochrome 2 and a cryptochrome interacting bHLH protein, we developed a split lexA transcriptional activation system for use in Drosophila that allows regulation of gene expression in vivo using blue light or two-photon excitation. We show that this system offers high spatiotemporal resolution by inducing gene expression in tissues at various developmental stages. In combination with two-photon excitation, gene expression can be manipulated at precise sites in embryos, potentially offering an important tool with which to examine developmental processes. PMID:26383635

  16. Expression of polarity genes in human cancer.

    PubMed

    Lin, Wan-Hsin; Asmann, Yan W; Anastasiadis, Panos Z

    2015-01-01

    Polarity protein complexes are crucial for epithelial apical-basal polarity and directed cell migration. Since alterations of these processes are common in cancer, polarity proteins have been proposed to function as tumor suppressors or oncogenic promoters. Here, we review the current understanding of polarity protein functions in epithelial homeostasis, as well as tumor formation and progression. As most previous studies focused on the function of single polarity proteins in simplified model systems, we used a genomics approach to systematically examine and identify the expression profiles of polarity genes in human cancer. The expression profiles of polarity genes were distinct in different human tissues and classified cancer types. Additionally, polarity expression profiles correlated with disease progression and aggressiveness, as well as with identified cancer types, where specific polarity genes were commonly altered. In the case of Scribble, gene expression analysis indicated its common amplification and upregulation in human cancer, suggesting a tumor promoting function.

  17. Regulation of Gene Expression in Protozoa Parasites

    PubMed Central

    Gomez, Consuelo; Esther Ramirez, M.; Calixto-Galvez, Mercedes; Medel, Olivia; Rodríguez, Mario A.

    2010-01-01

    Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis. PMID:20204171

  18. Dynamic modeling of gene expression data

    NASA Technical Reports Server (NTRS)

    Holter, N. S.; Maritan, A.; Cieplak, M.; Fedoroff, N. V.; Banavar, J. R.

    2001-01-01

    We describe the time evolution of gene expression levels by using a time translational matrix to predict future expression levels of genes based on their expression levels at some initial time. We deduce the time translational matrix for previously published DNA microarray gene expression data sets by modeling them within a linear framework by using the characteristic modes obtained by singular value decomposition. The resulting time translation matrix provides a measure of the relationships among the modes and governs their time evolution. We show that a truncated matrix linking just a few modes is a good approximation of the full time translation matrix. This finding suggests that the number of essential connections among the genes is small.

  19. Gene Positioning Effects on Expression in Eukaryotes.

    PubMed

    Nguyen, Huy Q; Bosco, Giovanni

    2015-01-01

    The packaging and organization of the genome within the eukaryotic interphase nucleus directly influence how the genes are expressed. An underappreciated aspect of genome structure is that it is highly dynamic and that the physical positioning of a gene can impart control over its transcriptional status. In this review, we assess the current knowledge of how gene positioning at different levels of genome organization can directly influence gene expression during interphase. The levels of organization discussed include chromatin looping, topologically associated domains, chromosome territories, and nuclear compartments. We discuss specific studies demonstrating that gene positioning is a dynamic and highly regulated feature of the eukaryotic genome that allows for the essential spatiotemporal regulation of genes.

  20. Homeobox genes expressed during echinoderm arm regeneration.

    PubMed

    Ben Khadra, Yousra; Said, Khaled; Thorndyke, Michael; Martinez, Pedro

    2014-04-01

    Regeneration in echinoderms has proved to be more amenable to study in the laboratory than the more classical vertebrate models, since the smaller genome size and the absence of multiple orthologs for different genes in echinoderms simplify the analysis of gene function during regeneration. In order to understand the role of homeobox-containing genes during arm regeneration in echinoderms, we isolated the complement of genes belonging to the Hox class that are expressed during this process in two major echinoderm groups: asteroids (Echinaster sepositus and Asterias rubens) and ophiuroids (Amphiura filiformis), both of which show an extraordinary capacity for regeneration. By exploiting the sequence conservation of the homeobox, putative orthologs of several Hox genes belonging to the anterior, medial, and posterior groups were isolated. We also report the isolation of a few Hox-like genes expressed in the same systems. PMID:24309817

  1. Endogenous Reference Genes for Gene Expression Studies on Bicuspid Aortic Valve Associated Aortopathy in Humans

    PubMed Central

    Harrison, Oliver J.; Moorjani, Narain; Torrens, Christopher; Ohri, Sunil K.; Cagampang, Felino R.

    2016-01-01

    Bicuspid aortic valve (BAV) disease is the most common congenital cardiac abnormality and predisposes patients to life-threatening aortic complications including aortic aneurysm. Quantitative real-time reverse transcription PCR (qRT-PCR) is one of the most commonly used methods to investigate underlying molecular mechanisms involved in aortopathy. The accuracy of the gene expression data is dependent on normalization by appropriate housekeeping (HK) genes, whose expression should remain constant regardless of aortic valve morphology, aortic diameter and other factors associated with aortopathy. Here, we identified an appropriate set of HK genes to be used as endogenous reference for quantifying gene expression in ascending aortic tissue using a spin column-based RNA extraction method. Ascending aortic biopsies were collected intra-operatively from patients undergoing aortic valve and/or ascending aortic surgery. These patients had BAV or tricuspid aortic valve (TAV), and the aortas were either dilated (≥4.5cm) or undilated. The cohort had an even distribution of gender, valve disease and hypertension. The expression stability of 12 reference genes were investigated (ATP5B, ACTB, B2M, CYC1, EIF4A2, GAPDH, SDHA, RPL13A, TOP1, UBC, YWHAZ, and 18S) using geNorm software. The most stable HK genes were found to be GAPDH, UBC and ACTB. Both GAPDH and UBC demonstrated relative stability regardless of valve morphology, aortic diameter, gender and age. The expression of B2M and SDHA were found to be the least stable HK genes. We propose the use of GAPDH, UBC and ACTB as reference genes for gene expression studies of BAV aortopathy using ascending aortic tissue. PMID:27727313

  2. Mechanisms of control of gene expression

    SciTech Connect

    Cullen, B.; Gage, L.P.; Siddiqui, M.A.Q.; Skalka, A.M.; Weissbach, H.

    1987-01-01

    This book examines an array of topics on the regulation of gene expression, including an examination of DNA-protein interactions and the role of oncogene proteins in normal and abnormal cellular responses. The book focuses on the control of mRNA transcription in eykaryotes and delineates other areas including gene regulation in prokaryotes and control of stable RNA synthesis.

  3. Perspectives: Gene Expression in Fisheries Management

    USGS Publications Warehouse

    Nielsen, Jennifer L.; Pavey, Scott A.

    2010-01-01

    Functional genes and gene expression have been connected to physiological traits linked to effective production and broodstock selection in aquaculture, selective implications of commercial fish harvest, and adaptive changes reflected in non-commercial fish populations subject to human disturbance and climate change. Gene mapping using single nucleotide polymorphisms (SNPs) to identify functional genes, gene expression (analogue microarrays and real-time PCR), and digital sequencing technologies looking at RNA transcripts present new concepts and opportunities in support of effective and sustainable fisheries. Genomic tools have been rapidly growing in aquaculture research addressing aspects of fish health, toxicology, and early development. Genomic technologies linking effects in functional genes involved in growth, maturation and life history development have been tied to selection resulting from harvest practices. Incorporating new and ever-increasing knowledge of fish genomes is opening a different perspective on local adaptation that will prove invaluable in wild fish conservation and management. Conservation of fish stocks is rapidly incorporating research on critical adaptive responses directed at the effects of human disturbance and climate change through gene expression studies. Genomic studies of fish populations can be generally grouped into three broad categories: 1) evolutionary genomics and biodiversity; 2) adaptive physiological responses to a changing environment; and 3) adaptive behavioral genomics and life history diversity. We review current genomic research in fisheries focusing on those that use microarrays to explore differences in gene expression among phenotypes and within or across populations, information that is critically important to the conservation of fish and their relationship to humans.

  4. Reading Genomes and Controlling Gene Expression

    NASA Astrophysics Data System (ADS)

    Libchaber, Albert

    2000-03-01

    Molecular recognition of DNA sequences is achieved by DNA hybridization of complementary sequences. We present various scenarios for optimization, leading to microarrays and global measurement. Gene expression can be controlled using gene constructs immobilized on a template with micron scale temperature heaters. We will discuss and present results on protein microarrays.

  5. Bayesian modeling of differential gene expression.

    PubMed

    Lewin, Alex; Richardson, Sylvia; Marshall, Clare; Glazier, Anne; Aitman, Tim

    2006-03-01

    We present a Bayesian hierarchical model for detecting differentially expressing genes that includes simultaneous estimation of array effects, and show how to use the output for choosing lists of genes for further investigation. We give empirical evidence that expression-level dependent array effects are needed, and explore different nonlinear functions as part of our model-based approach to normalization. The model includes gene-specific variances but imposes some necessary shrinkage through a hierarchical structure. Model criticism via posterior predictive checks is discussed. Modeling the array effects (normalization) simultaneously with differential expression gives fewer false positive results. To choose a list of genes, we propose to combine various criteria (for instance, fold change and overall expression) into a single indicator variable for each gene. The posterior distribution of these variables is used to pick the list of genes, thereby taking into account uncertainty in parameter estimates. In an application to mouse knockout data, Gene Ontology annotations over- and underrepresented among the genes on the chosen list are consistent with biological expectations.

  6. Inducible gene expression in transgenic Xenopus embryos.

    PubMed

    Wheeler, G N; Hamilton, F S; Hoppler, S

    2000-07-13

    The amphibian Xenopus laevis has been successfully used for many years as a model system for studying vertebrate development. Because of technical limitations, however, molecular investigations have mainly concentrated on early stages. We have developed a straightforward method for stage-specific induction of gene expression in transgenic Xenopus embryos [1] [2]. This method is based on the Xenopus heat shock protein 70 (Xhsp70 [3]) promoter driving the expression of desired gene products. We found that ubiquitous expression of the transgene is induced upon relatively mild heat treatment. Green fluorescent protein (GFP) was used as a marker to monitor successful induction of gene expression in transgenic embryos. We used this method to study the stage specificity of Wnt signalling function. Transient ectopic Wnt-8 expression during early neurulation was sufficient to repress anterior head development and this capacity was restricted to early stages of neurulation. By transient over-expression at different stages of development, we show that frizzled-7 disrupted morphogenesis sequentially from anterior to posterior along the dorsal axis as development proceeds. These results demonstrate that this method for inducible gene expression in transgenic Xenopus embryos will be a very powerful tool for temporal analysis of gene function and for studying molecular mechanisms of vertebrate organogenesis.

  7. Assessing Gene Expression of the Endocannabinoid System.

    PubMed

    Pucci, Mariangela; D'Addario, Claudio

    2016-01-01

    Real-time quantitative reverse transcription polymerase chain reaction (real-time qRT-PCR), a major development of PCR technology, is a powerful and sensitive gene analysis technique that revolutionized the field of measuring gene expression. Here, we describe in detail RNA extraction, reverse transcription (RT), and relative quantification of genes belonging to the endocannabinoid system in mouse, rat, or human samples. PMID:27245909

  8. Screening Reliable Reference Genes for RT-qPCR Analysis of Gene Expression in Moringa oleifera

    PubMed Central

    Deng, Li-Ting; Wu, Yu-Ling; Li, Jun-Cheng; OuYang, Kun-Xi; Ding, Mei-Mei; Zhang, Jun-Jie; Li, Shu-Qi; Lin, Meng-Fei; Chen, Han-Bin; Hu, Xin-Sheng; Chen, Xiao-Yang

    2016-01-01

    Moringa oleifera is a promising plant species for oil and forage, but its genetic improvement is limited. Our current breeding program in this species focuses on exploiting the functional genes associated with important agronomical traits. Here, we screened reliable reference genes for accurately quantifying the expression of target genes using the technique of real-time quantitative polymerase chain reaction (RT-qPCR) in M. oleifera. Eighteen candidate reference genes were selected from a transcriptome database, and their expression stabilities were examined in 90 samples collected from the pods in different developmental stages, various tissues, and the roots and leaves under different conditions (low or high temperature, sodium chloride (NaCl)- or polyethyleneglycol (PEG)- simulated water stress). Analyses with geNorm, NormFinder and BestKeeper algorithms revealed that the reliable reference genes differed across sample designs and that ribosomal protein L1 (RPL1) and acyl carrier protein 2 (ACP2) were the most suitable reference genes in all tested samples. The experiment results demonstrated the significance of using the properly validated reference genes and suggested the use of more than one reference gene to achieve reliable expression profiles. In addition, we applied three isotypes of the superoxide dismutase (SOD) gene that are associated with plant adaptation to abiotic stress to confirm the efficacy of the validated reference genes under NaCl and PEG water stresses. Our results provide a valuable reference for future studies on identifying important functional genes from their transcriptional expressions via RT-qPCR technique in M. oleifera. PMID:27541138

  9. Screening Reliable Reference Genes for RT-qPCR Analysis of Gene Expression in Moringa oleifera.

    PubMed

    Deng, Li-Ting; Wu, Yu-Ling; Li, Jun-Cheng; OuYang, Kun-Xi; Ding, Mei-Mei; Zhang, Jun-Jie; Li, Shu-Qi; Lin, Meng-Fei; Chen, Han-Bin; Hu, Xin-Sheng; Chen, Xiao-Yang

    2016-01-01

    Moringa oleifera is a promising plant species for oil and forage, but its genetic improvement is limited. Our current breeding program in this species focuses on exploiting the functional genes associated with important agronomical traits. Here, we screened reliable reference genes for accurately quantifying the expression of target genes using the technique of real-time quantitative polymerase chain reaction (RT-qPCR) in M. oleifera. Eighteen candidate reference genes were selected from a transcriptome database, and their expression stabilities were examined in 90 samples collected from the pods in different developmental stages, various tissues, and the roots and leaves under different conditions (low or high temperature, sodium chloride (NaCl)- or polyethyleneglycol (PEG)- simulated water stress). Analyses with geNorm, NormFinder and BestKeeper algorithms revealed that the reliable reference genes differed across sample designs and that ribosomal protein L1 (RPL1) and acyl carrier protein 2 (ACP2) were the most suitable reference genes in all tested samples. The experiment results demonstrated the significance of using the properly validated reference genes and suggested the use of more than one reference gene to achieve reliable expression profiles. In addition, we applied three isotypes of the superoxide dismutase (SOD) gene that are associated with plant adaptation to abiotic stress to confirm the efficacy of the validated reference genes under NaCl and PEG water stresses. Our results provide a valuable reference for future studies on identifying important functional genes from their transcriptional expressions via RT-qPCR technique in M. oleifera. PMID:27541138

  10. Application of multidisciplinary analysis to gene expression.

    SciTech Connect

    Wang, Xuefel; Kang, Huining; Fields, Chris; Cowie, Jim R.; Davidson, George S.; Haaland, David Michael; Sibirtsev, Valeriy; Mosquera-Caro, Monica P.; Xu, Yuexian; Martin, Shawn Bryan; Helman, Paul; Andries, Erik; Ar, Kerem; Potter, Jeffrey; Willman, Cheryl L.; Murphy, Maurice H.

    2004-01-01

    Molecular analysis of cancer, at the genomic level, could lead to individualized patient diagnostics and treatments. The developments to follow will signal a significant paradigm shift in the clinical management of human cancer. Despite our initial hopes, however, it seems that simple analysis of microarray data cannot elucidate clinically significant gene functions and mechanisms. Extracting biological information from microarray data requires a complicated path involving multidisciplinary teams of biomedical researchers, computer scientists, mathematicians, statisticians, and computational linguists. The integration of the diverse outputs of each team is the limiting factor in the progress to discover candidate genes and pathways associated with the molecular biology of cancer. Specifically, one must deal with sets of significant genes identified by each method and extract whatever useful information may be found by comparing these different gene lists. Here we present our experience with such comparisons, and share methods developed in the analysis of an infant leukemia cohort studied on Affymetrix HG-U95A arrays. In particular, spatial gene clustering, hyper-dimensional projections, and computational linguistics were used to compare different gene lists. In spatial gene clustering, different gene lists are grouped together and visualized on a three-dimensional expression map, where genes with similar expressions are co-located. In another approach, projections from gene expression space onto a sphere clarify how groups of genes can jointly have more predictive power than groups of individually selected genes. Finally, online literature is automatically rearranged to present information about genes common to multiple groups, or to contrast the differences between the lists. The combination of these methods has improved our understanding of infant leukemia. While the complicated reality of the biology dashed our initial, optimistic hopes for simple answers from

  11. Identification of housekeeping genes suitable for gene expression analysis in the pearl mussel, Hyriopsis cumingii, during biomineralization.

    PubMed

    Bai, Zhiyi; Lin, Jingyun; Ma, Keyi; Wang, Guiling; Niu, Donghong; Li, Jiale

    2014-08-01

    Quantitative real-time polymerase chain reaction is a sensitive technique for quantifying gene expression levels. One or more appropriate reference genes must be selected to accurately compare mRNA transcript levels across different samples and tissues. The freshwater pearl, Hyriopsis cumingii (Lea), is an important economic species cultured in China. To date, no reference genes for gene expression analysis in this species have been validated. This study aimed to compare the relative expression of seven housekeeping genes across different tissue types and in the mantle or pearl sac during three biomineralization processes: seasonal shell growth, shell healing and pearl-sac formation in H. cumingii. Three programs evaluated the expression stabilities of the seven genes: BestKeeper, geNorm and NormFinder. The beta actin gene (ACTB), commonly used as a housekeeping gene in many studies, was the least stable. The expressions of Ubiquitin (Ubi) and Ribosomal protein L18 (Rpl18) and Elongation factor 1-alpha (EF1α) were more stable than the remaining four genes. Therefore, we suggest that Ubi, Rpl18 and EF1α are suitable reference genes. The three selected reference genes are expected to facilitate analysis of gene expressions during shell or pearl formation in H. cumingii.

  12. Thyroid-specific gene expression in chondrocytes.

    PubMed

    Endo, Toyoshi; Kobayashi, Tetsuro

    2011-12-16

    Previously, we demonstrated that Runx2 (Cbfa1/AML3), a chondrocyte-specific transcription factor, is expressed in thyroid glands of mice, where it stimulates expression of the thyroglobulin (Tg) gene. Here, we reverse transcribed thyroid transcription factor-1 (TTF-1), Pax-8, Tg, thyroid peroxidase (TPO) and Na(+)/I(-) symporter (NIS) cDNAs from mouse trachea and bronchus RNA samples, but were unable to recover these cDNAs from mouse liver RNA samples. Tg mRNA levels in trachea and bronchus were about 5.1% and 2.1% of those in thyroid glands. ATDC-5 cells, cultured chondrocytes, expressed about 30-fold more Tg mRNA than undifferentiated cells. Gel shift and Tg gene reporter assay revealed that TTF-1 stimulated Tg gene expression in these cells. These results indicate that chondrocytes turn on some aspects of the thyroid gene expression program and that TTF-1 plays important roles in Tg gene expression in chondrocyte. PMID:21945616

  13. Regulation of gene expression in human tendinopathy

    PubMed Central

    2011-01-01

    Background Chronic tendon injuries, also known as tendinopathies, are common among professional and recreational athletes. These injuries result in a significant amount of morbidity and health care expenditure, yet little is known about the molecular mechanisms leading to tendinopathy. Methods We have used histological evaluation and molecular profiling to determine gene expression changes in 23 human patients undergoing surgical procedures for the treatment of chronic tendinopathy. Results Diseased tendons exhibit altered extracellular matrix, fiber disorientation, increased cellular content and vasculature, and the absence of inflammatory cells. Global gene expression profiling identified 983 transcripts with significantly different expression patterns in the diseased tendons. Global pathway analysis further suggested altered expression of extracellular matrix proteins and the lack of an appreciable inflammatory response. Conclusions Identification of the pathways and genes that are differentially regulated in tendinopathy samples will contribute to our understanding of the disease and the development of novel therapeutics. PMID:21539748

  14. Links between Transcription, Environmental Adaptation and Gene Variability in Escherichia coli: Correlations between Gene Expression and Gene Variability Reflect Growth Efficiencies.

    PubMed

    Feugeas, Jean-Paul; Tourret, Jerome; Launay, Adrien; Bouvet, Odile; Hoede, Claire; Denamur, Erick; Tenaillon, Olivier

    2016-10-01

    Gene expression is known to be the principle factor explaining how fast genes evolve. Highly transcribed genes evolve slowly because any negative impact caused by a particular mutation is magnified by protein abundance. However, gene expression is a phenotype that depends both on the environment and on the strains or species. We studied this phenotypic plasticity by analyzing the transcriptome profiles of four Escherichia coli strains grown in three different culture media, and explored how expression variability was linked to gene allelic diversity. Genes whose expression changed according to the media and not to the strains were less polymorphic than other genes. Genes for which transcription depended predominantly on the strain were more polymorphic than other genes and were involved in sensing and responding to environmental changes, with an overrepresentation of two-component system genes. Surprisingly, we found that the correlation between transcription and gene diversity was highly variable among growth conditions and could be used to quantify growth efficiency of a strain in a medium. Genetic variability was found to increase with gene expression in poor growth conditions. As such conditions are also characterized by down-regulation of all DNA repair systems, including transcription-coupled repair, we suggest that gene expression under stressful conditions may be mutagenic and thus leads to a variability in mutation rate among genes in the genome which contributes to the pattern of protein evolution.

  15. Intergrin gene expression profiles of humanhepatocellular carcinoma

    PubMed Central

    Liu, Lian-Xin; Jiang, Hong-Chi; Liu, Zhi-Hua; Zhou, Jing; Zhang, Wei-Hui; Zhu, An-Long; Wang, Xiu-Qin; Wu, Min

    2002-01-01

    AIM: To investigate gene expression profiles of intergrin genes in hepatocellular carcinoma (HCC) through the usage of Atlas Human Cancer Array membranes, semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and Northern blot. METHODS: Hybridization of cDNA array membrane was performed with α 32P-labeled cDNA probes synthesized from RNA isolated from hepatocellular carcinoma and adjacent non-cirrhotic liver. AtlasImage, which is a software specific to array, was used to analyze the result. RT-PCR of 24 pairs specimen and Northern blot of 4 pairs specimen were used to confirm the expression pattern of some intergrin genes identified by Atlas arrays hybridization. RESULTS: Among 588 genes spotted in membrane, 17 genes were related to intergrin. Four genes were up-regulated, such as intergrin alpha8, beta1, beta7 and beta8 in HCC. Whereas there were no genes down-regulated in HCC. RT-PCR and Northern blot analysis of intergrin beta1 gene gave results consistent with cDNA array findings. CONCLUSION: Investigation of these intergrin genes should help to disclose the molecular mechanism of the cell adhesion, invasive and metastasis of HCC. A few genes are reported to have changed in HCC for the first time. The quick and high-throughout method of profiling gene expression by cDNA array provides us overview of key factors that may involved in HCC, and may find the clue of the study of HCC metastasis and molecular targets of anti-metastasis therapy. The precise relationship between the altered genes and HCC is a matter of further investigation. PMID:12174369

  16. Systematic analysis of somatic mutations impacting gene expression in 12 tumour types

    PubMed Central

    Ding, Jiarui; McConechy, Melissa K.; Horlings, Hugo M.; Ha, Gavin; Chun Chan, Fong; Funnell, Tyler; Mullaly, Sarah C.; Reimand, Jüri; Bashashati, Ali; Bader, Gary D.; Huntsman, David; Aparicio, Samuel; Condon, Anne; Shah, Sohrab P.

    2015-01-01

    We present a novel hierarchical Bayes statistical model, xseq, to systematically quantify the impact of somatic mutations on expression profiles. We establish the theoretical framework and robust inference characteristics of the method using computational benchmarking. We then use xseq to analyse thousands of tumour data sets available through The Cancer Genome Atlas, to systematically quantify somatic mutations impacting expression profiles. We identify 30 novel cis-effect tumour suppressor gene candidates, enriched in loss-of-function mutations and biallelic inactivation. Analysis of trans-effects of mutations and copy number alterations with xseq identifies mutations in 150 genes impacting expression networks, with 89 novel predictions. We reveal two important novel characteristics of mutation impact on expression: (1) patients harbouring known driver mutations exhibit different downstream gene expression consequences; (2) expression patterns for some mutations are stable across tumour types. These results have critical implications for identification and interpretation of mutations with consequent impact on transcription in cancer. PMID:26436532

  17. Noise minimization in eukaryotic gene expression

    SciTech Connect

    Fraser, Hunter B.; Hirsh, Aaron E.; Giaever, Guri; Kumm, Jochen; Eisen, Michael B.

    2004-01-15

    All organisms have elaborate mechanisms to control rates of protein production. However, protein production is also subject to stochastic fluctuations, or noise. Several recent studies in Saccharomyces cerevisiae and Escherichia coli have investigated the relationship between transcription and translation rates and stochastic fluctuations in protein levels, or more generally, how such randomness is a function of intrinsic and extrinsic factors. However, the fundamental question of whether stochasticity in protein expression is generally biologically relevant has not been addressed, and it remains unknown whether random noise in the protein production rate of most genes significantly affects the fitness of any organism. We propose that organisms should be particularly sensitive to variation in the protein levels of two classes of genes: genes whose deletion is lethal to the organism and genes that encode subunits of multiprotein complexes. Using an experimentally verified model of stochastic gene expression in S. cerevisiae, we estimate the noise in protein production for nearly every yeast gene, and confirm our prediction that the production of essential and complex-forming proteins involves lower levels of noise than does the production of most other genes. Our results support the hypothesis that noise in gene expression is a biologically important variable, is generally detrimental to organismal fitness, and is subject to natural selection.

  18. Soybean physiology and gene expression during drought.

    PubMed

    Stolf-Moreira, R; Medri, M E; Neumaier, N; Lemos, N G; Pimenta, J A; Tobita, S; Brogin, R L; Marcelino-Guimarães, F C; Oliveira, M C N; Farias, J R B; Abdelnoor, R V; Nepomuceno, A L

    2010-10-05

    Soybean genotypes MG/BR46 (Conquista) and BR16, drought-tolerant and -sensitive, respectively, were compared in terms of morphophysiological and gene-expression responses to water stress during two stages of development. Gene-expression analysis showed differential responses in Gmdreb1a and Gmpip1b mRNA expression within 30 days of water-deficit initiation in MG/BR46 (Conquista) plants. Within 45 days of initiating stress, Gmp5cs and Gmpip1b had relatively higher expression. Initially, BR16 showed increased expression only for Gmdreb1a, and later (45 days) for Gmp5cs, Gmdefensin and Gmpip1b. Only BR16 presented down-regulated expression of genes, such as Gmp5cs and Gmpip1b, 30 days after the onset of moisture stress, and Gmgols after 45 days of stress. The faster perception of water stress in MG/BR46 (Conquista) and the better maintenance of up-regulated gene expression than in the sensitive BR16 genotype imply mechanisms by which the former is better adapted to tolerate moisture deficiency.

  19. Nanobarcode gene expression monitoring system for potential miniaturized space applications

    NASA Astrophysics Data System (ADS)

    Ruan, Weiming; Eastman, P. Scott; Cooke, Patrick A.; Park, Jennifer S.; Chu, Julia S. F.; Gray, Joe W.; Li, Song; Chen, Fanqing Frank

    Manned mission to space has been threatened by various cosmos risks including radiation, mirogravity, vacuum, confinement, etc., which may cause genetic variations of astronauts and eventually lead to damages of their health. Thus, the development of small biomedical devices, which can monitor astronaut gene expression changes, is useful for future long-term space missions. Using magnetic microbeads packed with nanocrystal quantum dots at controlled ratios, we were able to generate highly multiplexed nanobarcodes, which can encode a flexible panel of genes. Also, by using a reporter quantum dot, this nanobarcode platform can monitor and quantify gene expression level with improved speed and sensitivity. As a comparison, we studied TGF-β1 induced transcription changes in human bone marrow mesenchymal stem cells with both the nanobarcode microbead system and the Affymetrix GeneChip ® HTA system, which is currently considered as the industrial standard. Though using only 1/20 of the sample RNA, the nanobarcode system showed sensitivity equivalent to Affymetrix GeneChip ® system. The coefficient of variation, dynamic range, and accuracy of the nanobarcodes measurement is equivalent to that of the GeneChip ® HTA system. Therefore, this newly invented nanobarcode microbead platform is thought to be sensitive, flexible, cost-effective and accurate in a level equivalent to the conventional methods. As an extension of the use of this new platform, spacecrafts may carry this miniaturized system as a diagnostic tool for the astronauts.

  20. Reference genes for quantitative gene expression studies in multiple avian species.

    PubMed

    Olias, Philipp; Adam, Iris; Meyer, Anne; Scharff, Constance; Gruber, Achim D

    2014-01-01

    Quantitative real-time PCR (qPCR) rapidly and reliably quantifies gene expression levels across different experimental conditions. Selection of suitable reference genes is essential for meaningful normalization and thus correct interpretation of data. In recent years, an increasing number of avian species other than the chicken has been investigated molecularly, highlighting the need for an experimentally validated pan-avian primer set for reference genes. Here we report testing a set for 14 candidate reference genes (18S, ABL, GAPDH, GUSB, HMBS, HPRT, PGK1, RPL13, RPL19, RPS7, SDHA, TFRC, VIM, YWHAZ) on different tissues of the mallard (Anas platyrhynchos), domestic chicken (Gallus gallus domesticus), common crane (Grus grus), white-tailed eagle (Haliaeetus albicilla), domestic turkey (Meleagris gallopavo f. domestica), cockatiel (Nymphicus hollandicus), Humboldt penguin (Sphenicus humboldti), ostrich (Struthio camelus) and zebra finch (Taeniopygia guttata), spanning a broad range of the phylogenetic tree of birds. Primer pairs for six to 11 genes were successfully established for each of the nine species. As a proof of principle, we analyzed expression levels of 10 candidate reference genes as well as FOXP2 and the immediate early genes, EGR1 and CFOS, known to be rapidly induced by singing in the avian basal ganglia. We extracted RNA from microbiopsies of the striatal song nucleus Area X of adult male zebra finches after they had sang or remained silent. Using three different statistical algorithms, we identified five genes (18S, PGK1, RPS7, TFRC, YWHAZ) that were stably expressed within each group and also between the singing and silent conditions, establishing them as suitable reference genes. In conclusion, the newly developed pan-avian primer set allows accurate normalization and quantification of gene expression levels in multiple avian species. PMID:24926893

  1. Validation of reference genes for RT-qPCR studies of gene expression in banana fruit under different experimental conditions.

    PubMed

    Chen, Lei; Zhong, Hai-ying; Kuang, Jian-fei; Li, Jian-guo; Lu, Wang-jin; Chen, Jian-ye

    2011-08-01

    Reverse transcription quantitative real-time PCR (RT-qPCR) is a sensitive technique for quantifying gene expression, but its success depends on the stability of the reference gene(s) used for data normalization. Only a few studies on validation of reference genes have been conducted in fruit trees and none in banana yet. In the present work, 20 candidate reference genes were selected, and their expression stability in 144 banana samples were evaluated and analyzed using two algorithms, geNorm and NormFinder. The samples consisted of eight sample sets collected under different experimental conditions, including various tissues, developmental stages, postharvest ripening, stresses (chilling, high temperature, and pathogen), and hormone treatments. Our results showed that different suitable reference gene(s) or combination of reference genes for normalization should be selected depending on the experimental conditions. The RPS2 and UBQ2 genes were validated as the most suitable reference genes across all tested samples. More importantly, our data further showed that the widely used reference genes, ACT and GAPDH, were not the most suitable reference genes in many banana sample sets. In addition, the expression of MaEBF1, a gene of interest that plays an important role in regulating fruit ripening, under different experimental conditions was used to further confirm the validated reference genes. Taken together, our results provide guidelines for reference gene(s) selection under different experimental conditions and a foundation for more accurate and widespread use of RT-qPCR in banana. PMID:21505864

  2. Inferring differentiation pathways from gene expression

    PubMed Central

    Costa, Ivan G.; Roepcke, Stefan; Hafemeister, Christoph; Schliep, Alexander

    2008-01-01

    Motivation: The regulation of proliferation and differentiation of embryonic and adult stem cells into mature cells is central to developmental biology. Gene expression measured in distinguishable developmental stages helps to elucidate underlying molecular processes. In previous work we showed that functional gene modules, which act distinctly in the course of development, can be represented by a mixture of trees. In general, the similarities in the gene expression programs of cell populations reflect the similarities in the differentiation path. Results: We propose a novel model for gene expression profiles and an unsupervised learning method to estimate developmental similarity and infer differentiation pathways. We assess the performance of our model on simulated data and compare it with favorable results to related methods. We also infer differentiation pathways and predict functional modules in gene expression data of lymphoid development. Conclusions: We demonstrate for the first time how, in principal, the incorporation of structural knowledge about the dependence structure helps to reveal differentiation pathways and potentially relevant functional gene modules from microarray datasets. Our method applies in any area of developmental biology where it is possible to obtain cells of distinguishable differentiation stages. Availability: The implementation of our method (GPL license), data and additional results are available at http://algorithmics.molgen.mpg.de/Supplements/InfDif/ Contact: filho@molgen.mpg.de, schliep@molgen.mpg.de Supplementary information: Supplementary data is available at Bioinformatics online. PMID:18586709

  3. Human AZU-1 gene, variants thereof and expressed gene products

    DOEpatents

    Chen, Huei-Mei; Bissell, Mina

    2004-06-22

    A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

  4. [Expression and regulation of the SOST gene].

    PubMed

    Qin, Long-Juan; Ding, Da-Xia; Cui, Lu-Lu; Huang, Qing-Yang

    2013-08-01

    Sclerostin(SOST), mainly expressed in osteocytes, is a negative regulator of bone formation. Hormones PTH and E2 inhibit the expression of the SOST gene. Transcription factors Osterix, Runx2, and Mef2c promote the SOST expression, while Sirt1 negatively regulates the SOST expression. In addition, the expression of the SOST gene is regulated by epigenetic mechanisms, such as DNA methylation and microRNA. Mutations in the SOST gene, which cause sclerosteosis and Van Buchem diseases, are associated with osteoporosis. Wnt and BMP are two important signaling pathways in bone metabolic regulation. SOST can regulate osteoblastic differentiation and bone formation by binding type I/II receptors and co-receptor LRP5/6 to inhibit BMP and Wnt signaling pathways. Suppression of SOST provides a new approach for osteoporosis treatment. This review covers the structure, function and expression regulation of the SOST gene, human disease association, mechanism in the regulation of bone metabolism and prospect in clinical application.

  5. Alternative-splicing-mediated gene expression

    NASA Astrophysics Data System (ADS)

    Wang, Qianliang; Zhou, Tianshou

    2014-01-01

    Alternative splicing (AS) is a fundamental process during gene expression and has been found to be ubiquitous in eukaryotes. However, how AS impacts gene expression levels both quantitatively and qualitatively remains to be fully explored. Here, we analyze two common models of gene expression, each incorporating a simple splice mechanism that a pre-mRNA is spliced into two mature mRNA isoforms in a probabilistic manner. In the constitutive expression case, we show that the steady-state molecular numbers of two mature mRNA isoforms follow mutually independent Poisson distributions. In the bursting expression case, we demonstrate that the tail decay of the steady-state distribution for both mature mRNA isoforms that in general are not mutually independent can be characterized by the product of mean burst size and splicing probability. In both cases, we find that AS can efficiently modulate both the variability (measured by variance) and the noise level of the total mature mRNA, and in particular, the latter is always lower than the noise level of the pre-mRNA, implying that AS always reduces the noise. These results altogether reveal that AS is a mechanism of efficiently controlling the gene expression noise.

  6. Gene Expression Profiling of Infecting Microbes Using a Digital Bar-coding Platform

    PubMed Central

    Xu, Wenjie; Solis, Norma V.; Filler, Scott G.; Mitchell, Aaron P.

    2016-01-01

    For most mammalian pathogens, gene expression profiling studies have been limited by technical difficulties to accurately quantify pathogen gene transcripts from infected tissues. Pathogen RNA constitutes a tiny portion of the total RNA isolated from infected tissue samples. Both microarray and RNAseq technologies have difficulties in generating reliable reads for weakly expressed pathogen genes. Mutant pathogen strains with reduced in vivo proliferation pose an even bigger challenge. Here we describe an in vivo gene expression profiling protocol that is very fast, extremely sensitive and highly reproducible. We developed this protocol during our investigation of the fungal pathogen Candida albicans in a murine model of hematogenously disseminated candidiasis. Using this protocol, we have documented time courses of dynamically regulated C. albicans gene expression during kidney infection, and discovered unexpected features of gene expression responses to antifungal drug treatment in vivo. PMID:26863547

  7. Leveraging Distant Relatedness to Quantify Human Mutation and Gene-Conversion Rates

    PubMed Central

    Palamara, Pier Francesco; Francioli, Laurent C.; Wilton, Peter R.; Genovese, Giulio; Gusev, Alexander; Finucane, Hilary K.; Sankararaman, Sriram; Sunyaev, Shamil R.; de Bakker, Paul I.W.; Wakeley, John; Pe’er, Itsik; Price, Alkes L.

    2015-01-01

    The rate at which human genomes mutate is a central biological parameter that has many implications for our ability to understand demographic and evolutionary phenomena. We present a method for inferring mutation and gene-conversion rates by using the number of sequence differences observed in identical-by-descent (IBD) segments together with a reconstructed model of recent population-size history. This approach is robust to, and can quantify, the presence of substantial genotyping error, as validated in coalescent simulations. We applied the method to 498 trio-phased sequenced Dutch individuals and inferred a point mutation rate of 1.66 × 10−8 per base per generation and a rate of 1.26 × 10−9 for <20 bp indels. By quantifying how estimates varied as a function of allele frequency, we inferred the probability that a site is involved in non-crossover gene conversion as 5.99 × 10−6. We found that recombination does not have observable mutagenic effects after gene conversion is accounted for and that local gene-conversion rates reflect recombination rates. We detected a strong enrichment of recent deleterious variation among mismatching variants found within IBD regions and observed summary statistics of local sharing of IBD segments to closely match previously proposed metrics of background selection; however, we found no significant effects of selection on our mutation-rate estimates. We detected no evidence of strong variation of mutation rates in a number of genomic annotations obtained from several recent studies. Our analysis suggests that a mutation-rate estimate higher than that reported by recent pedigree-based studies should be adopted in the context of DNA-based demographic reconstruction. PMID:26581902

  8. Leveraging Distant Relatedness to Quantify Human Mutation and Gene-Conversion Rates.

    PubMed

    Palamara, Pier Francesco; Francioli, Laurent C; Wilton, Peter R; Genovese, Giulio; Gusev, Alexander; Finucane, Hilary K; Sankararaman, Sriram; Sunyaev, Shamil R; de Bakker, Paul I W; Wakeley, John; Pe'er, Itsik; Price, Alkes L

    2015-12-01

    The rate at which human genomes mutate is a central biological parameter that has many implications for our ability to understand demographic and evolutionary phenomena. We present a method for inferring mutation and gene-conversion rates by using the number of sequence differences observed in identical-by-descent (IBD) segments together with a reconstructed model of recent population-size history. This approach is robust to, and can quantify, the presence of substantial genotyping error, as validated in coalescent simulations. We applied the method to 498 trio-phased sequenced Dutch individuals and inferred a point mutation rate of 1.66 × 10(-8) per base per generation and a rate of 1.26 × 10(-9) for <20 bp indels. By quantifying how estimates varied as a function of allele frequency, we inferred the probability that a site is involved in non-crossover gene conversion as 5.99 × 10(-6). We found that recombination does not have observable mutagenic effects after gene conversion is accounted for and that local gene-conversion rates reflect recombination rates. We detected a strong enrichment of recent deleterious variation among mismatching variants found within IBD regions and observed summary statistics of local sharing of IBD segments to closely match previously proposed metrics of background selection; however, we found no significant effects of selection on our mutation-rate estimates. We detected no evidence of strong variation of mutation rates in a number of genomic annotations obtained from several recent studies. Our analysis suggests that a mutation-rate estimate higher than that reported by recent pedigree-based studies should be adopted in the context of DNA-based demographic reconstruction.

  9. Gene expression analysis of flax seed development

    PubMed Central

    2011-01-01

    Background Flax, Linum usitatissimum L., is an important crop whose seed oil and stem fiber have multiple industrial applications. Flax seeds are also well-known for their nutritional attributes, viz., omega-3 fatty acids in the oil and lignans and mucilage from the seed coat. In spite of the importance of this crop, there are few molecular resources that can be utilized toward improving seed traits. Here, we describe flax embryo and seed development and generation of comprehensive genomic resources for the flax seed. Results We describe a large-scale generation and analysis of expressed sequences in various tissues. Collectively, the 13 libraries we have used provide a broad representation of genes active in developing embryos (globular, heart, torpedo, cotyledon and mature stages) seed coats (globular and torpedo stages) and endosperm (pooled globular to torpedo stages) and genes expressed in flowers, etiolated seedlings, leaves, and stem tissue. A total of 261,272 expressed sequence tags (EST) (GenBank accessions LIBEST_026995 to LIBEST_027011) were generated. These EST libraries included transcription factor genes that are typically expressed at low levels, indicating that the depth is adequate for in silico expression analysis. Assembly of the ESTs resulted in 30,640 unigenes and 82% of these could be identified on the basis of homology to known and hypothetical genes from other plants. When compared with fully sequenced plant genomes, the flax unigenes resembled poplar and castor bean more than grape, sorghum, rice or Arabidopsis. Nearly one-fifth of these (5,152) had no homologs in sequences reported for any organism, suggesting that this category represents genes that are likely unique to flax. Digital analyses revealed gene expression dynamics for the biosynthesis of a number of important seed constituents during seed development. Conclusions We have developed a foundational database of expressed sequences and collection of plasmid clones that comprise

  10. Inducible gene expression systems for plants.

    PubMed

    Borghi, Lorenzo

    2010-01-01

    Several systems for induction of transgene expression in plants have been described recently. Inducible systems were used mainly in tobacco, rice, Arabidopsis, tomato, and maize. Inducible systems offer researchers the possibility to deregulate gene expression levels at particular stages of plant development and in particular tissues of interest. The more precise temporal and spatial control, obtained by providing the transgenic plant with the appropriate chemical compound or treatment, permits to analyze also the function of those genes required for plant viability. In addition, inducible systems allow promoting local changes in gene expression levels without causing gross alterations to the whole plant development. Here, protocols will be presented to work with five different inducible systems: AlcR/AlcA (ethanol inducible); GR fusions, GVG, and pOp/LhGR (dexamethasone inducible); XVE/OlexA (beta-estradiol inducible); and heat shock induction. PMID:20734254

  11. Heritability and genomics of gene expression in peripheral blood.

    PubMed

    Wright, Fred A; Sullivan, Patrick F; Brooks, Andrew I; Zou, Fei; Sun, Wei; Xia, Kai; Madar, Vered; Jansen, Rick; Chung, Wonil; Zhou, Yi-Hui; Abdellaoui, Abdel; Batista, Sandra; Butler, Casey; Chen, Guanhua; Chen, Ting-Huei; D'Ambrosio, David; Gallins, Paul; Ha, Min Jin; Hottenga, Jouke Jan; Huang, Shunping; Kattenberg, Mathijs; Kochar, Jaspreet; Middeldorp, Christel M; Qu, Ani; Shabalin, Andrey; Tischfield, Jay; Todd, Laura; Tzeng, Jung-Ying; van Grootheest, Gerard; Vink, Jacqueline M; Wang, Qi; Wang, Wei; Wang, Weibo; Willemsen, Gonneke; Smit, Johannes H; de Geus, Eco J; Yin, Zhaoyu; Penninx, Brenda W J H; Boomsma, Dorret I

    2014-05-01

    We assessed gene expression profiles in 2,752 twins, using a classic twin design to quantify expression heritability and quantitative trait loci (eQTLs) in peripheral blood. The most highly heritable genes (∼777) were grouped into distinct expression clusters, enriched in gene-poor regions, associated with specific gene function or ontology classes, and strongly associated with disease designation. The design enabled a comparison of twin-based heritability to estimates based on dizygotic identity-by-descent sharing and distant genetic relatedness. Consideration of sampling variation suggests that previous heritability estimates have been upwardly biased. Genotyping of 2,494 twins enabled powerful identification of eQTLs, which we further examined in a replication set of 1,895 unrelated subjects. A large number of non-redundant local eQTLs (6,756) met replication criteria, whereas a relatively small number of distant eQTLs (165) met quality control and replication standards. Our results provide a new resource toward understanding the genetic control of transcription.

  12. The Interrelationship between Promoter Strength, Gene Expression, and Growth Rate

    PubMed Central

    Klesmith, Justin R.; Detwiler, Emily E.; Tomek, Kyle J.; Whitehead, Timothy A.

    2014-01-01

    In exponentially growing bacteria, expression of heterologous protein impedes cellular growth rates. Quantitative understanding of the relationship between expression and growth rate will advance our ability to forward engineer bacteria, important for metabolic engineering and synthetic biology applications. Recently, a work described a scaling model based on optimal allocation of ribosomes for protein translation. This model quantitatively predicts a linear relationship between microbial growth rate and heterologous protein expression with no free parameters. With the aim of validating this model, we have rigorously quantified the fitness cost of gene expression by using a library of synthetic constitutive promoters to drive expression of two separate proteins (eGFP and amiE) in E. coli in different strains and growth media. In all cases, we demonstrate that the fitness cost is consistent with the previous findings. We expand upon the previous theory by introducing a simple promoter activity model to quantitatively predict how basal promoter strength relates to growth rate and protein expression. We then estimate the amount of protein expression needed to support high flux through a heterologous metabolic pathway and predict the sizable fitness cost associated with enzyme production. This work has broad implications across applied biological sciences because it allows for prediction of the interplay between promoter strength, protein expression, and the resulting cost to microbial growth rates. PMID:25286161

  13. The interrelationship between promoter strength, gene expression, and growth rate.

    PubMed

    Bienick, Matthew S; Young, Katherine W; Klesmith, Justin R; Detwiler, Emily E; Tomek, Kyle J; Whitehead, Timothy A

    2014-01-01

    In exponentially growing bacteria, expression of heterologous protein impedes cellular growth rates. Quantitative understanding of the relationship between expression and growth rate will advance our ability to forward engineer bacteria, important for metabolic engineering and synthetic biology applications. Recently, a work described a scaling model based on optimal allocation of ribosomes for protein translation. This model quantitatively predicts a linear relationship between microbial growth rate and heterologous protein expression with no free parameters. With the aim of validating this model, we have rigorously quantified the fitness cost of gene expression by using a library of synthetic constitutive promoters to drive expression of two separate proteins (eGFP and amiE) in E. coli in different strains and growth media. In all cases, we demonstrate that the fitness cost is consistent with the previous findings. We expand upon the previous theory by introducing a simple promoter activity model to quantitatively predict how basal promoter strength relates to growth rate and protein expression. We then estimate the amount of protein expression needed to support high flux through a heterologous metabolic pathway and predict the sizable fitness cost associated with enzyme production. This work has broad implications across applied biological sciences because it allows for prediction of the interplay between promoter strength, protein expression, and the resulting cost to microbial growth rates.

  14. Gene expression profiles in irradiated cancer cells

    NASA Astrophysics Data System (ADS)

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-01

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  15. Gene expression profiles in irradiated cancer cells

    SciTech Connect

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-26

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  16. Comparative gene expression profiling by oligonucleotide fingerprinting.

    PubMed Central

    Meier-Ewert, S; Lange, J; Gerst, H; Herwig, R; Schmitt, A; Freund, J; Elge, T; Mott, R; Herrmann, B; Lehrach, H

    1998-01-01

    The use of hybridisation of synthetic oligonucleotides to cDNAs under high stringency to characterise gene sequences has been demonstrated by a number of groups. We have used two cDNA libraries of 9 and 12 day mouse embryos (24 133 and 34 783 clones respectively) in a pilot study to characterise expressed genes by hybridisation with 110 hybridisation probes. We have identified 33 369 clusters of cDNA clones, that ranged in representation from 1 to 487 copies (0.7%). 737 were assigned to known rodent genes, and a further 13 845 showed significant homologies. A total of 404 clusters were identified as significantly differentially represented (P < 0.01) between the two cDNA libraries. This study demonstrates the utility of the fingerprinting approach for the generation of comparative gene expression profiles through the analysis of cDNAs derived from different biological materials. PMID:9547283

  17. Duplicate gene enrichment and expression pattern diversification in multicellularity

    PubMed Central

    Padawer, Timothy; Leighty, Ralph E.; Wang, Degeng

    2012-01-01

    The enrichment of duplicate genes, and therefore paralogs (proteins coded by duplicate genes), in multicellular versus unicellular organisms enhances genomic functional innovation. This study quantitatively examined relationships among paralog enrichment, expression pattern diversification and multicellularity, aiming to better understand genomic basis of multicellularity. Paralog abundance in specific cells was compared with those in unicellular proteomes and the whole proteomes of multicellular organisms. The budding yeast, Saccharomyces cerevisiae and the nematode, Caenorhabditis elegans, for which the gene sets expressed in specific cells are available, were used as uni and multicellular models, respectively. Paralog count (K) distributions [P(k)] follow a power-law relationship [P(k) ∝ k−α] in the whole proteomes of both species and in specific C. elegans cells. The value of the constant α can be used as a gauge of paralog abundance; the higher the value, the lower the paralog abundance. The α-value is indeed lower in the whole proteome of C. elegans (1.74) than in S. cerevisiae (2.34), quantifying the enrichment of paralogs in multicellular species. We also found that the power-law relationship applies to the proteomes of specific C. elegans cells. Strikingly, values of α in specific cells are higher and comparable to that in S. cerevisiae. Thus, paralog abundance in specific cells is lower and comparable to that in unicellular species. Furthermore, how much the expression level of a gene fluctuates across different C. elegans cells correlates positively with its paralog count, which is further confirmed by human gene-expression patterns across different tissues. Taken together, these results quantitatively and mechanistically establish enrichment of paralogs with diversifying expression patterns as genomic and evolutionary basis of multicellularity. PMID:22645319

  18. Visualizing Gene Expression In Situ

    SciTech Connect

    Burlage, R.S.

    1998-11-02

    Visualizing bacterial cells and describing their responses to the environment are difficult tasks. Their small size is the chief reason for the difficulty, which means that we must often use many millions of cells in a sample in order to determine what the average response of the bacteria is. However, an average response can sometimes mask important events in bacterial physiology, which means that our understanding of these organisms will suffer. We have used a variety of instruments to visualize bacterial cells, all of which tell us something different about the sample. We use a fluorescence activated cell sorter to sort cells based on the fluorescence provided by bioreporter genes, and these can be used to select for particular genetic mutations. Cells can be visualized by epifluorescent microscopy, and sensitive photodetectors can be added that allow us to find a single bacterial cell that is fluorescent or bioluminescent. We have also used standard photomultipliers to examine cell aggregates as field bioreporter microorganisms. Examples of each of these instruments show how our understanding of bacterial physiology has changed with the technology.

  19. Gene expression profile of Clonorchis sinensis metacercariae.

    PubMed

    Cho, Pyo Yun; Kim, Tae Im; Whang, Seong Man; Hong, Sung-Jong

    2008-01-01

    Clonorchis sinensis develop through miracidium, sporocyst, redia, cercaria, and metacercaria stages before becoming egg-laying adult flukes. The authors undertook this analysis of gene expression profiles during developmental stages to increase our understanding of the biology of C. sinensis and of host-parasite relationships. From a C. sinensis metacercariae complementary deoxyribonucleic acid library, 419 expressed sequence tags (ESTs) of average length of 668 bp were collected and assembled into 322 genes containing 70 clusters and 252 singletons. The genes were annotated using BLAST searches and categorized into ten major functional categories. Genes expressed abundantly were those of proteases and metabolic, transcription, and translation housekeeping proteins. Genes expressed higher in C. sinensis metacercariae than in adults coded structural and cytoskeletal proteins, transcription and translation machinery proteins, and energy metabolism-related proteins. This EST information supports the notion that C. sinensis metacercariae in fish hosts have a physiology and metabolism that is quite different from that of its adult form in mammals. PMID:17924144

  20. Optogenetics for gene expression in mammalian cells.

    PubMed

    Müller, Konrad; Naumann, Sebastian; Weber, Wilfried; Zurbriggen, Matias D

    2015-02-01

    Molecular switches that are controlled by chemicals have evolved as central research instruments in mammalian cell biology. However, these tools are limited in terms of their spatiotemporal resolution due to freely diffusing inducers. These limitations have recently been addressed by the development of optogenetic, genetically encoded, and light-responsive tools that can be controlled with the unprecedented spatiotemporal precision of light. In this article, we first provide a brief overview of currently available optogenetic tools that have been designed to control diverse cellular processes. Then, we focus on recent developments in light-controlled gene expression technologies and provide the reader with a guideline for choosing the most suitable gene expression system.

  1. Clustering of High Throughput Gene Expression Data

    PubMed Central

    Pirim, Harun; Ekşioğlu, Burak; Perkins, Andy; Yüceer, Çetin

    2012-01-01

    High throughput biological data need to be processed, analyzed, and interpreted to address problems in life sciences. Bioinformatics, computational biology, and systems biology deal with biological problems using computational methods. Clustering is one of the methods used to gain insight into biological processes, particularly at the genomics level. Clearly, clustering can be used in many areas of biological data analysis. However, this paper presents a review of the current clustering algorithms designed especially for analyzing gene expression data. It is also intended to introduce one of the main problems in bioinformatics - clustering gene expression data - to the operations research community. PMID:23144527

  2. Genes Expressed in Human Tumor Endothelium

    NASA Astrophysics Data System (ADS)

    St. Croix, Brad; Rago, Carlo; Velculescu, Victor; Traverso, Giovanni; Romans, Katharine E.; Montgomery, Elizabeth; Lal, Anita; Riggins, Gregory J.; Lengauer, Christoph; Vogelstein, Bert; Kinzler, Kenneth W.

    2000-08-01

    To gain a molecular understanding of tumor angiogenesis, we compared gene expression patterns of endothelial cells derived from blood vessels of normal and malignant colorectal tissues. Of over 170 transcripts predominantly expressed in the endothelium, 79 were differentially expressed, including 46 that were specifically elevated in tumor-associated endothelium. Several of these genes encode extracellular matrix proteins, but most are of unknown function. Most of these tumor endothelial markers were expressed in a wide range of tumor types, as well as in normal vessels associated with wound healing and corpus luteum formation. These studies demonstrate that tumor and normal endothelium are distinct at the molecular level, a finding that may have significant implications for the development of anti-angiogenic therapies.

  3. Chromatin modifications remodel cardiac gene expression.

    PubMed

    Mathiyalagan, Prabhu; Keating, Samuel T; Du, Xiao-Jun; El-Osta, Assam

    2014-07-01

    Signalling and transcriptional control involve precise programmes of gene activation and suppression necessary for cardiovascular physiology. Deep sequencing of DNA-bound transcription factors reveals a remarkable complexity of co-activators or co-repressors that serve to alter chromatin modification and regulate gene expression. The regulated complexes characterized by genome-wide mapping implicate the recruitment and exchange of proteins with specific enzymatic activities that include roles for histone acetylation and methylation in key developmental programmes of the heart. As for transcriptional changes in response to pathological stress, co-regulatory complexes are also differentially utilized to regulate genes in cardiac disease. Members of the histone deacetylase (HDAC) family catalyse the removal of acetyl groups from proteins whose pharmacological inhibition has profound effects preventing heart failure. HDACs interact with a complex co-regulatory network of transcription factors, chromatin-remodelling complexes, and specific histone modifiers to regulate gene expression in the heart. For example, the histone methyltransferase (HMT), enhancer of zeste homolog 2 (Ezh2), is regulated by HDAC inhibition and associated with pathological cardiac hypertrophy. The challenge now is to target the activity of enzymes involved in protein modification to prevent or reverse the expression of genes implicated with cardiac hypertrophy. In this review, we discuss the role of HDACs and HMTs with a focus on chromatin modification and gene function as well as the clinical treatment of heart failure. PMID:24812277

  4. Sequence and gene expression evolution of paralogous genes in willows.

    PubMed

    Harikrishnan, Srilakshmy L; Pucholt, Pascal; Berlin, Sofia

    2015-12-22

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows.

  5. Sequence and gene expression evolution of paralogous genes in willows

    PubMed Central

    Harikrishnan, Srilakshmy L.; Pucholt, Pascal; Berlin, Sofia

    2015-01-01

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows. PMID:26689951

  6. Gene expression profiling analysis of ovarian cancer

    PubMed Central

    YIN, JI-GANG; LIU, XIAN-YING; WANG, BIN; WANG, DAN-YANG; WEI, MAN; FANG, HUA; XIANG, MEI

    2016-01-01

    As a gynecological oncology, ovarian cancer has high incidence and mortality. To study the mechanisms of ovarian cancer, the present study analyzed the GSE37582 microarray. GSE37582 was downloaded from Gene Expression Omnibus and included data from 74 ovarian cancer cases and 47 healthy controls. The differentially-expressed genes (DEGs) were screened using linear models for microarray data package in R and were further screened for functional annotation. Next, Gene Ontology and pathway enrichment analysis of the DEGs was conducted. The interaction associations of the proteins encoded by the DEGs were searched using the Search Tool for the Retrieval of Interacting Genes, and the protein-protein interaction (PPI) network was visualized by Cytoscape. Moreover, module analysis of the PPI network was performed using the BioNet analysis tool in R. A total of 284 DEGs were screened, consisting of 145 upregulated genes and 139 downregulated genes. In particular, downregulated FBJ murine osteosarcoma viral oncogene homolog (FOS) was an oncogene, while downregulated cyclin-dependent kinase inhibitor 1A (CDKN1A) was a tumor suppressor gene and upregulated cluster of differentiation 44 (CD44) was classed as an ‘other’ gene. The enriched functions included collagen catabolic process, stress-activated mitogen-activated protein kinases cascade and insulin receptor signaling pathway. Meanwhile, FOS (degree, 15), CD44 (degree, 9), B-cell CLL/lymphoma 2 (BCL2; degree, 7), CDKN1A (degree, 7) and matrix metallopeptidase 3 (MMP3; degree, 6) had higher connectivity degrees in the PPI network for the DEGs. These genes may be involved in ovarian cancer by interacting with other genes in the module of the PPI network (e.g., BCL2-FOS, BCL2-CDKN1A, FOS-CDKN1A, FOS-CD44, MMP3-MMP7 and MMP7-CD44). Overall, BCL2, FOS, CDKN1A, CD44, MMP3 and MMP7 may be correlated with ovarian cancer. PMID:27347159

  7. Annotation of gene function in citrus using gene expression information and co-expression networks

    PubMed Central

    2014-01-01

    Background The genus Citrus encompasses major cultivated plants such as sweet orange, mandarin, lemon and grapefruit, among the world’s most economically important fruit crops. With increasing volumes of transcriptomics data available for these species, Gene Co-expression Network (GCN) analysis is a viable option for predicting gene function at a genome-wide scale. GCN analysis is based on a “guilt-by-association” principle whereby genes encoding proteins involved in similar and/or related biological processes may exhibit similar expression patterns across diverse sets of experimental conditions. While bioinformatics resources such as GCN analysis are widely available for efficient gene function prediction in model plant species including Arabidopsis, soybean and rice, in citrus these tools are not yet developed. Results We have constructed a comprehensive GCN for citrus inferred from 297 publicly available Affymetrix Genechip Citrus Genome microarray datasets, providing gene co-expression relationships at a genome-wide scale (33,000 transcripts). The comprehensive citrus GCN consists of a global GCN (condition-independent) and four condition-dependent GCNs that survey the sweet orange species only, all citrus fruit tissues, all citrus leaf tissues, or stress-exposed plants. All of these GCNs are clustered using genome-wide, gene-centric (guide) and graph clustering algorithms for flexibility of gene function prediction. For each putative cluster, gene ontology (GO) enrichment and gene expression specificity analyses were performed to enhance gene function, expression and regulation pattern prediction. The guide-gene approach was used to infer novel roles of genes involved in disease susceptibility and vitamin C metabolism, and graph-clustering approaches were used to investigate isoprenoid/phenylpropanoid metabolism in citrus peel, and citric acid catabolism via the GABA shunt in citrus fruit. Conclusions Integration of citrus gene co-expression networks

  8. Gene expression profiling of human erythroid progenitors by micro-serial analysis of gene expression.

    PubMed

    Fujishima, Naohito; Hirokawa, Makoto; Aiba, Namiko; Ichikawa, Yoshikazu; Fujishima, Masumi; Komatsuda, Atsushi; Suzuki, Yoshiko; Kawabata, Yoshinari; Miura, Ikuo; Sawada, Ken-ichi

    2004-10-01

    We compared the expression profiles of highly purified human CD34+ cells and erythroid progenitor cells by micro-serial analysis of gene expression (microSAGE). Human CD34+ cells were purified from granulocyte colony-stimulating factor-mobilized blood stem cells, and erythroid progenitors were obtained by cultivating these cells in the presence of stem cell factor, interleukin 3, and erythropoietin. Our 10,202 SAGE tags allowed us to identify 1354 different transcripts appearing more than once. Erythroid progenitor cells showed increased expression of LRBA, EEF1A1, HSPCA, PILRB, RANBP1, NACA, and SMURF. Overexpression of HSPCA was confirmed by real-time polymerase chain reaction analysis. MicroSAGE revealed an unexpected preferential expression of several genes in erythroid progenitor cells in addition to the known functional genes, including hemoglobins. Our results provide reference data for future studies of gene expression in various hematopoietic disorders, including myelodysplastic syndrome and leukemia.

  9. The low noise limit in gene expression

    SciTech Connect

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.; Razooky, Brandon S.

    2015-10-21

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiency can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. Lastly, these results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.

  10. The low noise limit in gene expression

    DOE PAGESBeta

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.; Razooky, Brandon S.

    2015-10-21

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiencymore » can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. Lastly, these results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.« less

  11. Trigger finger, tendinosis, and intratendinous gene expression.

    PubMed

    Lundin, A-C; Aspenberg, P; Eliasson, P

    2014-04-01

    The pathogenesis of trigger finger has generally been ascribed to primary changes in the first annular ligament. In contrast, we recently found histological changes in the tendons, similar to the findings in Achilles tendinosis or tendinopathy. We therefore hypothesized that trigger finger tendons would show differences in gene expression in comparison to normal tendons in a pattern similar to what is published for Achilles tendinosis. We performed quantitative real-time polymerase chain reaction on biopsies from finger flexor tendons, 13 trigger fingers and 13 apparently healthy control tendons, to assess the expression of 10 genes which have been described to be differently expressed in tendinosis (collagen type 1a1, collagen 3a1, MMP-2, MMP-3, ADAMTS-5, TIMP-3, aggrecan, biglycan, decorin, and versican). In trigger finger tendons, collagen types 1a1 and 3a1, aggrecan and biglycan were all up-regulated, and MMP-3and TIMP-3 were down-regulated. These changes were statistically significant and have been previously described for Achilles tendinosis. The remaining four genes were not significantly altered. The changes in gene expression support the hypothesis that trigger finger is a form of tendinosis. Because trigger finger is a common condition, often treated surgically, it could provide opportunities for clinical research on tendinosis. PMID:22882155

  12. Multiple Stochastic Point Processes in Gene Expression

    NASA Astrophysics Data System (ADS)

    Murugan, Rajamanickam

    2008-04-01

    We generalize the idea of multiple-stochasticity in chemical reaction systems to gene expression. Using Chemical Langevin Equation approach we investigate how this multiple-stochasticity can influence the overall molecular number fluctuations. We show that the main sources of this multiple-stochasticity in gene expression could be the randomness in transcription and translation initiation times which in turn originates from the underlying bio-macromolecular recognition processes such as the site-specific DNA-protein interactions and therefore can be internally regulated by the supra-molecular structural factors such as the condensation/super-coiling of DNA. Our theory predicts that (1) in case of gene expression system, the variances ( φ) introduced by the randomness in transcription and translation initiation-times approximately scales with the degree of condensation ( s) of DNA or mRNA as φ ∝ s -6. From the theoretical analysis of the Fano factor as well as coefficient of variation associated with the protein number fluctuations we predict that (2) unlike the singly-stochastic case where the Fano factor has been shown to be a monotonous function of translation rate, in case of multiple-stochastic gene expression the Fano factor is a turn over function with a definite minimum. This in turn suggests that the multiple-stochastic processes can also be well tuned to behave like a singly-stochastic point processes by adjusting the rate parameters.

  13. Population-level control of gene expression

    NASA Astrophysics Data System (ADS)

    Nevozhay, Dmitry; Adams, Rhys; van Itallie, Elizabeth; Bennett, Matthew; Balazsi, Gabor

    2011-03-01

    Gene expression is the process that translates genetic information into proteins, that determine the way cells live, function and even die. It was demonstrated that cells with identical genomes exposed to the same environment can differ in their protein composition and therefore phenotypes. Protein levels can vary between cells due to the stochastic nature of intracellular biochemical events, indicating that the genotype-phenotype connection is not deterministic at the cellular level. We asked whether genomes could encode isogenic cell populations more reliably than single cells. To address this question, we built two gene circuits to control three cell population-level characteristics: gene expression mean, coefficient of variation and non-genetic memory of previous expression states. Indeed, we found that these population-level characteristics were more predictable than the gene expression of single cells in a well-controlled environment. This research was supported by the NIH Director's New Innovator Award 1DP2 OD006481-01 and Welch Foundation Grant C-1729.

  14. Expression of mouse metallothionein genes in tobacco

    SciTech Connect

    Maiti, I.B.; Yeargan, R.; Wagner, G.J.; Hunt, A.G. )

    1990-05-01

    We have expressed a mouse metallothionein (NT) gene in tobacco under control of the cauliflower mosaic virus (CaMV) 35S promoter and a pea ribulose-1,5-bisphosphate carboxylase small subunit (rbcS) gene promoter. Seedlings in which MT gene expression is driven by the 35S promoter are resistant to toxic levels of cadmium. Mature plants carrying the 35S-MT gene accumulate less Cd in their leaves when exposed to low levels of Cd in laboratory growth conditions. Plants with the rbcS-MT construction express this gene in a light-regulated and tissue-specific manner, as expected. Moreover, the MT levels in leaves in these plants are about 20% of those seen in 35S-MT plants. These plants are currently being tested for Cd resistance. In addition, a small field evaluation of 35S-MT lines for Cd levels is being evaluated. These experiments will address the possibility of using MTs to alter Cd levels in crop species.

  15. Functionalization of a protosynaptic gene expression network

    PubMed Central

    Conaco, Cecilia; Bassett, Danielle S.; Zhou, Hongjun; Arcila, Mary Luz; Degnan, Sandie M.; Degnan, Bernard M.; Kosik, Kenneth S.

    2012-01-01

    Assembly of a functioning neuronal synapse requires the precisely coordinated synthesis of many proteins. To understand the evolution of this complex cellular machine, we tracked the developmental expression patterns of a core set of conserved synaptic genes across a representative sampling of the animal kingdom. Coregulation, as measured by correlation of gene expression over development, showed a marked increase as functional nervous systems emerged. In the earliest branching animal phyla (Porifera), in which a nearly complete set of synaptic genes exists in the absence of morphological synapses, these “protosynaptic” genes displayed a lack of global coregulation although small modules of coexpressed genes are readily detectable by using network analysis techniques. These findings suggest that functional synapses evolved by exapting preexisting cellular machines, likely through some modification of regulatory circuitry. Evolutionarily ancient modules continue to operate seamlessly within the synapses of modern animals. This work shows that the application of network techniques to emerging genomic and expression data can provide insights into the evolution of complex cellular machines such as the synapse. PMID:22723359

  16. Coordination of plastid and nuclear gene expression.

    PubMed Central

    Gray, John C; Sullivan, James A; Wang, Jun-Hui; Jerome, Cheryl A; MacLean, Daniel

    2003-01-01

    The coordinated expression of genes distributed between the nuclear and plastid genomes is essential for the assembly of functional chloroplasts. Although the nucleus has a pre-eminent role in controlling chloroplast biogenesis, there is considerable evidence that the expression of nuclear genes encoding photosynthesis-related proteins is regulated by signals from plastids. Perturbation of several plastid-located processes, by inhibitors or in mutants, leads to decreased transcription of a set of nuclear photosynthesis-related genes. Characterization of arabidopsis gun (genomes uncoupled) mutants, which express nuclear genes in the presence of norflurazon or lincomycin, has provided evidence for two separate signalling pathways, one involving tetrapyrrole biosynthesis intermediates and the other requiring plastid protein synthesis. In addition, perturbation of photosynthetic electron transfer produces at least two different redox signals, as part of the acclimation to altered light conditions. The recognition of multiple plastid signals requires a reconsideration of the mechanisms of regulation of transcription of nuclear genes encoding photosynthesis-related proteins. PMID:12594922

  17. Gene Expression Variability Underlies Adaptive Resistance in Phenotypically Heterogeneous Bacterial Populations.

    PubMed

    Erickson, Keesha E; Otoupal, Peter B; Chatterjee, Anushree

    2015-11-13

    The root cause of the antibiotic resistance crisis is the ability of bacteria to evolve resistance to a multitude of antibiotics and other environmental toxins. The regulation of adaptation is difficult to pinpoint due to extensive phenotypic heterogeneity arising during evolution. Here, we investigate the mechanisms underlying general bacterial adaptation by evolving wild-type Escherichia coli populations to dissimilar chemical toxins. We demonstrate the presence of extensive inter- and intrapopulation phenotypic heterogeneity across adapted populations in multiple traits, including minimum inhibitory concentration, growth rate, and lag time. To search for a common response across the heterogeneous adapted populations, we measured gene expression in three stress-response networks: the mar regulon, the general stress response, and the SOS response. While few genes were differentially expressed, clustering revealed that interpopulation gene expression variability in adapted populations was distinct from that of unadapted populations. Notably, we observed both increases and decreases in gene expression variability upon adaptation. Sequencing select genes revealed that the observed gene expression trends are not necessarily attributable to genetic changes. To further explore the connection between gene expression variability and adaptation, we propagated single-gene knockout and CRISPR (clustered regularly interspaced short palindromic repeats) interference strains and quantified impact on adaptation to antibiotics. We identified significant correlations that suggest genes with low expression variability have greater impact on adaptation. This study provides evidence that gene expression variability can be used as an indicator of bacterial adaptive resistance, even in the face of the pervasive phenotypic heterogeneity underlying adaptation. PMID:27623410

  18. Gene Expression Commons: An Open Platform for Absolute Gene Expression Profiling

    PubMed Central

    Seita, Jun; Sahoo, Debashis; Rossi, Derrick J.; Bhattacharya, Deepta; Serwold, Thomas; Inlay, Matthew A.; Ehrlich, Lauren I. R.; Fathman, John W.; Dill, David L.; Weissman, Irving L.

    2012-01-01

    Gene expression profiling using microarrays has been limited to comparisons of gene expression between small numbers of samples within individual experiments. However, the unknown and variable sensitivities of each probeset have rendered the absolute expression of any given gene nearly impossible to estimate. We have overcome this limitation by using a very large number (>10,000) of varied microarray data as a common reference, so that statistical attributes of each probeset, such as the dynamic range and threshold between low and high expression, can be reliably discovered through meta-analysis. This strategy is implemented in a web-based platform named “Gene Expression Commons” (https://gexc.stanford.edu/) which contains data of 39 distinct highly purified mouse hematopoietic stem/progenitor/differentiated cell populations covering almost the entire hematopoietic system. Since the Gene Expression Commons is designed as an open platform, investigators can explore the expression level of any gene, search by expression patterns of interest, submit their own microarray data, and design their own working models representing biological relationship among samples. PMID:22815738

  19. Validation of housekeeping genes as an internal control for gene expression studies in Giardia lamblia using quantitative real-time PCR.

    PubMed

    Marcial-Quino, Jaime; Fierro, Francisco; De la Mora-De la Mora, Ignacio; Enríquez-Flores, Sergio; Gómez-Manzo, Saúl; Vanoye-Carlo, America; Garcia-Torres, Itzhel; Sierra-Palacios, Edgar; Reyes-Vivas, Horacio

    2016-04-25

    The analysis of transcript levels of specific genes is important for understanding transcriptional regulation and for the characterization of gene function. Real-time quantitative reverse transcriptase PCR (RT-qPCR) has become a powerful tool to quantify gene expression. The objective of this study was to identify reliable housekeeping genes in Giardia lamblia. Twelve genes were selected for this purpose, and their expression was analyzed in the wild type WB strain and in two strains with resistance to nitazoxanide (NTZ) and metronidazole (MTZ), respectively. RefFinder software analysis showed that the expression of the genes is different in the three strains. The integrated data from the four analyses showed that the NADH oxidase (NADH) and aldolase (ALD) genes were the most steadily expressed genes, whereas the glyceraldehyde-3-phosphate dehydrogenase gene was the most unstable. Additionally, the relative expression of seven genes were quantified in the NTZ- and MTZ-resistant strains by RT-qPCR, using the aldolase gene as the internal control, and the results showed a consistent differential pattern of expression in both strains. The housekeeping genes found in this work will facilitate the analysis of mRNA expression levels of other genes of interest in G. lamblia.

  20. Selection of reliable reference genes for gene expression studies in Clonostachys rosea 67-1 under sclerotial induction.

    PubMed

    Sun, Zhan-Bin; Li, Shi-Dong; Sun, Man-Hong

    2015-07-01

    Reference genes are important to precisely quantify gene expression by real-time PCR. In order to identify stable and reliable expressed genes in mycoparasite Clonostachys rosea in different modes of nutrition, seven commonly used housekeeping genes, 18S rRNA, actin, β-tubulin, elongation factor 1, ubiquitin, ubiquitin-conjugating enzyme and glyceraldehyde-3-phosphate dehydrogenase, from the effective biocontrol isolate C. rosea 67-1 were tested for their expression under sclerotial induction and during vegetative growth on PDA medium. Analysis by three software programs showed that differences existed among the candidates. Elongation factor 1 was most stable; the M value in geNorm, SD value in Bestkeeper and stability value in Normfinder analysis were 0.405, 0.450 and 0.442, respectively, indicating that the gene elongation factor 1 could be used to normalize gene expression in C. rosea in both vegetative growth and parasitic process. By using elongation factor 1, the expression of a serine protease gene, sep, in different conditions was assessed, which was consistent with the transcriptomic data. This research provides an effective method to quantitate expression changes of target genes in C. rosea, and will assist in further investigation of parasitism-related genes of this fungus.

  1. Fluid Mechanics, Arterial Disease, and Gene Expression

    NASA Astrophysics Data System (ADS)

    Tarbell, John M.; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow-induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid mechanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs.

  2. Fluid Mechanics, Arterial Disease, and Gene Expression

    PubMed Central

    Tarbell, John M.; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow–induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs. PMID:25360054

  3. Fluid Mechanics, Arterial Disease, and Gene Expression.

    PubMed

    Tarbell, John M; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow-induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs.

  4. Control mechanisms of plastid gene expression

    SciTech Connect

    Gruissem, W.; Tonkyn, J.C.

    1993-12-31

    Plastid DNAs of higher plants contain approximately 150 genes that encode RNAs and proteins for genetic and photosynthetic functions of the organelle. Results published in the last few years illustrate that the spatial and temporal expression of these plastid genes is regulated, in part, at the transcriptional level, but that developmentally controlled changes in mRNA stability, translational activity, and protein phosphorylation also have an important role in the control of plastid functions. This comprehensive review summarizes and discusses the mechanisms by which regulation of gene expression is exerted at the transcriptional and post-transcriptional levels. It provides an overview of our current knowledge, but also emphasizes areas that are controversial and in which information on regulatory mechanisms is still incomplete. 455 refs., 3 figs., 3 tabs.

  5. Repression of gene expression by oxidative stress.

    PubMed Central

    Morel, Y; Barouki, R

    1999-01-01

    Gene expression is modulated by both physiological signals (hormones, cytokines, etc.) and environmental stimuli (physical parameters, xenobiotics, etc.). Oxidative stress appears to be a key pleiotropic modulator which may be involved in either pathway. Indeed, reactive oxygen species (ROS) have been described as second messengers for several growth factors and cytokines, but have also been shown to rise following cellular insults such as xenobiotic metabolism or enzymic deficiency. Extensive studies on the induction of stress-response genes by oxidative stress have been reported. In contrast, owing to the historical focus on gene induction, less attention has been paid to gene repression by ROS. However, a growing number of studies have shown that moderate (i.e. non-cytotoxic) oxidative stress specifically down-regulates the expression of various genes. In this review, we describe the alteration of several physiological functions resulting from oxidative-stress-mediated inhibition of gene transcription. We will then focus on the repressive oxidative modulation of various transcription factors elicited by ROS. PMID:10477257

  6. Using interpolation to estimate system uncertainty in gene expression experiments.

    PubMed

    Falin, Lee J; Tyler, Brett M

    2011-01-01

    The widespread use of high-throughput experimental assays designed to measure the entire complement of a cell's genes or gene products has led to vast stores of data that are extremely plentiful in terms of the number of items they can measure in a single sample, yet often sparse in the number of samples per experiment due to their high cost. This often leads to datasets where the number of treatment levels or time points sampled is limited, or where there are very small numbers of technical and/or biological replicates. Here we introduce a novel algorithm to quantify the uncertainty in the unmeasured intervals between biological measurements taken across a set of quantitative treatments. The algorithm provides a probabilistic distribution of possible gene expression values within unmeasured intervals, based on a plausible biological constraint. We show how quantification of this uncertainty can be used to guide researchers in further data collection by identifying which samples would likely add the most information to the system under study. Although the context for developing the algorithm was gene expression measurements taken over a time series, the approach can be readily applied to any set of quantitative systems biology measurements taken following quantitative (i.e. non-categorical) treatments. In principle, the method could also be applied to combinations of treatments, in which case it could greatly simplify the task of exploring the large combinatorial space of future possible measurements.

  7. From gene expressions to genetic networks

    NASA Astrophysics Data System (ADS)

    Cieplak, Marek

    2009-03-01

    A method based on the principle of entropy maximization is used to identify the gene interaction network with the highest probability of giving rise to experimentally observed transcript profiles [1]. In its simplest form, the method yields the pairwise gene interaction network, but it can also be extended to deduce higher order correlations. Analysis of microarray data from genes in Saccharomyces cerevisiae chemostat cultures exhibiting energy metabollic oscillations identifies a gene interaction network that reflects the intracellular communication pathways. These pathways adjust cellular metabolic activity and cell division to the limiting nutrient conditions that trigger metabolic oscillations. The success of the present approach in extracting meaningful genetic connections suggests that the maximum entropy principle is a useful concept for understanding living systems, as it is for other complex, nonequilibrium systems. The time-dependent behavior of the genetic network is found to involve only a few fundamental modes [2,3]. [4pt] REFERENCES:[0pt] [1] T. R. Lezon, J. R. Banavar, M. Cieplak, A. Maritan, and N. Fedoroff, Using the principle of entropy maximization to infer genetic interaction networks from gene expression patterns, Proc. Natl. Acad. Sci. (USA) 103, 19033-19038 (2006) [0pt] [2] N. S. Holter, M. Mitra, A. Maritan, M. Cieplak, J. R. Banavar, and N. V. Fedoroff, Fundamental patterns underlying gene expression profiles: simplicity from complexity, Proc. Natl. Acad. Sci. USA 97, 8409-8414 (2000) [0pt] [3] N. S. Holter, A. Maritan, M. Cieplak, N. V. Fedoroff, and J. R. Banavar, Dynamic modeling of gene expression data, Proc. Natl. Acad. Sci. USA 98, 1693-1698 (2001)

  8. Determining Physical Mechanisms of Gene Expression Regulation from Single Cell Gene Expression Data

    PubMed Central

    Moignard, Victoria; Göttgens, Berthold; Adryan, Boris

    2016-01-01

    Many genes are expressed in bursts, which can contribute to cell-to-cell heterogeneity. It is now possible to measure this heterogeneity with high throughput single cell gene expression assays (single cell qPCR and RNA-seq). These experimental approaches generate gene expression distributions which can be used to estimate the kinetic parameters of gene expression bursting, namely the rate that genes turn on, the rate that genes turn off, and the rate of transcription. We construct a complete pipeline for the analysis of single cell qPCR data that uses the mathematics behind bursty expression to develop more accurate and robust algorithms for analyzing the origin of heterogeneity in experimental samples, specifically an algorithm for clustering cells by their bursting behavior (Simulated Annealing for Bursty Expression Clustering, SABEC) and a statistical tool for comparing the kinetic parameters of bursty expression across populations of cells (Estimation of Parameter changes in Kinetics, EPiK). We applied these methods to hematopoiesis, including a new single cell dataset in which transcription factors (TFs) involved in the earliest branchpoint of blood differentiation were individually up- and down-regulated. We could identify two unique sub-populations within a seemingly homogenous group of hematopoietic stem cells. In addition, we could predict regulatory mechanisms controlling the expression levels of eighteen key hematopoietic transcription factors throughout differentiation. Detailed information about gene regulatory mechanisms can therefore be obtained simply from high throughput single cell gene expression data, which should be widely applicable given the rapid expansion of single cell genomics. PMID:27551778

  9. Coevolution of gene expression among interacting proteins

    SciTech Connect

    Fraser, Hunter B.; Hirsh, Aaron E.; Wall, Dennis P.; Eisen,Michael B.

    2004-03-01

    Physically interacting proteins or parts of proteins are expected to evolve in a coordinated manner that preserves proper interactions. Such coevolution at the amino acid-sequence level is well documented and has been used to predict interacting proteins, domains, and amino acids. Interacting proteins are also often precisely coexpressed with one another, presumably to maintain proper stoichiometry among interacting components. Here, we show that the expression levels of physically interacting proteins coevolve. We estimate average expression levels of genes from four closely related fungi of the genus Saccharomyces using the codon adaptation index and show that expression levels of interacting proteins exhibit coordinated changes in these different species. We find that this coevolution of expression is a more powerful predictor of physical interaction than is coevolution of amino acid sequence. These results demonstrate previously uncharacterized coevolution of gene expression, adding a different dimension to the study of the coevolution of interacting proteins and underscoring the importance of maintaining coexpression of interacting proteins over evolutionary time. Our results also suggest that expression coevolution can be used for computational prediction of protein protein interactions.

  10. Correlation between Gene Expression and Osteoarthritis Progression in Human

    PubMed Central

    Zhong, Leilei; Huang, Xiaobin; Karperien, Marcel; Post, Janine N.

    2016-01-01

    Osteoarthritis (OA) is a multifactorial disease characterized by gradual degradation of joint cartilage. This study aimed to quantify major pathogenetic factors during OA progression in human cartilage. Cartilage specimens were isolated from OA patients and scored 0–5 according to the Osteoarthritis Research Society International (OARSI) guidelines. Protein and gene expressions were measured by immunohistochemistry and qPCR, respectively. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays were used to detect apoptotic cells. Cartilage degeneration in OA is a gradual progress accompanied with gradual loss of collagen type II and a gradual decrease in mRNA expression of SOX9, ACAN and COL2A1. Expression of WNT antagonists DKK1 and FRZB was lost, while hypertrophic markers (RUNX2, COL10A1 and IHH) increased during OA progression. Moreover, DKK1 and FRZB negatively correlated with OA grading, while RUNX2 and IHH showed a significantly positive correlation with OA grading. The number of apoptotic cells was increased with the severity of OA. Taken together, our results suggested that genetic profiling of the gene expression could be used as markers for staging OA at the molecular level. This helps to understand the molecular pathology of OA and may lead to the development of therapies based on OA stage. PMID:27428952

  11. Gene expression regulation in roots under drought.

    PubMed

    Janiak, Agnieszka; Kwaśniewski, Mirosław; Szarejko, Iwona

    2016-02-01

    Stress signalling and regulatory networks controlling expression of target genes are the basis of plant response to drought. Roots are the first organs exposed to water deficiency in the soil and are the place of drought sensing. Signalling cascades transfer chemical signals toward the shoot and initiate molecular responses that lead to the biochemical and morphological changes that allow plants to be protected against water loss and to tolerate stress conditions. Here, we present an overview of signalling network and gene expression regulation pathways that are actively induced in roots under drought stress. In particular, the role of several transcription factor (TF) families, including DREB, AP2/ERF, NAC, bZIP, MYC, CAMTA, Alfin-like and Q-type ZFP, in the regulation of root response to drought are highlighted. The information provided includes available data on mutual interactions between these TFs together with their regulation by plant hormones and other signalling molecules. The most significant downstream target genes and molecular processes that are controlled by the regulatory factors are given. These data are also coupled with information about the influence of the described regulatory networks on root traits and root development which may translate to enhanced drought tolerance. This is the first literature survey demonstrating the gene expression regulatory machinery that is induced by drought stress, presented from the perspective of roots.

  12. Expression of bacterial genes in plant cells.

    PubMed Central

    Fraley, R T; Rogers, S G; Horsch, R B; Sanders, P R; Flick, J S; Adams, S P; Bittner, M L; Brand, L A; Fink, C L; Fry, J S; Galluppi, G R; Goldberg, S B; Hoffmann, N L; Woo, S C

    1983-01-01

    Chimeric bacterial genes conferring resistance to aminoglycoside antibiotics have been inserted into the Agrobacterium tumefaciens tumor-inducing (Ti) plasmid and introduced into plant cells by in vitro transformation techniques. The chimeric genes contain the nopaline synthase 5' and 3' regulatory regions joined to the genes for neomycin phosphotransferase type I or type II. The chimeric genes were cloned into an intermediate vector, pMON120, and inserted into pTiB6S3 by recombination and then introduced into petunia and tobacco cells by cocultivating A. tumefaciens cells with protoplast-derived cells. Southern hybridization was used to confirm the presence of the chimeric genes in the transformed plant tissues. Expression of the chimeric genes was determined by the ability of the transformed cells to proliferate on medium containing normally inhibitory levels of kanamycin (50 micrograms/ml) or other aminoglycoside antibiotics. Plant cells transformed by wild-type pTiB6S3 or derivatives carrying the bacterial neomycin phosphotransferase genes with their own promoters failed to grow under these conditions. The significance of these results for plant genetic engineering is discussed. Images PMID:6308651

  13. Gene expression pattern in canine mammary osteosarcoma.

    PubMed

    Pawłowski, K M; Majewska, A; Szyszko, K; Dolka, I; Motyl, T; Król, M

    2011-01-01

    Canine mammary sarcomas are usually very aggressive and easily metastasize. Unfortunately the biology of this type of tumor is not well known because they are a very rare type of tumors. The aim of this study was to find differences in gene expression patterns in canine mammary osteosarcomas (malignant) versus osteomas (benign) using DNA microarrays. Our microarray experiment showed that 11 genes were up-regulated in osteosarcoma in comparison to osteoma whereas 36 genes were down-regulated. Among the up-regulated genes were: PDK1, EXT1, and EIF4H which are involved in AKT/PI3K and GLI/Hedgehog pathways. These genes play an important role in cell biology (cancer cell proliferation) and may be essential in osteosarcoma formation and development. Analyzing the down-regulated genes, the most interesting seemed to be HSPB8 and SEPP1. HSPB8 is a small heat shock protein that plays an important role in cell cycle regulation, apoptosis, and breast carcinogenesis. Also SEPP1 may play a role in carcinogenesis, as its down-regulation may induce oxidative stress possibly resulting in carcinogenesis. The preliminary results of the present study indicate that the up-regulation of three genes EXT1, EIF4H, and PDK1 may play an essential role in osteosarcoma formation, development and proliferation. In our opinion the cross-talk between GLI/Hedgehog and PI3K/AKT pathways may be a key factor to increase tumor proliferation and malignancy. PMID:21528706

  14. Pathophysiological factors affecting CAR gene expression.

    PubMed

    Pascussi, Jean Marc; Dvorák, Zdenek; Gerbal-Chaloin, Sabine; Assenat, Eric; Maurel, Patrick; Vilarem, Marie José

    2003-11-01

    The body defends itself against potentially harmful compounds, such as drugs and toxic endogenous compounds and their metabolites, by inducing the expression of enzymes and transporters involved in their metabolism and elimination. The orphan nuclear receptor CAR (NR1I3 controls phase I (CYP2B, CYP2C, CYP3A), phase II (UGT1A1), and transporter (SLC21A6, MRP2) genes involved in drug metabolism and bilirubin clearance. Constitutive androstane receptor (CAR) is activated by xenobiotics, such as phenobarbital, but also by toxic endogenous compounds such as bilirubin metabolite(s). To better understand the inter- and intravariability in drug detoxification, we studied the molecular mechanisms involved in CAR gene expression in human hepatocytes. We clearly identified CAR as a glucocorticoid receptor (GR) target gene, and we proposed the hypothesis of a signal transduction where the activation of GR plays a critical function in CAR-mediated cellular response. According to our model, chemicals or pathophysiological factors that affect GR function should decrease CAR function. To test this hypothesis, we recently investigated the effect of microtubule disrupting agents (MIAs) or proinflammatory cytokines. These compounds are well-known inhibitors of GR transactivation property. MIAs activate c-Jun N-terminal kinase (JNK), which phosphorylates and inactivates GR, whereas proinflammatory cytokines, such as IL-6 or IL1beta, induce AP-1 or NF-kB activation, respectively, leading to GR inhibition. As expected, we observed that these molecules inhibit both CAR gene expression and phenobarbital-mediated CYP gene expression in human hepatocytes. PMID:14705859

  15. Gene expression profiles in skeletal muscle after gene electrotransfer

    PubMed Central

    Hojman, Pernille; Zibert, John R; Gissel, Hanne; Eriksen, Jens; Gehl, Julie

    2007-01-01

    Background Gene transfer by electroporation (DNA electrotransfer) to muscle results in high level long term transgenic expression, showing great promise for treatment of e.g. protein deficiency syndromes. However little is known about the effects of DNA electrotransfer on muscle fibres. We have therefore investigated transcriptional changes through gene expression profile analyses, morphological changes by histological analysis, and physiological changes by force generation measurements. DNA electrotransfer was obtained using a combination of a short high voltage pulse (HV, 1000 V/cm, 100 μs) followed by a long low voltage pulse (LV, 100 V/cm, 400 ms); a pulse combination optimised for efficient and safe gene transfer. Muscles were transfected with green fluorescent protein (GFP) and excised at 4 hours, 48 hours or 3 weeks after treatment. Results Differentially expressed genes were investigated by microarray analysis, and descriptive statistics were performed to evaluate the effects of 1) electroporation, 2) DNA injection, and 3) time after treatment. The biological significance of the results was assessed by gene annotation and supervised cluster analysis. Generally, electroporation caused down-regulation of structural proteins e.g. sarcospan and catalytic enzymes. Injection of DNA induced down-regulation of intracellular transport proteins e.g. sentrin. The effects on muscle fibres were transient as the expression profiles 3 weeks after treatment were closely related with the control muscles. Most interestingly, no changes in the expression of proteins involved in inflammatory responses or muscle regeneration was detected, indicating limited muscle damage and regeneration. Histological analysis revealed structural changes with loss of cell integrity and striation pattern in some fibres after DNA+HV+LV treatment, while HV+LV pulses alone showed preservation of cell integrity. No difference in the force generation capacity was observed in the muscles 2 weeks

  16. The Relationship between Gene Network Structure and Expression Variation among Individuals and Species.

    PubMed

    Sears, Karen E; Maier, Jennifer A; Rivas-Astroza, Marcelo; Poe, Rachel; Zhong, Sheng; Kosog, Kari; Marcot, Jonathan D; Behringer, Richard R; Cretekos, Chris J; Rasweiler, John J; Rapti, Zoi

    2015-08-01

    Variation among individuals is a prerequisite of evolution by natural selection. As such, identifying the origins of variation is a fundamental goal of biology. We investigated the link between gene interactions and variation in gene expression among individuals and species using the mammalian limb as a model system. We first built interaction networks for key genes regulating early (outgrowth; E9.5-11) and late (expansion and elongation; E11-13) limb development in mouse. This resulted in an Early (ESN) and Late (LSN) Stage Network. Computational perturbations of these networks suggest that the ESN is more robust. We then quantified levels of the same key genes among mouse individuals and found that they vary less at earlier limb stages and that variation in gene expression is heritable. Finally, we quantified variation in gene expression levels among four mammals with divergent limbs (bat, opossum, mouse and pig) and found that levels vary less among species at earlier limb stages. We also found that variation in gene expression levels among individuals and species are correlated for earlier and later limb development. In conclusion, results are consistent with the robustness of the ESN buffering among-individual variation in gene expression levels early in mammalian limb development, and constraining the evolution of early limb development among mammalian species. PMID:26317994

  17. The Relationship between Gene Network Structure and Expression Variation among Individuals and Species

    PubMed Central

    Sears, Karen E.; Maier, Jennifer A.; Rivas-Astroza, Marcelo; Poe, Rachel; Zhong, Sheng; Kosog, Kari; Marcot, Jonathan D.; Behringer, Richard R.; Rasweiler, John J.; Rapti, Zoi

    2015-01-01

    Abstract Variation among individuals is a prerequisite of evolution by natural selection. As such, identifying the origins of variation is a fundamental goal of biology. We investigated the link between gene interactions and variation in gene expression among individuals and species using the mammalian limb as a model system. We first built interaction networks for key genes regulating early (outgrowth; E9.5–11) and late (expansion and elongation; E11-13) limb development in mouse. This resulted in an Early (ESN) and Late (LSN) Stage Network. Computational perturbations of these networks suggest that the ESN is more robust. We then quantified levels of the same key genes among mouse individuals and found that they vary less at earlier limb stages and that variation in gene expression is heritable. Finally, we quantified variation in gene expression levels among four mammals with divergent limbs (bat, opossum, mouse and pig) and found that levels vary less among species at earlier limb stages. We also found that variation in gene expression levels among individuals and species are correlated for earlier and later limb development. In conclusion, results are consistent with the robustness of the ESN buffering among-individual variation in gene expression levels early in mammalian limb development, and constraining the evolution of early limb development among mammalian species. PMID:26317994

  18. Decomposition of Gene Expression State Space Trajectories

    PubMed Central

    Mar, Jessica C.; Quackenbush, John

    2009-01-01

    Representing and analyzing complex networks remains a roadblock to creating dynamic network models of biological processes and pathways. The study of cell fate transitions can reveal much about the transcriptional regulatory programs that underlie these phenotypic changes and give rise to the coordinated patterns in expression changes that we observe. The application of gene expression state space trajectories to capture cell fate transitions at the genome-wide level is one approach currently used in the literature. In this paper, we analyze the gene expression dataset of Huang et al. (2005) which follows the differentiation of promyelocytes into neutrophil-like cells in the presence of inducers dimethyl sulfoxide and all-trans retinoic acid. Huang et al. (2005) build on the work of Kauffman (2004) who raised the attractor hypothesis, stating that cells exist in an expression landscape and their expression trajectories converge towards attractive sites in this landscape. We propose an alternative interpretation that explains this convergent behavior by recognizing that there are two types of processes participating in these cell fate transitions—core processes that include the specific differentiation pathways of promyelocytes to neutrophils, and transient processes that capture those pathways and responses specific to the inducer. Using functional enrichment analyses, specific biological examples and an analysis of the trajectories and their core and transient components we provide a validation of our hypothesis using the Huang et al. (2005) dataset. PMID:20041215

  19. Insights into SAGA function during gene expression

    PubMed Central

    Rodríguez-Navarro, Susana

    2009-01-01

    Histone modifications are a crucial source of epigenetic control. SAGA (Spt–Ada–Gcn5 acetyltransferase) is a chromatin-modifying complex that contains two distinct enzymatic activities, Gcn5 and Ubp8, through which it acetylates and deubiquitinates histone residues, respectively, thereby enforcing a pattern of modifications that is decisive in regulating gene expression. Here, I discuss the latest contributions to understanding the roles of the SAGA complex, highlighting the characterization of the SAGA-deubiquitination module, and emphasizing the functions newly ascribed to SAGA during transcription elongation and messenger-RNA export. These findings suggest that a crosstalk exists between chromatin remodelling, transcription and messenger-RNA export, which could constitute a checkpoint for accurate gene expression. I focus particularly on the new components of human SAGA, which was recently discovered and confirms the conservation of the SAGA complex throughout evolution. PMID:19609321

  20. Structure, expression and functions of MTA genes.

    PubMed

    Kumar, Rakesh; Wang, Rui-An

    2016-05-15

    Metastatic associated proteins (MTA) are integrators of upstream regulatory signals with the ability to act as master coregulators for modifying gene transcriptional activity. The MTA family includes three genes and multiple alternatively spliced variants. The MTA proteins neither have their own enzymatic activity nor have been shown to directly interact with DNA. However, MTA proteins interact with a variety of chromatin remodeling factors and complexes with enzymatic activities for modulating the plasticity of nucleosomes, leading to the repression or derepression of target genes or other extra-nuclear and nucleosome remodeling and histone deacetylase (NuRD)-complex independent activities. The functions of MTA family members are driven by the steady state levels and subcellular localization of MTA proteins, the dynamic nature of modifying signals and enzymes, the structural features and post-translational modification of protein domains, interactions with binding proteins, and the nature of the engaged and resulting features of nucleosomes in the proximity of target genes. In general, MTA1 and MTA2 are the most upregulated genes in human cancer and correlate well with aggressive phenotypes, therapeutic resistance, poor prognosis and ultimately, unfavorable survival of cancer patients. Here we will discuss the structure, expression and functions of the MTA family of genes in the context of cancer cells. PMID:26869315

  1. Identifying driver genes in cancer by triangulating gene expression, gene location, and survival data.

    PubMed

    Rouam, Sigrid; Miller, Lance D; Karuturi, R Krishna Murthy

    2014-01-01

    Driver genes are directly responsible for oncogenesis and identifying them is essential in order to fully understand the mechanisms of cancer. However, it is difficult to delineate them from the larger pool of genes that are deregulated in cancer (ie, passenger genes). In order to address this problem, we developed an approach called TRIAngulating Gene Expression (TRIAGE through clinico-genomic intersects). Here, we present a refinement of this approach incorporating a new scoring methodology to identify putative driver genes that are deregulated in cancer. TRIAGE triangulates - or integrates - three levels of information: gene expression, gene location, and patient survival. First, TRIAGE identifies regions of deregulated expression (ie, expression footprints) by deriving a newly established measure called the Local Singular Value Decomposition (LSVD) score for each locus. Driver genes are then distinguished from passenger genes using dual survival analyses. Incorporating measurements of gene expression and weighting them according to the LSVD weight of each tumor, these analyses are performed using the genes located in significant expression footprints. Here, we first use simulated data to characterize the newly established LSVD score. We then present the results of our application of this refined version of TRIAGE to gene expression data from five cancer types. This refined version of TRIAGE not only allowed us to identify known prominent driver genes, such as MMP1, IL8, and COL1A2, but it also led us to identify several novel ones. These results illustrate that TRIAGE complements existing tools, allows for the identification of genes that drive cancer and could perhaps elucidate potential future targets of novel anticancer therapeutics.

  2. Gene expression: RNA interference in adult mice

    NASA Astrophysics Data System (ADS)

    McCaffrey, Anton P.; Meuse, Leonard; Pham, Thu-Thao T.; Conklin, Douglas S.; Hannon, Gregory J.; Kay, Mark A.

    2002-07-01

    RNA interference is an evolutionarily conserved surveillance mechanism that responds to double-stranded RNA by sequence-specific silencing of homologous genes. Here we show that transgene expression can be suppressed in adult mice by synthetic small interfering RNAs and by small-hairpin RNAs transcribed in vivo from DNA templates. We also show the therapeutic potential of this technique by demonstrating effective targeting of a sequence from hepatitis C virus by RNA interference in vivo.

  3. Imaging gene expression in single living cells

    PubMed Central

    Shav-Tal, Yaron; Singer, Robert H.; Darzacq, Xavier

    2016-01-01

    Technical advances in the field of live-cell imaging have introduced the cell biologist to a new, dynamic, subcellular world. The static world of molecules in fixed cells has now been extended to the time dimension. This allows the visualization and quantification of gene expression and intracellular trafficking events of the studied molecules and the associated enzymatic processes in individual cells, in real time. PMID:15459666

  4. The systemic control of circadian gene expression.

    PubMed

    Gerber, A; Saini, C; Curie, T; Emmenegger, Y; Rando, G; Gosselin, P; Gotic, I; Gos, P; Franken, P; Schibler, U

    2015-09-01

    The mammalian circadian timing system consists of a central pacemaker in the brain's suprachiasmatic nucleus (SCN) and subsidiary oscillators in nearly all body cells. The SCN clock, which is adjusted to geophysical time by the photoperiod, synchronizes peripheral clocks through a wide variety of systemic cues. The latter include signals depending on feeding cycles, glucocorticoid hormones, rhythmic blood-borne signals eliciting daily changes in actin dynamics and serum response factor (SRF) activity, and sensors of body temperature rhythms, such as heat shock transcription factors and the cold-inducible RNA-binding protein CIRP. To study these systemic signalling pathways, we designed and engineered a novel, highly photosensitive apparatus, dubbed RT-Biolumicorder. This device enables us to record circadian luciferase reporter gene expression in the liver and other organs of freely moving mice over months in real time. Owing to the multitude of systemic signalling pathway involved in the phase resetting of peripheral clocks the disruption of any particular one has only minor effects on the steady state phase of circadian gene expression in organs such as the liver. Nonetheless, the implication of specific pathways in the synchronization of clock gene expression can readily be assessed by monitoring the phase-shifting kinetics using the RT-Biolumicorder.

  5. Carbon Nanomaterials Alter Global Gene Expression Profiles.

    PubMed

    Woodman, Sara; Short, John C W; McDermott, Hyoeun; Linan, Alexander; Bartlett, Katelyn; Gadila, Shiva Kumar Goud; Schmelzle, Katie; Wanekaya, Adam; Kim, Kyoungtae

    2016-05-01

    Carbon nanomaterials (CNMs), which include carbon nanotubes (CNTs) and their derivatives, have diverse technological and biomedical applications. The potential toxicity of CNMs to cells and tissues has become an important emerging question in nanotechnology. To assess the toxicity of CNTs and fullerenol C60(OH)24, we in the present work used the budding yeast Saccharomyces cerevisiae, one of the simplest eukaryotic organisms that share fundamental aspects of eukaryotic cell biology. We found that treatment with CNMs, regardless of their physical shape, negatively affected the growth rates, end-point cell densities and doubling times of CNM-exposed yeast cells when compared to unexposed cells. To investigate potential mechanisms behind the CNMs-induced growth defects, we performed RNA-Seq dependent transcriptional analysis and constructed global gene expression profiles of fullerenol C60(OH)24- and CNT-treated cells. When compared to non-treated control cells, CNM-treated cells displayed differential expression of genes whose functions are implicated in membrane transporters and stress response, although differentially expressed genes were not consistent between CNT- and fullerenol C60(OH)24-treated groups, leading to our conclusion that CNMs could serve as environmental toxic factors to eukaryotic cells. PMID:27483901

  6. Expression of foreign genes in filamentous cyanobacteria

    SciTech Connect

    Kuritz, T.; Wolk, C.P. )

    1993-06-01

    Several advantages make cyanobacteria attractive hosts for biodegradative genes and possibly for other exogenous genes that have practical uses. The authors have obtained expression in Anabaena sp. strain PCC 7120 and Nostoc ellipsosporum of a dechlorination operon, fcbAB, from Arthrobacter globiformis, and have also developed a simple method for qualitative assessment of dechlorination by microorganisms, such as cyanobacteria, whose metabolism is dependent on the presence of chloride in the medium. Transcription of fcbAB under the control of a variety of promoters was monitored by placing luxAB (encoding luciferase) downstream from fcbAB, and by measuring light emission from luciferase. They believe that the system that they have described has value as a means to screen for factors influencing transcription of foreign genes in cyanobacteria.

  7. [Transcriptomes for serial analysis of gene expression].

    PubMed

    Marti, Jacques; Piquemal, David; Manchon, Laurent; Commes, Thérèse

    2002-01-01

    The availability of the sequences for whole genomes is changing our understanding of cell biology. Functional genomics refers to the comprehensive analysis, at the protein level (proteome) and at the mRNA level (transcriptome) of all events associated with the expression of whole sets of genes. New methods have been developed for transcriptome analysis. Serial Analysis of Gene Expression (SAGE) is based on the massive sequential analysis of short cDNA sequence tags. Each tag is derived from a defined position within a transcript. Its size (14 bp) is sufficient to identify the corresponding gene and the number of times each tag is observed provides an accurate measurement of its expression level. Since tag populations can be widely amplified without altering their relative proportions, SAGE may be performed with minute amounts of biological extract. Dealing with the mass of data generated by SAGE necessitates computer analysis. A software is required to automatically detect and count tags from sequence files. Criterias allowing to assess the quality of experimental data can be included at this stage. To identify the corresponding genes, a database is created registering all virtual tags susceptible to be observed, based on the present status of the genome knowledge. By using currently available database functions, it is easy to match experimental and virtual tags, thus generating a new database registering identified tags, together with their expression levels. As an open system, SAGE is able to reveal new, yet unknown, transcripts. Their identification will become increasingly easier with the progress of genome annotation. However, their direct characterization can be attempted, since tag information may be sufficient to design primers allowing to extend unknown sequences. A major advantage of SAGE is that, by measuring expression levels without reference to an arbitrary standard, data are definitively acquired and cumulative. All publicly available data can thus

  8. Reduced expression of Autographa californica nucleopolyhedrovirus ORF34, an essential gene, enhances heterologous gene expression

    SciTech Connect

    Salem, Tamer Z.; Zhang, Fengrui; Thiem, Suzanne M.

    2013-01-20

    Autographa californica multiple nucleopolyhedrovirus ORF34 is part of a transcriptional unit that includes ORF32, encoding a viral fibroblast growth factor (FGF) and ORF33. We identified ORF34 as a candidate for deletion to improve protein expression in the baculovirus expression system based on enhanced reporter gene expression in an RNAi screen of virus genes. However, ORF34 was shown to be an essential gene. To explore ORF34 function, deletion (KO34) and rescue bacmids were constructed and characterized. Infection did not spread from primary KO34 transfected cells and supernatants from KO34 transfected cells could not infect fresh Sf21 cells whereas the supernatant from the rescue bacmids transfection could recover the infection. In addition, budded viruses were not observed in KO34 transfected cells by electron microscopy, nor were viral proteins detected from the transfection supernatants by western blots. These demonstrate that ORF34 is an essential gene with a possible role in infectious virus production.

  9. Screening of differentially expressed genes in pathological scar tissues using expression microarray.

    PubMed

    Huang, L P; Mao, Z; Zhang, L; Liu, X X; Huang, C; Jia, Z S

    2015-01-01

    Pathological scar tissues and normal skin tissues were differentiated by screening for differentially expressed genes in pathologic scar tissues via gene expression microarray. The differentially expressed gene data was analyzed by gene ontology and pathway analyses. There were 5001 up- or down-regulated genes in 2-fold differentially expressed genes, 956 up- or down-regulated genes in 5-fold differentially expressed genes, and 114 up- or down-regulated genes in 20-fold differentially expressed genes. Therefore, significant differences were observed in the gene expression in pathological scar tissues and normal foreskin tissues. The development of pathological scar tissues has been correlated to changes in multiple genes and pathways, which are believed to form a dynamic network connection.

  10. Comparative analysis of Phytophthora infestans induced gene expression in potato cultivars with different levels of resistance.

    PubMed

    Ros, B; Thümmler, F; Wenzel, G

    2005-11-01

    Differential gene expression was analyzed after infection with Phytophthora infestans in six potato cultivars with different levels of resistance to late blight. To verify the infection of the potato leaflets, the amount of phytopathogen mRNA within the plant material was quantified by real-time quantitative PCR. The expression of 182 genes selected from two subtracted cDNA libraries was studied with cDNA array hybridization using RNA from non-infected and infected potato leaflets. Gene up- and down-regulation were clearly detectable in all cultivars 72 h post inoculation. Gene expression patterns in susceptible cultivars differed from those in potato varieties with a higher level of resistance. In general, a stronger gene induction was observed in the susceptible cultivars compared to the moderately to highly resistant potato varieties. Five genes with the highest homology to stress and/or defence-related genes were induced specifically in the susceptible cultivars. Four genes responded to pathogen attack independently of the level of resistance of the cultivar used, and three genes were repressed in infected tissue of most cultivars. Even in the absence of P. infestans infection, six genes showed higher expression levels in the somewhat resistant cultivars Bettina and Matilda. Possible reasons for the different levels of gene expression are discussed.

  11. Murine erythropoietin gene: cloning, expression, and human gene homology.

    PubMed Central

    Shoemaker, C B; Mitsock, L D

    1986-01-01

    The gene for murine erythropoietin (EPO) was isolated from a mouse genomic library with a human EPO cDNA probe. Nucleotide sequence analysis permitted the identification of the murine EPO coding sequence and the prediction of the encoded amino acid sequence based on sequence conservation between the mouse and human EPO genes. Both the coding DNA and the amino acid sequences were 80% conserved between the two species. Transformation of COS-1 cells with a mammalian cell expression vector containing the murine EPO coding region resulted in secretion of murine EPO with biological activity on both murine and human erythroid progenitor cells. The transcription start site for the murine EPO gene in kidneys was determined. This permitted tentative identification of the transcription control region. The region included 140 base pairs upstream of the cap site which was over 90% conserved between the murine and human genes. Surprisingly, the first intron and much of the 5'- and 3'-untranslated sequences were also substantially conserved between the genes of the two species. Images PMID:3773894

  12. Identification of common prognostic gene expression signatures with biological meanings from microarray gene expression datasets.

    PubMed

    Yao, Jun; Zhao, Qi; Yuan, Ying; Zhang, Li; Liu, Xiaoming; Yung, W K Alfred; Weinstein, John N

    2012-01-01

    Numerous prognostic gene expression signatures for breast cancer were generated previously with few overlap and limited insight into the biology of the disease. Here we introduce a novel algorithm named SCoR (Survival analysis using Cox proportional hazard regression and Random resampling) to apply random resampling and clustering methods in identifying gene features correlated with time to event data. This is shown to reduce overfitting noises involved in microarray data analysis and discover functional gene sets linked to patient survival. SCoR independently identified a common poor prognostic signature composed of cell proliferation genes from six out of eight breast cancer datasets. Furthermore, a sequential SCoR analysis on highly proliferative breast cancers repeatedly identified T/B cell markers as favorable prognosis factors. In glioblastoma, SCoR identified a common good prognostic signature of chromosome 10 genes from two gene expression datasets (TCGA and REMBRANDT), recapitulating the fact that loss of one copy of chromosome 10 (which harbors the tumor suppressor PTEN) is linked to poor survival in glioblastoma patients. SCoR also identified prognostic genes on sex chromosomes in lung adenocarcinomas, suggesting patient gender might be used to predict outcome in this disease. These results demonstrate the power of SCoR to identify common and biologically meaningful prognostic gene expression signatures.

  13. Identification of Common Prognostic Gene Expression Signatures with Biological Meanings from Microarray Gene Expression Datasets

    PubMed Central

    Yao, Jun; Zhao, Qi; Yuan, Ying; Zhang, Li; Liu, Xiaoming; Yung, W. K. Alfred; Weinstein, John N.

    2012-01-01

    Numerous prognostic gene expression signatures for breast cancer were generated previously with few overlap and limited insight into the biology of the disease. Here we introduce a novel algorithm named SCoR (Survival analysis using Cox proportional hazard regression and Random resampling) to apply random resampling and clustering methods in identifying gene features correlated with time to event data. This is shown to reduce overfitting noises involved in microarray data analysis and discover functional gene sets linked to patient survival. SCoR independently identified a common poor prognostic signature composed of cell proliferation genes from six out of eight breast cancer datasets. Furthermore, a sequential SCoR analysis on highly proliferative breast cancers repeatedly identified T/B cell markers as favorable prognosis factors. In glioblastoma, SCoR identified a common good prognostic signature of chromosome 10 genes from two gene expression datasets (TCGA and REMBRANDT), recapitulating the fact that loss of one copy of chromosome 10 (which harbors the tumor suppressor PTEN) is linked to poor survival in glioblastoma patients. SCoR also identified prognostic genes on sex chromosomes in lung adenocarcinomas, suggesting patient gender might be used to predict outcome in this disease. These results demonstrate the power of SCoR to identify common and biologically meaningful prognostic gene expression signatures. PMID:23029298

  14. X chromosome regulation of autosomal gene expression in bovine blastocysts.

    PubMed

    Itoh, Yuichiro; Arnold, Arthur P

    2014-10-01

    Although X chromosome inactivation in female mammals evolved to balance the expression of X chromosome and autosomal genes in the two sexes, female embryos pass through developmental stages in which both X chromosomes are active in somatic cells. Bovine blastocysts show higher expression of many X genes in XX than XY embryos, suggesting that X inactivation is not complete. Here, we reanalyzed bovine blastocyst microarray expression data from a network perspective with a focus on interactions between X chromosome and autosomal genes. Whereas male-to-female ratios of expression of autosomal genes were distributed around a mean of 1, X chromosome genes were clearly shifted towards higher expression in females. We generated gene coexpression networks and identified a major module of genes with correlated gene expression that includes female-biased X genes and sexually dimorphic autosomal genes for which the sexual dimorphism is likely driven by the X genes. In this module, expression of X chromosome genes correlates with autosome genes, more than the expression of autosomal genes with each other. Our study identifies correlated patterns of autosomal and X-linked genes that are likely influenced by the sexual imbalance of X gene expression when X inactivation is inefficient.

  15. A gene expression signature for insulin resistance.

    PubMed

    Konstantopoulos, Nicky; Foletta, Victoria C; Segal, David H; Shields, Katherine A; Sanigorski, Andrew; Windmill, Kelly; Swinton, Courtney; Connor, Tim; Wanyonyi, Stephen; Dyer, Thomas D; Fahey, Richard P; Watt, Rose A; Curran, Joanne E; Molero, Juan-Carlos; Krippner, Guy; Collier, Greg R; James, David E; Blangero, John; Jowett, Jeremy B; Walder, Ken R

    2011-02-11

    Insulin resistance is a heterogeneous disorder caused by a range of genetic and environmental factors, and we hypothesize that its etiology varies considerably between individuals. This heterogeneity provides significant challenges to the development of effective therapeutic regimes for long-term management of type 2 diabetes. We describe a novel strategy, using large-scale gene expression profiling, to develop a gene expression signature (GES) that reflects the overall state of insulin resistance in cells and patients. The GES was developed from 3T3-L1 adipocytes that were made "insulin resistant" by treatment with tumor necrosis factor-α (TNF-α) and then reversed with aspirin and troglitazone ("resensitized"). The GES consisted of five genes whose expression levels best discriminated between the insulin-resistant and insulin-resensitized states. We then used this GES to screen a compound library for agents that affected the GES genes in 3T3-L1 adipocytes in a way that most closely resembled the changes seen when insulin resistance was successfully reversed with aspirin and troglitazone. This screen identified both known and new insulin-sensitizing compounds including nonsteroidal anti-inflammatory agents, β-adrenergic antagonists, β-lactams, and sodium channel blockers. We tested the biological relevance of this GES in participants in the San Antonio Family Heart Study (n = 1,240) and showed that patients with the lowest GES scores were more insulin resistant (according to HOMA_IR and fasting plasma insulin levels; P < 0.001). These findings show that GES technology can be used for both the discovery of insulin-sensitizing compounds and the characterization of patients into subtypes of insulin resistance according to GES scores, opening the possibility of developing a personalized medicine approach to type 2 diabetes.

  16. Covariance Structure Models for Gene Expression Microarray Data

    ERIC Educational Resources Information Center

    Xie, Jun; Bentler, Peter M.

    2003-01-01

    Covariance structure models are applied to gene expression data using a factor model, a path model, and their combination. The factor model is based on a few factors that capture most of the expression information. A common factor of a group of genes may represent a common protein factor for the transcript of the co-expressed genes, and hence, it…

  17. Consistently altered expression of gene sets in postmortem brains of individuals with major psychiatric disorders

    PubMed Central

    Darby, M M; Yolken, R H; Sabunciyan, S

    2016-01-01

    The measurement of gene expression in postmortem brain is an important tool for understanding the pathogenesis of serious psychiatric disorders. We hypothesized that major molecular deficits associated with psychiatric disease would affect the entire brain, and such deficits may be shared across disorders. We performed RNA sequencing and quantified gene expression in the hippocampus of 100 brains in the Stanley Array Collection followed by replication in the orbitofrontal cortex of 57 brains in the Stanley Neuropathology Consortium. We then identified genes and canonical pathway gene sets with significantly altered expression in schizophrenia and bipolar disorder in the hippocampus and in schizophrenia, bipolar disorder and major depression in the orbitofrontal cortex. Although expression of individual genes varied, gene sets were significantly enriched in both of the brain regions, and many of these were consistent across diagnostic groups. Further examination of core gene sets with consistently increased or decreased expression in both of the brain regions and across target disorders revealed that ribosomal genes are overexpressed while genes involved in neuronal processes, GABAergic signaling, endocytosis and antigen processing have predominantly decreased expression in affected individuals compared to controls without a psychiatric disorder. Our results highlight pathways of central importance to psychiatric health and emphasize messenger RNA processing and protein synthesis as potential therapeutic targets for all three of the disorders. PMID:27622934

  18. Consistently altered expression of gene sets in postmortem brains of individuals with major psychiatric disorders.

    PubMed

    Darby, M M; Yolken, R H; Sabunciyan, S

    2016-01-01

    The measurement of gene expression in postmortem brain is an important tool for understanding the pathogenesis of serious psychiatric disorders. We hypothesized that major molecular deficits associated with psychiatric disease would affect the entire brain, and such deficits may be shared across disorders. We performed RNA sequencing and quantified gene expression in the hippocampus of 100 brains in the Stanley Array Collection followed by replication in the orbitofrontal cortex of 57 brains in the Stanley Neuropathology Consortium. We then identified genes and canonical pathway gene sets with significantly altered expression in schizophrenia and bipolar disorder in the hippocampus and in schizophrenia, bipolar disorder and major depression in the orbitofrontal cortex. Although expression of individual genes varied, gene sets were significantly enriched in both of the brain regions, and many of these were consistent across diagnostic groups. Further examination of core gene sets with consistently increased or decreased expression in both of the brain regions and across target disorders revealed that ribosomal genes are overexpressed while genes involved in neuronal processes, GABAergic signaling, endocytosis and antigen processing have predominantly decreased expression in affected individuals compared to controls without a psychiatric disorder. Our results highlight pathways of central importance to psychiatric health and emphasize messenger RNA processing and protein synthesis as potential therapeutic targets for all three of the disorders. PMID:27622934

  19. Gene Expression in First Trimester Preeclampsia Placenta

    PubMed Central

    Founds, Sandra A.; Terhorst, Lauren A.; Conrad, Kirk P.; Hogge, W. Allen; Jeyabalan, Arun; Conley, Yvette P.

    2013-01-01

    Background The goal of this study was to further validate eight candidate genes identified in a microarray analysis of first trimester placentas in preeclampsia. Material and method Surplus chorionic villus sampling (CVS) specimens of 4 women subsequently diagnosed with preeclampsia (PE) and 8 control women (C) without preeclampsia analyzed previously by microarray and 24 independent additional control samples (AS) were submitted for confirmatory studies by quantitative real-time polymerase chain reaction (qRT-PCR). Results Downregulation was significant in FSTL3 in PE as compared to C and AS (p = .04). PAEP was downregulated, but the difference was only significant between C and AS (p = .002) rather than between PE and either of the control groups. Expression levels for CFH, EPAS1, IGFBP1, MMP12, and SEMA3C were not statistically different among groups, but trends were consistent with microarray results; there was no anti-correlation. S100A8 was not measurable in all samples, probably because different probes and primers were needed. Conclusions This study corroborates reduced FSTL3 expression in the first trimester of preeclampsia. Nonsignificant trends in the other genes may require follow-up in studies powered for medium or medium/large effect sizes. qRT-PCR verification of the prior microarray of CVS may support the placental origins of preeclampsia hypothesis. Replication is needed for the candidate genes as potential biomarkers of susceptibility, early detection, and/or individualized care of maternal–infant preeclampsia. PMID:21044967

  20. Nuclear AXIN2 represses MYC gene expression

    SciTech Connect

    Rennoll, Sherri A.; Konsavage, Wesley M.; Yochum, Gregory S.

    2014-01-03

    Highlights: •AXIN2 localizes to cytoplasmic and nuclear compartments in colorectal cancer cells. •Nuclear AXIN2 represses the activity of Wnt-responsive luciferase reporters. •β-Catenin bridges AXIN2 to TCF transcription factors. •AXIN2 binds the MYC promoter and represses MYC gene expression. -- Abstract: The β-catenin transcriptional coactivator is the key mediator of the canonical Wnt signaling pathway. In the absence of Wnt, β-catenin associates with a cytosolic and multi-protein destruction complex where it is phosphorylated and targeted for proteasomal degradation. In the presence of Wnt, the destruction complex is inactivated and β-catenin translocates into the nucleus. In the nucleus, β-catenin binds T-cell factor (TCF) transcription factors to activate expression of c-MYC (MYC) and Axis inhibition protein 2 (AXIN2). AXIN2 is a member of the destruction complex and, thus, serves in a negative feedback loop to control Wnt/β-catenin signaling. AXIN2 is also present in the nucleus, but its function within this compartment is unknown. Here, we demonstrate that AXIN2 localizes to the nuclei of epithelial cells within normal and colonic tumor tissues as well as colorectal cancer cell lines. In the nucleus, AXIN2 represses expression of Wnt/β-catenin-responsive luciferase reporters and forms a complex with β-catenin and TCF. We demonstrate that AXIN2 co-occupies β-catenin/TCF complexes at the MYC promoter region. When constitutively localized to the nucleus, AXIN2 alters the chromatin structure at the MYC promoter and directly represses MYC gene expression. These findings suggest that nuclear AXIN2 functions as a rheostat to control MYC expression in response to Wnt/β-catenin signaling.

  1. Integrating constitutive gene expression and chemoactivity: mining the NCI60 anticancer screen.

    PubMed

    Covell, David G

    2012-01-01

    Studies into the genetic origins of tumor cell chemoactivity pose significant challenges to bioinformatic mining efforts. Connections between measures of gene expression and chemoactivity have the potential to identify clinical biomarkers of compound response, cellular pathways important to efficacy and potential toxicities; all vital to anticancer drug development. An investigation has been conducted that jointly explores tumor-cell constitutive NCI60 gene expression profiles and small-molecule NCI60 growth inhibition chemoactivity profiles, viewed from novel applications of self-organizing maps (SOMs) and pathway-centric analyses of gene expressions, to identify subsets of over- and under-expressed pathway genes that discriminate chemo-sensitive and chemo-insensitive tumor cell types. Linear Discriminant Analysis (LDA) is used to quantify the accuracy of discriminating genes to predict tumor cell chemoactivity. LDA results find 15% higher prediction accuracies, using ∼30% fewer genes, for pathway-derived discriminating genes when compared to genes derived using conventional gene expression-chemoactivity correlations. The proposed pathway-centric data mining procedure was used to derive discriminating genes for ten well-known compounds. Discriminating genes were further evaluated using gene set enrichment analysis (GSEA) to reveal a cellular genetic landscape, comprised of small numbers of key over and under expressed on- and off-target pathway genes, as important for a compound's tumor cell chemoactivity. Literature-based validations are provided as support for chemo-important pathways derived from this procedure. Qualitatively similar results are found when using gene expression measurements derived from different microarray platforms. The data used in this analysis is available at http://pubchem.ncbi.nlm.nih.gov/andhttp://www.ncbi.nlm.nih.gov/projects/geo (GPL96, GSE32474).

  2. Molecular mechanisms of curcumin action: gene expression.

    PubMed

    Shishodia, Shishir

    2013-01-01

    Curcumin derived from the tropical plant Curcuma longa has a long history of use as a dietary agent, food preservative, and in traditional Asian medicine. It has been used for centuries to treat biliary disorders, anorexia, cough, diabetic wounds, hepatic disorders, rheumatism, and sinusitis. The preventive and therapeutic properties of curcumin are associated with its antioxidant, anti-inflammatory, and anticancer properties. Extensive research over several decades has attempted to identify the molecular mechanisms of curcumin action. Curcumin modulates numerous molecular targets by altering their gene expression, signaling pathways, or through direct interaction. Curcumin regulates the expression of inflammatory cytokines (e.g., TNF, IL-1), growth factors (e.g., VEGF, EGF, FGF), growth factor receptors (e.g., EGFR, HER-2, AR), enzymes (e.g., COX-2, LOX, MMP9, MAPK, mTOR, Akt), adhesion molecules (e.g., ELAM-1, ICAM-1, VCAM-1), apoptosis related proteins (e.g., Bcl-2, caspases, DR, Fas), and cell cycle proteins (e.g., cyclin D1). Curcumin modulates the activity of several transcription factors (e.g., NF-κB, AP-1, STAT) and their signaling pathways. Based on its ability to affect multiple targets, curcumin has the potential for the prevention and treatment of various diseases including cancers, arthritis, allergies, atherosclerosis, aging, neurodegenerative disease, hepatic disorders, obesity, diabetes, psoriasis, and autoimmune diseases. This review summarizes the molecular mechanisms of modulation of gene expression by curcumin.

  3. Molecular mechanisms of curcumin action: gene expression.

    PubMed

    Shishodia, Shishir

    2013-01-01

    Curcumin derived from the tropical plant Curcuma longa has a long history of use as a dietary agent, food preservative, and in traditional Asian medicine. It has been used for centuries to treat biliary disorders, anorexia, cough, diabetic wounds, hepatic disorders, rheumatism, and sinusitis. The preventive and therapeutic properties of curcumin are associated with its antioxidant, anti-inflammatory, and anticancer properties. Extensive research over several decades has attempted to identify the molecular mechanisms of curcumin action. Curcumin modulates numerous molecular targets by altering their gene expression, signaling pathways, or through direct interaction. Curcumin regulates the expression of inflammatory cytokines (e.g., TNF, IL-1), growth factors (e.g., VEGF, EGF, FGF), growth factor receptors (e.g., EGFR, HER-2, AR), enzymes (e.g., COX-2, LOX, MMP9, MAPK, mTOR, Akt), adhesion molecules (e.g., ELAM-1, ICAM-1, VCAM-1), apoptosis related proteins (e.g., Bcl-2, caspases, DR, Fas), and cell cycle proteins (e.g., cyclin D1). Curcumin modulates the activity of several transcription factors (e.g., NF-κB, AP-1, STAT) and their signaling pathways. Based on its ability to affect multiple targets, curcumin has the potential for the prevention and treatment of various diseases including cancers, arthritis, allergies, atherosclerosis, aging, neurodegenerative disease, hepatic disorders, obesity, diabetes, psoriasis, and autoimmune diseases. This review summarizes the molecular mechanisms of modulation of gene expression by curcumin. PMID:22996381

  4. Combined clustering models for the analysis of gene expression

    SciTech Connect

    Angelova, M. Ellman, J.

    2010-02-15

    Clustering has become one of the fundamental tools for analyzing gene expression and producing gene classifications. Clustering models enable finding patterns of similarity in order to understand gene function, gene regulation, cellular processes and sub-types of cells. The clustering results however have to be combined with sequence data or knowledge about gene functionality in order to make biologically meaningful conclusions. In this work, we explore a new model that integrates gene expression with sequence or text information.

  5. A real-time one-step reverse transcriptase-polymerase chain reaction method to quantify c-erbB-2 expression in human breast cancer.

    PubMed

    Pawlowski, V; Révillion, F; Hornez, L; Peyrat, J P

    2000-01-01

    We developed a real-time one-step reverse transcriptase-polymerase chain reaction (RT-PCR) method for the routine quantification of c-erbB-2 oncogene expression in breast cancer, using a 7700 ABI PRISM Sequence Detector System (Perkin Elmer-Applied Biosystems, Courtaboeuf, France). The real-time quantification of the polymerase chain reaction products is based on the TaqMan 5' nuclease assay. The optimal experimental conditions we determined were as follows: 6 mM MgCl2, 200 nM of fluorogenic probe, 200 nM of each primer, and 12.5 units MuLV reverse transcriptase. The GAPDH housekeeping gene was used for normalization of c-erbB-2 expression. In human breast cancer cell lines, the normalized expression of c-erbB-2 ranged from 8 x 10(-6) to 2,600 x 10(-6), the two highest values corresponding to the c-erbB-2 overexpressing cells MDA-MB-453 and SK-BR-3. In a series of 100 breast cancer samples, c-erbB-2 normalized expression was found to range from 0.4 x 10(-6) to 350 x 10(-6). A close correlation was observed between this real-time one-step quantitative RT-PCR method and both semiquantitative conventional RT-PCR (N = 22; r = 0.8543; P < .0001) and c-erbB-2 protein expression (p185) quantified by an enzyme immunoassay (EIA) (N = 27; r = 0.71; P < .0001). The current realtime RT-PCR assay is rapid, sensitive, and reproducible and appears particularly suitable to quantify gene expression in large series of samples.

  6. Using PCR to Target Misconceptions about Gene Expression

    PubMed Central

    Wright, Leslie K.; Newman, Dina L.

    2013-01-01

    We present a PCR-based laboratory exercise that can be used with first- or second-year biology students to help overcome common misconceptions about gene expression. Biology students typically do not have a clear understanding of the difference between genes (DNA) and gene expression (mRNA/protein) and often believe that genes exist in an organism or cell only when they are expressed. This laboratory exercise allows students to carry out a PCR-based experiment designed to challenge their misunderstanding of the difference between genes and gene expression. Students first transform E. coli with an inducible GFP gene containing plasmid and observe induced and un-induced colonies. The following exercise creates cognitive dissonance when actual PCR results contradict their initial (incorrect) predictions of the presence of the GFP gene in transformed cells. Field testing of this laboratory exercise resulted in learning gains on both knowledge and application questions on concepts related to genes and gene expression. PMID:23858358

  7. Inferring gene expression dynamics via functional regression analysis

    PubMed Central

    Müller, Hans-Georg; Chiou, Jeng-Min; Leng, Xiaoyan

    2008-01-01

    Background Temporal gene expression profiles characterize the time-dynamics of expression of specific genes and are increasingly collected in current gene expression experiments. In the analysis of experiments where gene expression is obtained over the life cycle, it is of interest to relate temporal patterns of gene expression associated with different developmental stages to each other to study patterns of long-term developmental gene regulation. We use tools from functional data analysis to study dynamic changes by relating temporal gene expression profiles of different developmental stages to each other. Results We demonstrate that functional regression methodology can pinpoint relationships that exist between temporary gene expression profiles for different life cycle phases and incorporates dimension reduction as needed for these high-dimensional data. By applying these tools, gene expression profiles for pupa and adult phases are found to be strongly related to the profiles of the same genes obtained during the embryo phase. Moreover, one can distinguish between gene groups that exhibit relationships with positive and others with negative associations between later life and embryonal expression profiles. Specifically, we find a positive relationship in expression for muscle development related genes, and a negative relationship for strictly maternal genes for Drosophila, using temporal gene expression profiles. Conclusion Our findings point to specific reactivation patterns of gene expression during the Drosophila life cycle which differ in characteristic ways between various gene groups. Functional regression emerges as a useful tool for relating gene expression patterns from different developmental stages, and avoids the problems with large numbers of parameters and multiple testing that affect alternative approaches. PMID:18226220

  8. TNF-α gene polymorphisms and expression.

    PubMed

    El-Tahan, Radwa R; Ghoneim, Ahmed M; El-Mashad, Noha

    2016-01-01

    Tumor necrosis factor alpha (TNF-α) is a proinflammatory cytokine with an important role in the pathogenesis of several diseases. Its encoding gene is located in the short arm of chromosome 6 in the major histocompatibility complex class III region. Most of the TNF-α gene polymorphisms are located in its promoter region and they are thought to affect the susceptibility and/or severity of different human diseases. This review summarizes the data related to the association between TNF-α gene and its receptor polymorphisms, and the development of autoimmune diseases. Among these polymorphisms the -308G/A TNF-α promotor polymorphism has been associated several times with the the development of autoimmune diseases, however some discrepant results have been recorded. The other TNF-α gene polymorphisms had little or no association with autoimmune diseases. Current results about the molecules controlling TNF-α expression are also presented. The discrepancy between different records could be related partly to either the differences in the ethnic origin or number of the studied individuals, or the abundance and activation of other molecules that interact with the TNF-α promotor region or other elements. PMID:27652081

  9. Hyperbaric oxygen treatment induces antioxidant gene expression.

    PubMed

    Godman, Cassandra A; Joshi, Rashmi; Giardina, Charles; Perdrizet, George; Hightower, Lawrence E

    2010-06-01

    Although the underlying molecular causes of aging are not entirely clear, hormetic agents like exercise, heat, and calorie restriction may generate a mild pro-oxidant stress that induces cell protective responses to promote healthy aging. As an individual ages, many cellular and physiological processes decline, including wound healing and reparative angiogenesis. This is particularly critical in patients with chronic non-healing wounds who tend to be older. We are interested in the potential beneficial effects of hyperbaric oxygen as a mild hormetic stress on human microvascular endothelial cells. We analyzed global gene expression changes in human endothelial cells following a hyperbaric exposure comparable to a clinical treatment. Our analysis revealed an upregulation of antioxidant, cytoprotective, and immediate early genes. This increase coincided with an increased resistance to a lethal oxidative stress. Our data indicate that hyperbaric oxygen can induce protection against oxidative insults in endothelial cells and may provide an easily administered hormetic treatment to help promote healthy aging.

  10. Expressing exogenous genes in newts by transgenesis.

    PubMed

    Casco-Robles, Martin Miguel; Yamada, Shouta; Miura, Tomoya; Nakamura, Kenta; Haynes, Tracy; Maki, Nobuyasu; Del Rio-Tsonis, Katia; Tsonis, Panagiotis A; Chiba, Chikafumi

    2011-05-01

    The great regenerative abilities of newts provide the impetus for studies at the molecular level. However, efficient methods for gene regulation have historically been quite limited. Here we describe a protocol for transgenically expressing exogenous genes in the newt Cynops pyrrhogaster. This method is simple: a reaction mixture of I-SceI meganuclease and a plasmid DNA carrying a transgene cassette flanked by the enzyme recognition sites is directly injected into fertilized eggs. The protocol achieves a high efficiency of transgenesis, comparable to protocols used in other animal systems, and it provides a practical number of transgenic newts (∼20% of injected embryos) that survive beyond metamorphosis and that can be applied to regenerative studies. The entire protocol for obtaining transgenic adult newts takes 4-5 months.

  11. Retrotransposons as regulators of gene expression.

    PubMed

    Elbarbary, Reyad A; Lucas, Bronwyn A; Maquat, Lynne E

    2016-02-12

    Transposable elements (TEs) are both a boon and a bane to eukaryotic organisms, depending on where they integrate into the genome and how their sequences function once integrated. We focus on two types of TEs: long interspersed elements (LINEs) and short interspersed elements (SINEs). LINEs and SINEs are retrotransposons; that is, they transpose via an RNA intermediate. We discuss how LINEs and SINEs have expanded in eukaryotic genomes and contribute to genome evolution. An emerging body of evidence indicates that LINEs and SINEs function to regulate gene expression by affecting chromatin structure, gene transcription, pre-mRNA processing, or aspects of mRNA metabolism. We also describe how adenosine-to-inosine editing influences SINE function and how ongoing retrotransposition is countered by the body's defense mechanisms.

  12. Approaches for gene targeting and targeted gene expression in plants.

    PubMed

    Husaini, Amjad Masood; Rashid, Zerka; Mir, Reyaz-ul Rouf; Aquil, Bushra

    2011-01-01

    Transgenic science and technology are fundamental to state-of-the-art plant molecular genetics and crop improvement. The new generation of technology endeavors to introduce genes 'stably' into 'site-specific' locations and in 'single copy' without the integration of extraneous vector 'backbone' sequences or selectable markers and with a 'predictable and consistent' expression. Several similar strategies and technologies, which can push the development of 'smart' genetically modified plants with desirable attributes, as well as enhance their consumer acceptability, are discussed in this review.

  13. Pathway network inference from gene expression data

    PubMed Central

    2014-01-01

    Background The development of high-throughput omics technologies enabled genome-wide measurements of the activity of cellular elements and provides the analytical resources for the progress of the Systems Biology discipline. Analysis and interpretation of gene expression data has evolved from the gene to the pathway and interaction level, i.e. from the detection of differentially expressed genes, to the establishment of gene interaction networks and the identification of enriched functional categories. Still, the understanding of biological systems requires a further level of analysis that addresses the characterization of the interaction between functional modules. Results We present a novel computational methodology to study the functional interconnections among the molecular elements of a biological system. The PANA approach uses high-throughput genomics measurements and a functional annotation scheme to extract an activity profile from each functional block -or pathway- followed by machine-learning methods to infer the relationships between these functional profiles. The result is a global, interconnected network of pathways that represents the functional cross-talk within the molecular system. We have applied this approach to describe the functional transcriptional connections during the yeast cell cycle and to identify pathways that change their connectivity in a disease condition using an Alzheimer example. Conclusions PANA is a useful tool to deepen in our understanding of the functional interdependences that operate within complex biological systems. We show the approach is algorithmically consistent and the inferred network is well supported by the available functional data. The method allows the dissection of the molecular basis of the functional connections and we describe the different regulatory mechanisms that explain the network's topology obtained for the yeast cell cycle data. PMID:25032889

  14. Regulation of gene expression by hypoxia.

    PubMed

    Kenneth, Niall Steven; Rocha, Sonia

    2008-08-15

    Hypoxia induces profound changes in the cellular gene expression profile. The discovery of a major transcription factor family activated by hypoxia, HIF (hypoxia-inducible factor), and the factors that contribute to HIF regulation have greatly enhanced our knowledge of the molecular aspects of the hypoxic response. However, in addition to HIF, other transcription factors and cellular pathways are activated by exposure to reduced oxygen. In the present review, we summarize the current knowledge of how additional hypoxia-responsive transcription factors integrate with HIF and how other cellular pathways such as chromatin remodelling, translation regulation and microRNA induction, contribute to the co-ordinated cellular response observed following hypoxic stress.

  15. Network Completion for Static Gene Expression Data

    PubMed Central

    Nakajima, Natsu

    2014-01-01

    We tackle the problem of completing and inferring genetic networks under stationary conditions from static data, where network completion is to make the minimum amount of modifications to an initial network so that the completed network is most consistent with the expression data in which addition of edges and deletion of edges are basic modification operations. For this problem, we present a new method for network completion using dynamic programming and least-squares fitting. This method can find an optimal solution in polynomial time if the maximum indegree of the network is bounded by a constant. We evaluate the effectiveness of our method through computational experiments using synthetic data. Furthermore, we demonstrate that our proposed method can distinguish the differences between two types of genetic networks under stationary conditions from lung cancer and normal gene expression data. PMID:24826192

  16. Gene Expression During the Life Cycle of Drosophila melanogaster

    NASA Astrophysics Data System (ADS)

    Arbeitman, Michelle N.; Furlong, Eileen E. M.; Imam, Farhad; Johnson, Eric; Null, Brian H.; Baker, Bruce S.; Krasnow, Mark A.; Scott, Matthew P.; Davis, Ronald W.; White, Kevin P.

    2002-09-01

    Molecular genetic studies of Drosophila melanogaster have led to profound advances in understanding the regulation of development. Here we report gene expression patterns for nearly one-third of all Drosophila genes during a complete time course of development. Mutations that eliminate eye or germline tissue were used to further analyze tissue-specific gene expression programs. These studies define major characteristics of the transcriptional programs that underlie the life cycle, compare development in males and females, and show that large-scale gene expression data collected from whole animals can be used to identify genes expressed in particular tissues and organs or genes involved in specific biological and biochemical processes.

  17. Regulatory Divergence between Parental Alleles Determines Gene Expression Patterns in Hybrids

    PubMed Central

    Combes, Marie-Christine; Hueber, Yann; Dereeper, Alexis; Rialle, Stéphanie; Herrera, Juan-Carlos; Lashermes, Philippe

    2015-01-01

    Both hybridization and allopolyploidization generate novel phenotypes by conciliating divergent genomes and regulatory networks in the same cellular context. To understand the rewiring of gene expression in hybrids, the total expression of 21,025 genes and the allele-specific expression of over 11,000 genes were quantified in interspecific hybrids and their parental species, Coffea canephora and Coffea eugenioides using RNA-seq technology. Between parental species, cis- and trans-regulatory divergences affected around 32% and 35% of analyzed genes, respectively, with nearly 17% of them showing both. The relative importance of trans-regulatory divergences between both species could be related to their low genetic divergence and perennial habit. In hybrids, among divergently expressed genes between parental species and hybrids, 77% was expressed like one parent (expression level dominance), including 65% like C. eugenioides. Gene expression was shown to result from the expression of both alleles affected by intertwined parental trans-regulatory factors. A strong impact of C. eugenioides trans-regulatory factors on the upregulation of C. canephora alleles was revealed. The gene expression patterns appeared determined by complex combinations of cis- and trans-regulatory divergences. In particular, the observed biased expression level dominance seemed to be derived from the asymmetric effects of trans-regulatory parental factors on regulation of alleles. More generally, this study illustrates the effects of divergent trans-regulatory parental factors on the gene expression pattern in hybrids. The characteristics of the transcriptional response to hybridization appear to be determined by the compatibility of gene regulatory networks and therefore depend on genetic divergences between the parental species and their evolutionary history. PMID:25819221

  18. Streptomyces coelicolor as an expression host for heterologous gene clusters.

    PubMed

    Gomez-Escribano, Juan Pablo; Bibb, Mervyn J

    2012-01-01

    The expression of a gene or a set of genes from one organism in a different species is known as "heterologous expression." In actinomycetes, prolific producers of natural products, heterologous gene expression has been used to confirm the clustering of secondary metabolite biosynthetic genes, to analyze natural product biosynthesis, to produce variants of natural products by genetic engineering, and to discover new compounds by screening genomic libraries. Recent advances in DNA sequencing have enabled the rapid and affordable sequencing of actinomycete genomes and revealed a large number of secondary metabolite gene clusters with no known products. Heterologous expression of these cryptic gene clusters combined with comparative metabolic profiling provides an important means to identify potentially novel compounds. In this chapter, the methods and strategies used to heterologously express actinomycete gene clusters, including the techniques used for cloning secondary metabolite gene clusters, the Streptomyces hosts used for their expression, and the techniques employed to analyze their products by metabolic profiling, are described.

  19. Quantifying the constraining influence of gene flow on adaptive divergence in the lake-stream threespine stickleback system.

    PubMed

    Moore, Jean-Sébastien; Gow, Jennifer L; Taylor, Eric B; Hendry, Andrew P

    2007-08-01

    The constraining effect of gene flow on adaptive divergence is often inferred but rarely quantified. We illustrate ways of doing so using stream populations of threespine stickleback (Gasterosteus aculeatus) that experience different levels of gene flow from a parapatric lake population. In the Misty Lake watershed (British Columbia, Canada), the inlet stream population is morphologically divergent from the lake population, and presumably experiences little gene flow from the lake. The outlet stream population, however, shows an intermediate phenotype and may experience more gene flow from the lake. We first used microsatellite data to demonstrate that gene flow from the lake is low into the inlet but high into the outlet, and that gene flow from the lake remains relatively constant with distance along the outlet. We next combined gene flow data with morphological and habitat data to quantify the effect of gene flow on morphological divergence. In one approach, we assumed that inlet stickleback manifest well-adapted phenotypic trait values not constrained by gene flow. We then calculated the deviation between the observed and expected phenotypes for a given habitat in the outlet. In a second approach, we parameterized a quantitative genetic model of adaptive divergence. Both approaches suggest a large impact of gene flow, constraining adaptation by 80-86% in the outlet (i.e., only 14-20% of the expected morphological divergence in the absence of gene flow was observed). Such approaches may be useful in other taxa to estimate how important gene flow is in constraining adaptive divergence in nature. PMID:17683442

  20. Neuroinformatics for genome-wide 3D gene expression mapping in the mouse brain.

    PubMed

    Ng, Lydia; Pathak, Sayan D; Kuan, Chihchau; Lau, Chris; Dong, Hongwei; Sodt, Andrew; Dang, Chinh; Avants, Brian; Yushkevich, Paul; Gee, James C; Haynor, David; Lein, Ed; Jones, Allan; Hawrylycz, Mike

    2007-01-01

    Large scale gene expression studies in the mammalian brain offer the promise of understanding the topology, networks and ultimately the function of its complex anatomy, opening previously unexplored avenues in neuroscience. High-throughput methods permit genome-wide searches to discover genes that are uniquely expressed in brain circuits and regions that control behavior. Previous gene expression mapping studies in model organisms have employed situ hybridization (ISH), a technique that uses labeled nucleic acid probes to bind to specific mRNA transcripts in tissue sections. A key requirement for this effort is the development of fast and robust algorithms for anatomically mapping and quantifying gene expression for ISH. We describe a neuroinformatics pipeline for automatically mapping expression profiles of ISH data and its use to produce the first genomic scale 3-D mapping of gene expression in a mammalian brain. The pipeline is fully automated and adaptable to other organisms and tissues. Our automated study of over 20,000 genes indicates that at least 78.8 percent are expressed at some level in the adult C56BL/6J mouse brain. In addition to providing a platform for genomic scale search, high-resolution images and visualization tools for expression analysis are available at the Allen Brain Atlas web site (http://www.brain-map.org).

  1. Obesity alters the expression profile of clock genes in peripheral blood mononuclear cells

    PubMed Central

    Tahira, Kazunobu; Fukuda, Noboru; Aoyama, Takahiko; Tsunemi, Akiko; Matsumoto, Siroh; Nagura, Chinami; Matsumoto, Taro; Soma, Masayoshi; Shimba, Shigeki; Matsumoto, Yoshiaki

    2011-01-01

    Introduction The aim of this study was to investigate the association between the variation in expression profile of clock genes and obesity using peripheral blood mononuclear (PMN) cells. Material and methods The subjects comprised 10 obese patients and 10 healthy volunteers. Blood was collected at different time-points during the day and levels of blood sugar, IRI, adiponectin and leptin were determined. Peripheral blood mononuclear cells were sampled, and expression levels of brain and muscle Arnt-like protein-1 (BMAL1), Period (PER)1, PER2, Cryptochrome (CRY)1, CRY2, and REV-ERBα mRNA were quantified. Results During the day, the expression levels of BMAL1, CRY1, CRY2 and PER2 genes in PMN cells of the obese group were all significantly higher compared to those in the non-obese group. In addition, expression of BMAL1, CRY1, CRY2 and PER2 genes in PMN cells increased between 12:00 and 21:00 in the obese group. In PMN cells of both groups, PER1 gene expression showed a bimodal pattern, with high expression at 9:00 and 18:00. Conclusions Differences were observed in the expression profile variation of clock genes between the obese and non-obese groups. This study reveals the differences in clock gene expression profiles between obese and non-obese subjects, with evidence for two distinct chronotypes, and suggests a contribution of these chronotypes to fat accumulation in humans. PMID:22328874

  2. Live-cell, temporal gene expression analysis of osteogenic differentiation in adipose-derived stem cells.

    PubMed

    Desai, Hetal V; Voruganti, Indu S; Jayasuriya, Chathuraka; Chen, Qian; Darling, Eric M

    2014-03-01

    Adipose-derived stem cells (ASCs) are a widely investigated type of mesenchymal stem cells with great potential for musculoskeletal regeneration. However, the use of ASCs is complicated by their cellular heterogeneity, which exists at both the population and single-cell levels. This study demonstrates a live-cell assay to investigate gene expression in ASCs undergoing osteogenesis using fluorescently tagged DNA hybridization probes called molecular beacons. A molecular beacon was designed to target the mRNA sequence for alkaline phosphatase (ALPL), a gene characteristically expressed during early osteogenesis. The percentage of cells expressing this gene in a population was monitored daily to quantify the uniformity of the differentiation process. Differentiating ASC populations were repeatedly measured in a nondestructive fashion over a 10-day period to obtain temporal gene expression data. Results showed consistent expression patterns for the investigated osteogenic genes in response to induction medium. Peak signal level, indicating when the most cells expressed ALPL at once, was observed on days 3-5. The differentiation response of sample populations was generally uniform when assessed on a well-by-well basis over time. The expression of alkaline phosphatase is consistent with previous studies of osteogenic differentiation, suggesting that molecular beacons are a viable means of monitoring the spatiotemporal gene expression of live, differentiating ASCs.

  3. Live-Cell, Temporal Gene Expression Analysis of Osteogenic Differentiation in Adipose-Derived Stem Cells

    PubMed Central

    Desai, Hetal V.; Voruganti, Indu S.; Jayasuriya, Chathuraka; Chen, Qian

    2014-01-01

    Adipose-derived stem cells (ASCs) are a widely investigated type of mesenchymal stem cells with great potential for musculoskeletal regeneration. However, the use of ASCs is complicated by their cellular heterogeneity, which exists at both the population and single-cell levels. This study demonstrates a live-cell assay to investigate gene expression in ASCs undergoing osteogenesis using fluorescently tagged DNA hybridization probes called molecular beacons. A molecular beacon was designed to target the mRNA sequence for alkaline phosphatase (ALPL), a gene characteristically expressed during early osteogenesis. The percentage of cells expressing this gene in a population was monitored daily to quantify the uniformity of the differentiation process. Differentiating ASC populations were repeatedly measured in a nondestructive fashion over a 10-day period to obtain temporal gene expression data. Results showed consistent expression patterns for the investigated osteogenic genes in response to induction medium. Peak signal level, indicating when the most cells expressed ALPL at once, was observed on days 3–5. The differentiation response of sample populations was generally uniform when assessed on a well-by-well basis over time. The expression of alkaline phosphatase is consistent with previous studies of osteogenic differentiation, suggesting that molecular beacons are a viable means of monitoring the spatiotemporal gene expression of live, differentiating ASCs. PMID:24367991

  4. Imaging gene expression in real-time using aptamers

    SciTech Connect

    Shin, Ilchung

    2012-01-01

    Signal transduction pathways are usually activated by external stimuli and are transient. The downstream changes such as transcription of the activated genes are also transient. Real-time detection of promoter activity is useful for understanding changes in gene expression, especially during cell differentiation and in development. A simple and reliable method for viewing gene expression in real time is not yet available. Reporter proteins such as fluorescent proteins and luciferase allow for non-invasive detection of the products of gene expression in living cells. However, current reporter systems do not provide for real-time imaging of promoter activity in living cells. This is because of the long time period after transcription required for fluorescent protein synthesis and maturation. We have developed an RNA reporter system for imaging in real-time to detect changes in promoter activity as they occur. The RNA reporter uses strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags), which can be expressed from a promoter of choice. The tobramycin, neomycin and PDC RNA aptamers have been utilized for this system and expressed in yeast from the GAL1 promoter. The IMAGEtag RNA kinetics were quantified by RT-qPCR. In yeast precultured in raffinose containing media the GAL1 promoter responded faster than in yeast precultured in glucose containing media. IMAGEtag RNA has relatively short half-life (5.5 min) in yeast. For imaging, the yeast cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal to noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after activation of the promoter by galactose. FRET was confirmed with three different approaches, which were sensitized emission, acceptor photobleaching and donor lifetime by FLIM (fluorescence lifetime imaging

  5. Imaging gene expression in real-time using aptamers

    SciTech Connect

    Shin, Il Chung

    2011-01-01

    Signal transduction pathways are usually activated by external stimuli and are transient. The downstream changes such as transcription of the activated genes are also transient. Real-time detection of promoter activity is useful for understanding changes in gene expression, especially during cell differentiation and in development. A simple and reliable method for viewing gene expression in real time is not yet available. Reporter proteins such as fluorescent proteins and luciferase allow for non-invasive detection of the products of gene expression in living cells. However, current reporter systems do not provide for real-time imaging of promoter activity in living cells. This is because of the long time period after transcription required for fluorescent protein synthesis and maturation. We have developed an RNA reporter system for imaging in real-time to detect changes in promoter activity as they occur. The RNA reporter uses strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags), which can be expressed from a promoter of choice. The tobramycin, neomycin and PDC RNA aptamers have been utilized for this system and expressed in yeast from the GAL1 promoter. The IMAGEtag RNA kinetics were quantified by RT-qPCR. In yeast precultured in raffinose containing media the GAL1 promoter responded faster than in yeast precultured in glucose containing media. IMAGEtag RNA has relatively short half-life (5.5 min) in yeast. For imaging, the yeast cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal to noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after activation of the promoter by galactose. FRET was confirmed with three different approaches, which were sensitized emission, acceptor photobleaching and donor lifetime by FLIM (fluorescence lifetime imaging

  6. Repeatability of published microarray gene expression analyses.

    PubMed

    Ioannidis, John P A; Allison, David B; Ball, Catherine A; Coulibaly, Issa; Cui, Xiangqin; Culhane, Aedín C; Falchi, Mario; Furlanello, Cesare; Game, Laurence; Jurman, Giuseppe; Mangion, Jon; Mehta, Tapan; Nitzberg, Michael; Page, Grier P; Petretto, Enrico; van Noort, Vera

    2009-02-01

    Given the complexity of microarray-based gene expression studies, guidelines encourage transparent design and public data availability. Several journals require public data deposition and several public databases exist. However, not all data are publicly available, and even when available, it is unknown whether the published results are reproducible by independent scientists. Here we evaluated the replication of data analyses in 18 articles on microarray-based gene expression profiling published in Nature Genetics in 2005-2006. One table or figure from each article was independently evaluated by two teams of analysts. We reproduced two analyses in principle and six partially or with some discrepancies; ten could not be reproduced. The main reason for failure to reproduce was data unavailability, and discrepancies were mostly due to incomplete data annotation or specification of data processing and analysis. Repeatability of published microarray studies is apparently limited. More strict publication rules enforcing public data availability and explicit description of data processing and analysis should be considered.

  7. Regulation of gene expression by hypoxia.

    PubMed

    Millhorn, D E; Czyzyk-Krzeska, M; Bayliss, D A; Lawson, E E

    1993-12-01

    The present study was undertaken to determine if gene expression for tyrosine hydroxylase (TH), the rate limiting enzyme in the biosynthesis of catecholamines, is regulated in the carotid body, sympathetic ganglia and adrenal medulla by hypoxia. We found that a reduction in oxygen tension from 21% to 10% caused a substantial increase (200% at 1 hour and 500% at 6 hours exposure) in the concentration of TH mRNA in carotid body type I cells but not in either the sympathetic ganglia or adrenal gland. In addition, we found that hypercapnia, another natural stimulus of carotid body activity, failed to enhance TH mRNA in type I cells. Removal of the sensory and sympathetic innervation of the carotid body failed to prevent the induction of TH mRNA by hypoxia in type I cells. Our results show that TH gene expression is regulated by hypoxia in the carotid body but not in other peripheral catecholamine synthesizing tissue and that the regulatory mechanism is intrinsic to type I cells. PMID:7909954

  8. Insulin-glycerolipid mediators and gene expression

    SciTech Connect

    Standaert, M.L.; Pollet, R.J. )

    1988-06-01

    Insulin is an anabolic polypeptide hormone with pleiotrophic effects. During the decades since the initial description by Banting and Best, the acute effects of insulin have been widely studied with particular focus on the mechanism or mechanisms of insulin activation of hexose transport and regulation of metabolic enzyme activity. However, recently there has been a major expansion of investigation to include insulin regulation of gene expression with multiple insulin-sensitive specific mRNAs now reported. In this review, we explore the involvement of insulin-induced changes in plasma membrane glycerolipid metabolism in the transmembrane signaling process required for insulin regulation of mRNA levels. Insulin increase diacylglycerol levels in insulin-responsive cells, and synthetic diacylglycerols or their phorbol ester diacylglycerol analogs, such as 4{beta}, 9{alpha}, 12{beta}, 13{alpha}, 20-pentahydroxytiglia-1,6-dien-3-one 12{beta}-myristate 13-acetate (TPA), mimic insulin regulation of ornithine decarboxylase mRNA, c-fos mRNA, and phosphoenolpyruvate carboxykinase mRNA levels. This suggests that insulin regulation of specific mRNA levels may be mediated by insulin-induced changes in phospholipid metabolism and that diacylglycerol may play a pivotal role in insulin regulation of gene expression.

  9. Gene transfer and expression in plants.

    PubMed

    Lorence, Argelia; Verpoorte, Robert

    2004-01-01

    Until recently, agriculture and plant breeding relied solely on the accumulated experience of generations of farmers and breeders that is, on sexual transfer of genes between plant species. However, recent developments in plant molecular biology and genomics now give us access to knowledge and understanding of plant genomes and the possibility of modifying them. This chapter presents an updated overview of the two most powerful technologies for transferring genetic material (DNA) into plants: Agrobacterium-mediated transformation and microparticle bombardment (biolistics). Some of the topics that are discussed in detail are the main variables controlling the transformation efficiency that can be achieved using each one of these approaches; the advantages and limitations of each methodology; transient versus stable transformation approaches; the potential of some in planta transformation systems; alternatives to developing transgenic plants without selection markers; the availability of diverse genetic tools generated as part of the genome sequencing of different plant species; transgene expression, gene silencing, and their association with regulatory elements; and prospects and ways to possibly overcome some transgene expression difficulties, in particular the use of matrix-attachment regions (MARs).

  10. Coactivators in PPAR-Regulated Gene Expression

    PubMed Central

    Viswakarma, Navin; Jia, Yuzhi; Bai, Liang; Vluggens, Aurore; Borensztajn, Jayme; Xu, Jianming; Reddy, Janardan K.

    2010-01-01

    Peroxisome proliferator-activated receptor (PPAR)α, β (also known as δ), and γ function as sensors for fatty acids and fatty acid derivatives and control important metabolic pathways involved in the maintenance of energy balance. PPARs also regulate other diverse biological processes such as development, differentiation, inflammation, and neoplasia. In the nucleus, PPARs exist as heterodimers with retinoid X receptor-α bound to DNA with corepressor molecules. Upon ligand activation, PPARs undergo conformational changes that facilitate the dissociation of corepressor molecules and invoke a spatiotemporally orchestrated recruitment of transcription cofactors including coactivators and coactivator-associated proteins. While a given nuclear receptor regulates the expression of a prescribed set of target genes, coactivators are likely to influence the functioning of many regulators and thus affect the transcription of many genes. Evidence suggests that some of the coactivators such as PPAR-binding protein (PBP/PPARBP), thyroid hormone receptor-associated protein 220 (TRAP220), and mediator complex subunit 1 (MED1) may exert a broader influence on the functions of several nuclear receptors and their target genes. Investigations into the role of coactivators in the function of PPARs should strengthen our understanding of the complexities of metabolic diseases associated with energy metabolism. PMID:20814439

  11. Posttranscriptional Control of Gene Expression in Yeast

    PubMed Central

    McCarthy, John E. G.

    1998-01-01

    Studies of the budding yeast Saccharomyces cerevisiae have greatly advanced our understanding of the posttranscriptional steps of eukaryotic gene expression. Given the wide range of experimental tools applicable to S. cerevisiae and the recent determination of its complete genomic sequence, many of the key challenges of the posttranscriptional control field can be tackled particularly effectively by using this organism. This article reviews the current knowledge of the cellular components and mechanisms related to translation and mRNA decay, with the emphasis on the molecular basis for rate control and gene regulation. Recent progress in characterizing translation factors and their protein-protein and RNA-protein interactions has been rapid. Against the background of a growing body of structural information, the review discusses the thermodynamic and kinetic principles that govern the translation process. As in prokaryotic systems, translational initiation is a key point of control. Modulation of the activities of translational initiation factors imposes global regulation in the cell, while structural features of particular 5′ untranslated regions, such as upstream open reading frames and effector binding sites, allow for gene-specific regulation. Recent data have revealed many new details of the molecular mechanisms involved while providing insight into the functional overlaps and molecular networking that are apparently a key feature of evolving cellular systems. An overall picture of the mechanisms governing mRNA decay has only very recently begun to develop. The latest work has revealed new information about the mRNA decay pathways, the components of the mRNA degradation machinery, and the way in which these might relate to the translation apparatus. Overall, major challenges still to be addressed include the task of relating principles of posttranscriptional control to cellular compartmentalization and polysome structure and the role of molecular channelling

  12. Predicting cellular growth from gene expression signatures.

    PubMed

    Airoldi, Edoardo M; Huttenhower, Curtis; Gresham, David; Lu, Charles; Caudy, Amy A; Dunham, Maitreya J; Broach, James R; Botstein, David; Troyanskaya, Olga G

    2009-01-01

    Maintaining balanced growth in a changing environment is a fundamental systems-level challenge for cellular physiology, particularly in microorganisms. While the complete set of regulatory and functional pathways supporting growth and cellular proliferation are not yet known, portions of them are well understood. In particular, cellular proliferation is governed by mechanisms that are highly conserved from unicellular to multicellular organisms, and the disruption of these processes in metazoans is a major factor in the development of cancer. In this paper, we develop statistical methodology to identify quantitative aspects of the regulatory mechanisms underlying cellular proliferation in Saccharomyces cerevisiae. We find that the expression levels of a small set of genes can be exploited to predict the instantaneous growth rate of any cellular culture with high accuracy. The predictions obtained in this fashion are robust to changing biological conditions, experimental methods, and technological platforms. The proposed model is also effective in predicting growth rates for the related yeast Saccharomyces bayanus and the highly diverged yeast Schizosaccharomyces pombe, suggesting that the underlying regulatory signature is conserved across a wide range of unicellular evolution. We investigate the biological significance of the gene expression signature that the predictions are based upon from multiple perspectives: by perturbing the regulatory network through the Ras/PKA pathway, observing strong upregulation of growth rate even in the absence of appropriate nutrients, and discovering putative transcription factor binding sites, observing enrichment in growth-correlated genes. More broadly, the proposed methodology enables biological insights about growth at an instantaneous time scale, inaccessible by direct experimental methods. Data and tools enabling others to apply our methods are available at http://function.princeton.edu/growthrate.

  13. Social regulation of cortisol receptor gene expression

    PubMed Central

    Korzan, Wayne J.; Grone, Brian P.; Fernald, Russell D.

    2014-01-01

    In many social species, individuals influence the reproductive capacity of conspecifics. In a well-studied African cichlid fish species, Astatotilapia burtoni, males are either dominant (D) and reproductively competent or non-dominant (ND) and reproductively suppressed as evidenced by reduced gonadotropin releasing hormone (GnRH1) release, regressed gonads, lower levels of androgens and elevated levels of cortisol. Here, we asked whether androgen and cortisol levels might regulate this reproductive suppression. Astatotilapia burtoni has four glucocorticoid receptors (GR1a, GR1b, GR2 and MR), encoded by three genes, and two androgen receptors (ARα and ARβ), encoded by two genes. We previously showed that ARα and ARβ are expressed in GnRH1 neurons in the preoptic area (POA), which regulates reproduction, and that the mRNA levels of these receptors are regulated by social status. Here, we show that GR1, GR2 and MR mRNAs are also expressed in GnRH1 neurons in the POA, revealing potential mechanisms for both androgens and cortisol to influence reproductive capacity. We measured AR, MR and GR mRNA expression levels in a microdissected region of the POA containing GnRH1 neurons, comparing D and ND males. Using quantitative PCR (qPCR), we found D males had higher mRNA levels of ARα, MR, total GR1a and GR2 in the POA compared with ND males. In contrast, ND males had significantly higher levels of GR1b mRNA, a receptor subtype with a reduced transcriptional response to cortisol. Through this novel regulation of receptor type, neurons in the POA of an ND male will be less affected by the higher levels of cortisol typical of low status, suggesting GR receptor type change as a potential adaptive mechanism to mediate high cortisol levels during social suppression. PMID:25013108

  14. Identification of suitable reference genes for gene expression normalization in qRT-PCR analysis in watermelon.

    PubMed

    Kong, Qiusheng; Yuan, Jingxian; Gao, Lingyun; Zhao, Shuang; Jiang, Wei; Huang, Yuan; Bie, Zhilong

    2014-01-01

    Watermelon is one of the major Cucurbitaceae crops and the recent availability of genome sequence greatly facilitates the fundamental researches on it. Quantitative real-time reverse transcriptase PCR (qRT-PCR) is the preferred method for gene expression analyses, and using validated reference genes for normalization is crucial to ensure the accuracy of this method. However, a systematic validation of reference genes has not been conducted on watermelon. In this study, transcripts of 15 candidate reference genes were quantified in watermelon using qRT-PCR, and the stability of these genes was compared using geNorm and NormFinder. geNorm identified ClTUA and ClACT, ClEF1α and ClACT, and ClCAC and ClTUA as the best pairs of reference genes in watermelon organs and tissues under normal growth conditions, abiotic stress, and biotic stress, respectively. NormFinder identified ClYLS8, ClUBCP, and ClCAC as the best single reference genes under the above experimental conditions, respectively. ClYLS8 and ClPP2A were identified as the best reference genes across all samples. Two to nine reference genes were required for more reliable normalization depending on the experimental conditions. The widely used watermelon reference gene 18SrRNA was less stable than the other reference genes under the experimental conditions. Catalase family genes were identified in watermelon genome, and used to validate the reliability of the identified reference genes. ClCAT1and ClCAT2 were induced and upregulated in the first 24 h, whereas ClCAT3 was downregulated in the leaves under low temperature stress. However, the expression levels of these genes were significantly overestimated and misinterpreted when 18SrRNA was used as a reference gene. These results provide a good starting point for reference gene selection in qRT-PCR analyses involving watermelon.

  15. Identification of Suitable Reference Genes for Gene Expression Normalization in qRT-PCR Analysis in Watermelon

    PubMed Central

    Gao, Lingyun; Zhao, Shuang; Jiang, Wei; Huang, Yuan; Bie, Zhilong

    2014-01-01

    Watermelon is one of the major Cucurbitaceae crops and the recent availability of genome sequence greatly facilitates the fundamental researches on it. Quantitative real-time reverse transcriptase PCR (qRT–PCR) is the preferred method for gene expression analyses, and using validated reference genes for normalization is crucial to ensure the accuracy of this method. However, a systematic validation of reference genes has not been conducted on watermelon. In this study, transcripts of 15 candidate reference genes were quantified in watermelon using qRT–PCR, and the stability of these genes was compared using geNorm and NormFinder. geNorm identified ClTUA and ClACT, ClEF1α and ClACT, and ClCAC and ClTUA as the best pairs of reference genes in watermelon organs and tissues under normal growth conditions, abiotic stress, and biotic stress, respectively. NormFinder identified ClYLS8, ClUBCP, and ClCAC as the best single reference genes under the above experimental conditions, respectively. ClYLS8 and ClPP2A were identified as the best reference genes across all samples. Two to nine reference genes were required for more reliable normalization depending on the experimental conditions. The widely used watermelon reference gene 18SrRNA was less stable than the other reference genes under the experimental conditions. Catalase family genes were identified in watermelon genome, and used to validate the reliability of the identified reference genes. ClCAT1and ClCAT2 were induced and upregulated in the first 24 h, whereas ClCAT3 was downregulated in the leaves under low temperature stress. However, the expression levels of these genes were significantly overestimated and misinterpreted when 18SrRNA was used as a reference gene. These results provide a good starting point for reference gene selection in qRT–PCR analyses involving watermelon. PMID:24587403

  16. T gene isoform expression pattern is significantly different between chordomas and notochords.

    PubMed

    Wang, Ke; Hu, Qingtao; Wang, Liang; Chen, Wei; Tian, Kaibing; Cao, Chunwei; Wu, Zhen; Jia, Guijun; Zhang, Liwei; Zeng, Changqing; Zhang, Junting

    2015-11-13

    The T gene plays a key role in chordoma pathology. To investigate the role of T gene isoforms in chordoma, 22 skull base chordomas, three chordoma cell lines and 9 infant notochords, which were used as normal controls, were collected. We first conducted droplet digital PCR to quantify the absolute expression levels of the long and short isoforms of the T gene (T-long and T-short, respectively) and revealed that T-long was dominantly expressed in all chordomas and chordoma cell lines, but not in the notochords. The T-long/T-short ratio was significantly different between the chordomas and the notochords. Next, we validated the isoform expression pattern at protein expression level using Western blot in 9 chordomas. Furthermore, the T gene single nucleotide polymorphism site rs2305089, which is the only marker reported to be associated with chordomas, was sequenced in all of the chordoma samples. Association between rs2305089 and T-long/T-short ratio was not significant, indicating it was not involved in T gene alternative splicing. In conclusion, two T gene isoforms were investigated in skull base chordomas and chordoma cell lines, and the longer isoform was dominantly expressed. The distinct expression patterns of these T gene isoforms may contribute to the pathogenesis of skull base chordomas. However, further studies on the function of these isoforms are needed. PMID:26435504

  17. Patterns of homoeologous gene expression shown by RNA sequencing in hexaploid bread wheat

    PubMed Central

    2014-01-01

    Background Bread wheat (Triticum aestivum) has a large, complex and hexaploid genome consisting of A, B and D homoeologous chromosome sets. Therefore each wheat gene potentially exists as a trio of A, B and D homoeoloci, each of which may contribute differentially to wheat phenotypes. We describe a novel approach combining wheat cytogenetic resources (chromosome substitution ‘nullisomic-tetrasomic’ lines) with next generation deep sequencing of gene transcripts (RNA-Seq), to directly and accurately identify homoeologue-specific single nucleotide variants and quantify the relative contribution of individual homoeoloci to gene expression. Results We discover, based on a sample comprising ~5-10% of the total wheat gene content, that at least 45% of wheat genes are expressed from all three distinct homoeoloci. Most of these genes show strikingly biased expression patterns in which expression is dominated by a single homoeolocus. The remaining ~55% of wheat genes are expressed from either one or two homoeoloci only, through a combination of extensive transcriptional silencing and homoeolocus loss. Conclusions We conclude that wheat is tending towards functional diploidy, through a variety of mechanisms causing single homoeoloci to become the predominant source of gene transcripts. This discovery has profound consequences for wheat breeding and our understanding of wheat evolution. PMID:24726045

  18. Differential Expression of Inflammation-Related Genes in Children with Down Syndrome

    PubMed Central

    Silva, Cláudia Regina Santos; Biselli-Périco, Joice Matos; Zampieri, Bruna Lancia; Silva, Wilson Araujo; de Souza, Jorge Estefano Santana; Bürger, Matheus Carvalho; Goloni-Bertollo, Eny Maria; Pavarino, Érika Cristina

    2016-01-01

    Objective. The aim of the study was to investigate the expression patterns of a specific set of genes involved in the inflammation process in children with Down Syndrome (DS) and children without the syndrome (control group) to identify differences that may be related to the immune abnormalities observed in DS individuals. Method. RNA samples were obtained from peripheral blood, and gene expression was quantified using the TaqMan® Array Plate Human Inflammation Kit, which facilitated the investigation into 92 inflammation-related genes and four reference genes using real-time polymerase chain reaction (qPCR). Results. Twenty genes showed differential expression in children with DS; 12 were overexpressed (PLA2G2D, CACNA1D, ALOX12, VCAM1, ICAM1, PLCD1, ADRB1, HTR3A, PDE4C, CASP1, PLA2G5, and PLCB4), and eight were underexpressed (LTA4H, BDKRB1, ADRB2, CD40LG, ITGAM, TNFRSF1B, ITGB1, and TBXAS1). After statistically correcting for the false discovery rate, only the genes BDKRB1 and LTA4H showed differential expression, and both were underexpressed within the DS group. Conclusion. DS children showed differential expression of inflammation-related genes that were not located on chromosome 21 compared with children without DS. The BDKRB1 and LTA4H genes may differentiate the case and control groups based on the inflammatory response, which plays an important role in DS pathogenesis. PMID:27293319

  19. Serial Analysis of Gene Expression: Applications in Malaria Parasite, Yeast, Plant, and Animal Studies

    PubMed Central

    Tuteja, Renu

    2004-01-01

    The serial analysis of gene expression (SAGE) method is based on the isolation of unique sequence tags from individual transcripts and concatenation of tags serially into long DNA molecules. SAGE is an innovative technique that offers the potential of cataloging both the identity and relative frequencies of mRNA transcripts in a given RNA preparation. It can quantify low-abundance transcripts and reliably detect relatively small differences in transcript abundance between cell populations. SAGE data can be used to complement studies in cases where other gene expression methods may be more convenient or efficient. SAGE can be used in a wide variety of applications to identify disease-related genes, to analyze the effect of drugs on tissues, and to provide insights into the disease pathways. The most important application of SAGE is the identification of differentially expressed genes. In this review, we describe various applications of this powerful technology in malarial parasite, yeast, plant, and animal systems. PMID:15240921

  20. Sequence determinants of prokaryotic gene expression level under heat stress.

    PubMed

    Xiong, Heng; Yang, Yi; Hu, Xiao-Pan; He, Yi-Ming; Ma, Bin-Guang

    2014-11-01

    Prokaryotic gene expression is environment-dependent and temperature plays an important role in shaping the gene expression profile. Revealing the regulation mechanisms of gene expression pertaining to temperature has attracted tremendous efforts in recent years particularly owning to the yielding of transcriptome and proteome data by high-throughput techniques. However, most of the previous works concentrated on the characterization of the gene expression profile of individual organism and little effort has been made to disclose the commonality among organisms, especially for the gene sequence features. In this report, we collected the transcriptome and proteome data measured under heat stress condition from recently published literature and studied the sequence determinants for the expression level of heat-responsive genes on multiple layers. Our results showed that there indeed exist commonness and consistent patterns of the sequence features among organisms for the differentially expressed genes under heat stress condition. Some features are attributed to the requirement of thermostability while some are dominated by gene function. The revealed sequence determinants of bacterial gene expression level under heat stress complement the knowledge about the regulation factors of prokaryotic gene expression responding to the change of environmental conditions. Furthermore, comparisons to thermophilic adaption have been performed to reveal the similarity and dissimilarity of the sequence determinants for the response to heat stress and for the adaption to high habitat temperature, which elucidates the complex landscape of gene expression related to the same physical factor of temperature.

  1. Gene Expression Switching of Receptor Subunits in Human Brain Development

    PubMed Central

    Bar-Shira, Ossnat; Maor, Ronnie; Chechik, Gal

    2015-01-01

    Synaptic receptors in the human brain consist of multiple protein subunits, many of which have multiple variants, coded by different genes, and are differentially expressed across brain regions and developmental stages. The brain can tune the electrophysiological properties of synapses to regulate plasticity and information processing by switching from one protein variant to another. Such condition-dependent variant switch during development has been demonstrated in several neurotransmitter systems including NMDA and GABA. Here we systematically detect pairs of receptor-subunit variants that switch during the lifetime of the human brain by analyzing postmortem expression data collected in a population of donors at various ages and brain regions measured using microarray and RNA-seq. To further detect variant pairs that co-vary across subjects, we present a method to quantify age-corrected expression correlation in face of strong temporal trends. This is achieved by computing the correlations in the residual expression beyond a cubic-spline model of the population temporal trend, and can be seen as a nonlinear version of partial correlations. Using these methods, we detect multiple new pairs of context dependent variants. For instance, we find a switch from GLRA2 to GLRA3 that differs from the known switch in the rat. We also detect an early switch from HTR1A to HTR5A whose trends are negatively correlated and find that their age-corrected expression is strongly positively correlated. Finally, we observe that GRIN2B switch to GRIN2A occurs mostly during embryonic development, presumably earlier than observed in rodents. These results provide a systematic map of developmental switching in the neurotransmitter systems of the human brain. PMID:26636753

  2. Gene Expression Switching of Receptor Subunits in Human Brain Development.

    PubMed

    Bar-Shira, Ossnat; Maor, Ronnie; Chechik, Gal

    2015-12-01

    Synaptic receptors in the human brain consist of multiple protein subunits, many of which have multiple variants, coded by different genes, and are differentially expressed across brain regions and developmental stages. The brain can tune the electrophysiological properties of synapses to regulate plasticity and information processing by switching from one protein variant to another. Such condition-dependent variant switch during development has been demonstrated in several neurotransmitter systems including NMDA and GABA. Here we systematically detect pairs of receptor-subunit variants that switch during the lifetime of the human brain by analyzing postmortem expression data collected in a population of donors at various ages and brain regions measured using microarray and RNA-seq. To further detect variant pairs that co-vary across subjects, we present a method to quantify age-corrected expression correlation in face of strong temporal trends. This is achieved by computing the correlations in the residual expression beyond a cubic-spline model of the population temporal trend, and can be seen as a nonlinear version of partial correlations. Using these methods, we detect multiple new pairs of context dependent variants. For instance, we find a switch from GLRA2 to GLRA3 that differs from the known switch in the rat. We also detect an early switch from HTR1A to HTR5A whose trends are negatively correlated and find that their age-corrected expression is strongly positively correlated. Finally, we observe that GRIN2B switch to GRIN2A occurs mostly during embryonic development, presumably earlier than observed in rodents. These results provide a systematic map of developmental switching in the neurotransmitter systems of the human brain.

  3. Quantifying penetrance in a dominant disease gene using large population control cohorts

    PubMed Central

    Minikel, Eric Vallabh; Vallabh, Sonia M.; Lek, Monkol; Estrada, Karol; Samocha, Kaitlin E.; Sathirapongsasuti, J. Fah; McLean, Cory Y.; Tung, Joyce Y.; Yu, Linda P.C.; Gambetti, Pierluigi; Blevins, Janis; Zhang, Shulin; Cohen, Yvonne; Chen, Wei; Yamada, Masahito; Hamaguchi, Tsuyoshi; Sanjo, Nobuo; Mizusawa, Hidehiro; Nakamura, Yosikazu; Kitamoto, Tetsuyuki; Collins, Steven J.; Boyd, Alison; Will, Robert G.; Knight, Richard; Ponto, Claudia; Zerr, Inga; Kraus, Theo F.J.; Eigenbrod, Sabina; Giese, Armin; Calero, Miguel; de Pedro-Cuesta, Jesús; Haïk, Stéphane; Laplanche, Jean-Louis; Bouaziz-Amar, Elodie; Brandel, Jean-Philippe; Capellari, Sabina; Parchi, Piero; Poleggi, Anna; Ladogana, Anna; O'Donnell-Luria, Anne H.; Karczewski, Konrad J.; Marshall, Jamie L.; Boehnke, Michael; Laakso, Markku; Mohlke, Karen L.; Kähler, Anna; Chambert, Kimberly; McCarroll, Steven; Sullivan, Patrick F.; Hultman, Christina M.; Purcell, Shaun M.; Sklar, Pamela; van der Lee, Sven J.; Rozemuller, Annemieke; Jansen, Casper; Hofman, Albert; Kraaij, Robert; van Rooij, Jeroen G.J.; Ikram, M. Arfan; Uitterlinden, André G.; van Duijn, Cornelia M.; Daly, Mark J.; MacArthur, Daniel G.

    2016-01-01

    More than 100,000 genetic variants are reported to cause Mendelian disease in humans, but the penetrance - the probability that a carrier of the purported disease-causing genotype will indeed develop the disease - is generally unknown. Here we assess the impact of variants in the prion protein gene (PRNP) on the risk of prion disease by analyzing 16,025 prion disease cases, 60,706 population control exomes, and 531,575 individuals genotyped by 23andMe, Inc. We show that missense variants in PRNP previously reported to be pathogenic are at least 30× more common in the population than expected based on genetic prion disease prevalence. While some of this excess can be attributed to benign variants falsely assigned as pathogenic, other variants have genuine effects on disease susceptibility but confer lifetime risks ranging from <0.1% to ~100%. We also show that truncating variants in PRNP have position-dependent effects, with true loss-of-function alleles found in healthy older individuals, supporting the safety of therapeutic suppression of prion protein expression. PMID:26791950

  4. Global analysis of patterns of gene expression during Drosophila embryogenesis

    PubMed Central

    Tomancak, Pavel; Berman, Benjamin P; Beaton, Amy; Weiszmann, Richard; Kwan, Elaine; Hartenstein, Volker; Celniker, Susan E; Rubin, Gerald M

    2007-01-01

    Background Cell and tissue specific gene expression is a defining feature of embryonic development in multi-cellular organisms. However, the range of gene expression patterns, the extent of the correlation of expression with function, and the classes of genes whose spatial expression are tightly regulated have been unclear due to the lack of an unbiased, genome-wide survey of gene expression patterns. Results We determined and documented embryonic expression patterns for 6,003 (44%) of the 13,659 protein-coding genes identified in the Drosophila melanogaster genome with over 70,000 images and controlled vocabulary annotations. Individual expression patterns are extraordinarily diverse, but by supplementing qualitative in situ hybridization data with quantitative microarray time-course data using a hybrid clustering strategy, we identify groups of genes with similar expression. Of 4,496 genes with detectable expression in the embryo, 2,549 (57%) fall into 10 clusters representing broad expression patterns. The remaining 1,947 (43%) genes fall into 29 clusters representing restricted expression, 20% patterned as early as blastoderm, with the majority restricted to differentiated cell types, such as epithelia, nervous system, or muscle. We investigate the relationship between expression clusters and known molecular and cellular-physiological functions. Conclusion Nearly 60% of the genes with detectable expression exhibit broad patterns reflecting quantitative rather than qualitative differences between tissues. The other 40% show tissue-restricted expression; the expression patterns of over 1,500 of these genes are documented here for the first time. Within each of these categories, we identified clusters of genes associated with particular cellular and developmental functions. PMID:17645804

  5. Accurate Gene Expression-Based Biodosimetry Using a Minimal Set of Human Gene Transcripts

    SciTech Connect

    Tucker, James D.; Joiner, Michael C.; Thomas, Robert A.; Grever, William E.; Bakhmutsky, Marina V.; Chinkhota, Chantelle N.; Smolinski, Joseph M.; Divine, George W.; Auner, Gregory W.

    2014-03-15

    Purpose: Rapid and reliable methods for conducting biological dosimetry are a necessity in the event of a large-scale nuclear event. Conventional biodosimetry methods lack the speed, portability, ease of use, and low cost required for triaging numerous victims. Here we address this need by showing that polymerase chain reaction (PCR) on a small number of gene transcripts can provide accurate and rapid dosimetry. The low cost and relative ease of PCR compared with existing dosimetry methods suggest that this approach may be useful in mass-casualty triage situations. Methods and Materials: Human peripheral blood from 60 adult donors was acutely exposed to cobalt-60 gamma rays at doses of 0 (control) to 10 Gy. mRNA expression levels of 121 selected genes were obtained 0.5, 1, and 2 days after exposure by reverse-transcriptase real-time PCR. Optimal dosimetry at each time point was obtained by stepwise regression of dose received against individual gene transcript expression levels. Results: Only 3 to 4 different gene transcripts, ASTN2, CDKN1A, GDF15, and ATM, are needed to explain ≥0.87 of the variance (R{sup 2}). Receiver-operator characteristics, a measure of sensitivity and specificity, of 0.98 for these statistical models were achieved at each time point. Conclusions: The actual and predicted radiation doses agree very closely up to 6 Gy. Dosimetry at 8 and 10 Gy shows some effect of saturation, thereby slightly diminishing the ability to quantify higher exposures. Analyses of these gene transcripts may be advantageous for use in a field-portable device designed to assess exposures in mass casualty situations or in clinical radiation emergencies.

  6. Expression Quantitative Trait Loci Analysis Identifies Associations Between Genotype and Gene Expression in Human Intestine

    PubMed Central

    KABAKCHIEV, BOYKO; SILVERBERG, MARK S.

    2013-01-01

    BACKGROUND & AIMS Genome-wide association studies have greatly increased our understanding of intestinal disease. However, little is known about how genetic variations result in phenotypic changes. Some polymorphisms have been shown to modulate quantifiable phenotypic traits; these are called quantitative trait loci. Quantitative trait loci that affect levels of gene expression are called expression quantitative trait loci (eQTL), which can provide insight into the biological relevance of data from genome-wide association studies. We performed a comprehensive eQTL scan of intestinal tissue. METHODS Total RNA was extracted from ileal biopsy specimens and genomic DNA was obtained from whole-blood samples from the same cohort of individuals. Cis- and trans-eQTL analyses were performed using a custom software pipeline for samples from 173 subjects. The analyses determined the expression levels of 19,047 unique autosomal genes listed in the US National Center for Biotechnology Information database and more than 580,000 variants from the Single Nucleotide Polymorphism database. RESULTS The presence of more than 15,000 cis- and trans-eQTL was detected with statistical significance. eQTL associated with the same expression trait were in high linkage disequilibrium. Comparative analysis with previous eQTL studies showed that 30% to 40% of genes identified as eQTL in monocytes, liver tissue, lymphoblastoid cell lines, T cells, and fibroblasts are also eQTL in ileal tissue. Conversely, most of the significant eQTL have not been previously identified and could be tissue specific. These are involved in many cell functions, including division and antigen processing and presentation. Our analysis confirmed that previously published cis-eQTL are single nucleotide polymorphisms associated with inflammatory bowel disease: rs2298428/UBE2L3, rs1050152/SLC22A4, and SLC22A5. We identified many new associations between inflammatory bowel disease susceptibility loci and gene expression

  7. Application of qRT-PCR and RNA-Seq analysis for the identification of housekeeping genes useful for normalization of gene expression values during Striga hermonthica development.

    PubMed

    Fernández-Aparicio, M; Huang, K; Wafula, E K; Honaas, L A; Wickett, N J; Timko, M P; Depamphilis, C W; Yoder, J I; Westwood, J H

    2013-04-01

    Striga is a root parasitic weed that attacks many of the staple crops in Africa, India and Southeast Asia, inflicting tremendous losses in yield and for which there are few effective control measures. Studies of parasitic plant virulence and host resistance will be greatly facilitated by the recent emergence of genomic resources that include extensive transcriptome sequence datasets spanning all life stages of S. hermonthica. Functional characterization of Striga genes will require detailed analyses of gene expression patterns. Quantitative real-time PCR is a powerful tool for quantifying gene expression, but correct normalization of expression levels requires identification of control genes that have stable expression across tissues and life stages. Since no S. hermonthica housekeeping genes have been established for this purpose, we evaluated the suitability of six candidate housekeeping genes across key life stages of S. hermonthica from seed conditioning to flower initiation using qRT-PCR and high-throughput cDNA sequencing. Based on gene expression analysis by qRT-PCR and RNA-Seq across heterogeneous Striga life stages, we determined that using the combination of three genes, UBQ1, PP2A and TUB1 provides the best normalization for gene expression throughout the parasitic life cycle. The housekeeping genes characterized here provide robust standards that will facilitate powerful descriptions of parasite gene expression patterns.

  8. Melatonin regulation of antioxidant enzyme gene expression.

    PubMed

    Mayo, J C; Sainz, R M; Antoli, I; Herrera, F; Martin, V; Rodriguez, C

    2002-10-01

    Antioxidant enzymes (AOEs) are part of the primary cellular defense against free radicals induced by toxins and/or spontaneously formed in cells. Melatonin (MLT) has received much attention in recent years due to its direct free radical scavenging and antioxidant properties. In the present work we report that MLT, at physiological serum concentrations (1 nM), increases the mRNA of both superoxide dismutases (SODs) and glutathione peroxidase (GPx) in two neuronal cell lines. The MLT effect on both SODs and GPx mRNA was mediated by a de novo synthesized protein. MLT alters mRNA stability for Cu-Zn SOD and GPx. Experiments with a short time treatment (pulse action) of MLT suggest that the regulation of AOE gene expression is likely to be receptor mediated, because 1-h treatment with MLT results in the same response as a 24-h treatment.

  9. Gene expression during fruit ripening in avocado.

    PubMed

    Christoffersen, R E; Warm, E; Laties, G G

    1982-06-01

    The poly(A) (+)RNA populations from avocado fruit (Persea americana Mill cv. Hass) at four stages of ripening were isolated by two cycles of oligo-dT-cellulose chromatography and examined by invitro translation, using the rabbit reticulocyte lysate system, followed by two-dimensional gel electrophoresis (isoelectric focusing followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis) of the resulting translation products. Three mRNAs increased dramatically with the climacteric rise in respiration and ethylene production. The molecular weights of the corresponding translation products from the ripening-related mRNAs are 80,000, 36,000, and 16,500. These results indicate that ripening may be linked to the expression of specific genes.

  10. Monoallelic expression of the human FOXP2 speech gene.

    PubMed

    Adegbola, Abidemi A; Cox, Gerald F; Bradshaw, Elizabeth M; Hafler, David A; Gimelbrant, Alexander; Chess, Andrew

    2015-06-01

    The recent descriptions of widespread random monoallelic expression (RMAE) of genes distributed throughout the autosomal genome indicate that there are more genes subject to RMAE on autosomes than the number of genes on the X chromosome where X-inactivation dictates RMAE of X-linked genes. Several of the autosomal genes that undergo RMAE have independently been implicated in human Mendelian disorders. Thus, parsing the relationship between allele-specific expression of these genes and disease is of interest. Mutations in the human forkhead box P2 gene, FOXP2, cause developmental verbal dyspraxia with profound speech and language deficits. Here, we show that the human FOXP2 gene undergoes RMAE. Studying an individual with developmental verbal dyspraxia, we identify a deletion 3 Mb away from the FOXP2 gene, which impacts FOXP2 gene expression in cis. Together these data suggest the intriguing possibility that RMAE impacts the haploinsufficiency phenotypes observed for FOXP2 mutations.

  11. Optimizing retroviral gene expression for effective therapies.

    PubMed

    Antoniou, Michael N; Skipper, Kristian Alsbjerg; Anakok, Omer

    2013-04-01

    With their ability to integrate their genetic material into the target cell genome, retroviral vectors (RV) of both the gamma-retroviral (γ-RV) and lentiviral vector (LV) classes currently remain the most efficient and thus the system of choice for achieving transgene retention and therefore potentially long-term expression and therapeutic benefit. However, γ-RV and LV integration comes at a cost in that transcription units will be present within a native chromatin environment and thus be subject to epigenetic effects (DNA methylation, histone modifications) that can negatively impact on their function. Indeed, highly variable expression and silencing of γ-RV and LV transgenes especially resulting from promoter DNA methylation is well documented and was the cause of the failure of gene therapy in a clinical trial for X-linked chronic granulomatous disease. This review will critically explore the use of different classes of genetic control elements that can in principle reduce vector insertion site position effects and epigenetic-mediated silencing. These transcriptional regulatory elements broadly divide themselves into either those with a chromatin boundary or border function (scaffold/matrix attachment regions, insulators) or those with a dominant chromatin remodeling and transcriptional activating capability (locus control regions,, ubiquitous chromatin opening elements). All these types of elements have their strengths and weaknesses within the constraints of a γ-RV and LV backbone, showing varying degrees of efficacy in improving reproducibility and stability of transgene function. Combinations of boundary and chromatin remodeling; transcriptional activating elements, which do not impede vector production; transduction efficiency; and stability are most likely to meet the requirements within a gene therapy context especially when targeting a stem cell population.

  12. Phenotypic plasticity and divergence in gene expression.

    PubMed

    Healy, Timothy M; Schulte, Patricia M

    2015-07-01

    The extent to which phenotypic plasticity, or the ability of a single genotype to produce different phenotypes in different environments, impedes or promotes genetic divergence has been a matter of debate within evolutionary biology for many decades (see, for example, Ghalambor et al. ; Pfennig et al. ). Similarly, the role of evolution in shaping phenotypic plasticity remains poorly understood (Pigliucci ). In this issue of Molecular Ecology, Dayan et al. () provide empirical data relevant to these questions by assessing the extent of plasticity and divergence in the expression levels of 2272 genes in muscle tissue from killifish (genus Fundulus) exposed to different temperatures. F. heteroclitus (Fig. A) and F. grandis are minnows that inhabit estuarine marshes (Fig. B) along the coasts of the Atlantic Ocean and Gulf of Mexico in North America. These habitats undergo large variations in temperature both daily and seasonally, and these fish are known to demonstrate substantial phenotypic plasticity in response to temperature change (e.g. Fangue et al. ). Furthermore, the range of F. heteroclitus spans a large latitudinal gradient of temperatures, such that northern populations experience temperatures that are on average ~10°C colder than do southern populations (Schulte ). By comparing gene expression patterns between populations of these fish from different thermal habitats held in the laboratory at three different temperatures, Dayan et al. () address two important questions regarding the interacting effects of plasticity and evolution: (i) How does phenotypic plasticity affect adaptive divergence? and (ii) How does adaptive divergence affect plasticity? PMID:26096949

  13. Many body theory of stochastic gene expression

    NASA Astrophysics Data System (ADS)

    Walczak, Aleksandra M.

    The regulation of expression states of genes in cells is a stochastic process. The relatively small numbers of protein molecules of a given type present in the cell and the nonlinear nature of chemical reactions result in behaviours, which are hard to anticipate without an appropriate mathematical development. In this dissertation, I develop theoretical approaches based on methods of statistical physics and many-body theory, in which protein and operator state dynamics are treated stochastically and on an equal footing. This development allows me to study the general principles of how noise arising on different levels of the regulatory system affects the complex collective characteristics of systems observed experimentally. I discuss simple models and approximations, which allow for, at least some, analytical progress in these problems. These have allowed us to understand how the operator state fluctuations may influence the steady state properties and lifetimes of attractors of simple gene systems. I show, that for fast binding and unbinding from the DNA, the operator state may be taken to be in equilibrium for highly cooperative binding, when predicting steady state properties as is traditionally done. Nevertheless, if proteins are produced in bursts, the DNA binding state fluctuations must be taken into account explicitly. Furthermore, even when the steady state probability distributions are weakly influenced by the operator state fluctuations, the escape rate in biologically relevant regimes strongly depends on transcription factor-DNA binding rates.

  14. Selection of reference genes for expression analyses of red-fleshed sweet orange (Citrus sinensis).

    PubMed

    Pinheiro, T T; Nishimura, D S; De Nadai, F B; Figueira, A; Latado, R R

    2015-01-01

    Red-fleshed oranges (Citrus sinensis) contain high levels of carotenoids and lycopene. The growing consumer demand for products with health benefits has increased interest in these types of Citrus cultivars as a potential source of nutraceuticals. However, little is known about the physiology of these cultivars under Brazilian conditions. Transcriptome and gene expression analyses are important tools in the breeding and management of red-fleshed sweet orange cultivars. Reverse transcription quantitative polymerase chain reaction is a method of quantifying gene expression, but various standardizations are required to obtain precise, accurate, and specific results. Among the standardizations required, the choice of suitable stable reference genes is fundamental. The objective of this study was to evaluate the stability of 11 candidate genes using various tissue and organ samples from healthy plants or leaves from citrus greening disease (Huanglongbing)-symptomatic plants of a Brazilian red-fleshed cultivar ('Sanguínea de Mombuca'), in order to select the most suitable reference gene for investigating gene expression under these conditions. geNorm and NormFinder identified genes that encoded translation initiation factor 3, ribosomal protein L35, and translation initiation factor 5A as the most stable genes under the biological conditions tested, and genes coding actin (ACT) and the subunit of the PSI reaction center subunit III were the least stable. Phosphatase, malate dehydrogenase, and ACT were the most stable genes in the leaf samples of infected plants. PMID:26782492

  15. Selection of reference genes for expression analyses of red-fleshed sweet orange (Citrus sinensis).

    PubMed

    Pinheiro, T T; Nishimura, D S; De Nadai, F B; Figueira, A; Latado, R R

    2015-12-28

    Red-fleshed oranges (Citrus sinensis) contain high levels of carotenoids and lycopene. The growing consumer demand for products with health benefits has increased interest in these types of Citrus cultivars as a potential source of nutraceuticals. However, little is known about the physiology of these cultivars under Brazilian conditions. Transcriptome and gene expression analyses are important tools in the breeding and management of red-fleshed sweet orange cultivars. Reverse transcription quantitative polymerase chain reaction is a method of quantifying gene expression, but various standardizations are required to obtain precise, accurate, and specific results. Among the standardizations required, the choice of suitable stable reference genes is fundamental. The objective of this study was to evaluate the stability of 11 candidate genes using various tissue and organ samples from healthy plants or leaves from citrus greening disease (Huanglongbing)-symptomatic plants of a Brazilian red-fleshed cultivar ('Sanguínea de Mombuca'), in order to select the most suitable reference gene for investigating gene expression under these conditions. geNorm and NormFinder identified genes that encoded translation initiation factor 3, ribosomal protein L35, and translation initiation factor 5A as the most stable genes under the biological conditions tested, and genes coding actin (ACT) and the subunit of the PSI reaction center subunit III were the least stable. Phosphatase, malate dehydrogenase, and ACT were the most stable genes in the leaf samples of infected plants.

  16. Expression profiling identifies genes involved in emphysema severity.

    PubMed

    Francis, Santiyagu M Savarimuthu; Larsen, Jill E; Pavey, Sandra J; Bowman, Rayleen V; Hayward, Nicholas K; Fong, Kwun M; Yang, Ian A

    2009-01-01

    Chronic obstructive pulmonary disease (COPD) is a major public health problem. The aim of this study was to identify genes involved in emphysema severity in COPD patients.Gene expression profiling was performed on total RNA extracted from non-tumor lung tissue from 30 smokers with emphysema. Class comparison analysis based on gas transfer measurement was performed to identify differentially expressed genes. Genes were then selected for technical validation by quantitative reverse transcriptase-PCR (qRT-PCR) if also represented on microarray platforms used in previously published emphysema studies. Genes technically validated advanced to tests of biological replication by qRT-PCR using an independent test set of 62 lung samples.Class comparison identified 98 differentially expressed genes (p < 0.01). Fifty-one of those genes had been previously evaluated in differentiation between normal and severe emphysema lung. qRT-PCR confirmed the direction of change in expression in 29 of the 51 genes and 11 of those validated, remaining significant at p < 0.05. Biological replication in an independent cohort confirmed the altered expression of eight genes, with seven genes differentially expressed by greater than 1.3 fold, identifying these as candidate determinants of emphysema severity.Gene expression profiling of lung from emphysema patients identified seven candidate genes associated with emphysema severity including COL6A3, SERPINF1, ZNHIT6, NEDD4, CDKN2A, NRN1 and GSTM3. PMID:19723343

  17. Expression profiling identifies genes involved in emphysema severity

    PubMed Central

    2009-01-01

    Chronic obstructive pulmonary disease (COPD) is a major public health problem. The aim of this study was to identify genes involved in emphysema severity in COPD patients. Gene expression profiling was performed on total RNA extracted from non-tumor lung tissue from 30 smokers with emphysema. Class comparison analysis based on gas transfer measurement was performed to identify differentially expressed genes. Genes were then selected for technical validation by quantitative reverse transcriptase-PCR (qRT-PCR) if also represented on microarray platforms used in previously published emphysema studies. Genes technically validated advanced to tests of biological replication by qRT-PCR using an independent test set of 62 lung samples. Class comparison identified 98 differentially expressed genes (p < 0.01). Fifty-one of those genes had been previously evaluated in differentiation between normal and severe emphysema lung. qRT-PCR confirmed the direction of change in expression in 29 of the 51 genes and 11 of those validated, remaining significant at p < 0.05. Biological replication in an independent cohort confirmed the altered expression of eight genes, with seven genes differentially expressed by greater than 1.3 fold, identifying these as candidate determinants of emphysema severity. Gene expression profiling of lung from emphysema patients identified seven candidate genes associated with emphysema severity including COL6A3, SERPINF1, ZNHIT6, NEDD4, CDKN2A, NRN1 and GSTM3. PMID:19723343

  18. Structure and expression of the ATFa gene.

    PubMed

    Goetz, J; Chatton, B; Mattei, M G; Kedinger, C

    1996-11-22

    The human ATFa proteins belong to the ATF/CREB family of transcription factors. We have previously shown that they mediate the transcriptional activation by the largest E1a protein and can heterodimerize with members of the Jun/Fos family. ATFa proteins have also been found tightly associated with JNK2, a stress-activated kinase. We now report on the structure of the ATFa gene, which mapped to chromosome 12 (band 12q13). Sequence analysis revealed that ATFa isoforms are generated by alternative splice donor site usage. A minimal promoter region of approximately 200 base pairs was identified that retained nearly full transcriptional activity. Binding sites for potential transcription factors were delineated within a GC-rich segment by DNase I footprinting. Expression studies revealed that ATFa accumulates in the nuclei of transfected cells, and the nuclear localization signal was defined next to the leucine zipper domain. As revealed by hybridization with mouse ATFa sequences, low levels of ATFa mRNAs were ubiquitously distributed in fetal or adult mice, with enhanced expression in particular tissues, like squamous epithelia and specific brain cell layers. The possible significance of coexpression of ATFa, ATF-2, and Jun at similar sites in the brain is discussed. PMID:8939888

  19. Laser capture microdissection for gene expression analysis.

    PubMed

    Bidarimath, Mallikarjun; Edwards, Andrew K; Tayade, Chandrakant

    2015-01-01

    Laser capture microdissection (LCM) is an excellent and perhaps the only platform to isolate homogeneous cell populations from specific microscopic regions of heterogeneous tissue section, under direct microscopic visualization. The basic operations of the LCM system are based on (a) microscopic visualization of phenotypically identified cells of interest, (b) selective adherence of cells to a melting thermolabile film/membrane using a low-energy infrared laser (IR system) or photovolatization of cells within a selected region (UV system), (c) capturing or catapulting of structurally intact cells from a stained tissue section. RNA/DNA or protein can be extracted from the cell or tissue fragments for downstream applications to quantitatively study gene expression. This method can be applied to many downstream analyses including but not limited to quantitative real-time polymerase chain reaction (PCR), microarray, DNA genotyping, RNA transcript profiling, generation of cDNA library, mass spectrometry analysis, and proteomic discovery.The application of LCM is described here to specifically and reliably obtain a homogeneous cell population in order to extract RNA to study microRNA expression by quantitative real-time PCR.

  20. Laser capture microdissection for gene expression analysis.

    PubMed

    Bidarimath, Mallikarjun; Edwards, Andrew K; Tayade, Chandrakant

    2015-01-01

    Laser capture microdissection (LCM) is an excellent and perhaps the only platform to isolate homogeneous cell populations from specific microscopic regions of heterogeneous tissue section, under direct microscopic visualization. The basic operations of the LCM system are based on (a) microscopic visualization of phenotypically identified cells of interest, (b) selective adherence of cells to a melting thermolabile film/membrane using a low-energy infrared laser (IR system) or photovolatization of cells within a selected region (UV system), (c) capturing or catapulting of structurally intact cells from a stained tissue section. RNA/DNA or protein can be extracted from the cell or tissue fragments for downstream applications to quantitatively study gene expression. This method can be applied to many downstream analyses including but not limited to quantitative real-time polymerase chain reaction (PCR), microarray, DNA genotyping, RNA transcript profiling, generation of cDNA library, mass spectrometry analysis, and proteomic discovery.The application of LCM is described here to specifically and reliably obtain a homogeneous cell population in order to extract RNA to study microRNA expression by quantitative real-time PCR. PMID:25308266

  1. Human enteric defensins. Gene structure and developmental expression.

    PubMed

    Mallow, E B; Harris, A; Salzman, N; Russell, J P; DeBerardinis, R J; Ruchelli, E; Bevins, C L

    1996-02-23

    Paneth cells, secretory epithelial cells of the small intestinal crypts, are proposed to contribute to local host defense. Both mouse and human Paneth cells express a collection of antimicrobial proteins, including members of a family of antimicrobial peptides named defensins. In this study, data from an anchored polymerase chain reaction (PCR) strategy suggest that only two defensin mRNA isoforms are expressed in the human small intestine, far fewer than the number expressed in the mouse. The two isoforms detected by this PCR approach were human defensin family members, HD-5 and HD-6. The gene encoding HD-6 was cloned and characterized. HD-6 has a genomic organization similar to HD-5, and the two genes have a striking pattern of sequence similarity localized chiefly in their proximal 5'-flanking regions. Analysis of human fetal RNA by reverse transcriptase-PCR detected enteric defensin HD-5 mRNA at 13.5 weeks of gestation in the small intestine and the colon, but by 17 weeks HD-5 was restricted to the small intestine. HD-6 mRNA was detectable at 13.5-17 weeks of gestation in the small intestine but not in the colon. This pattern of expression coincides with the previously described appearance of Paneth cells as determined by ultrastructural approaches. Northern analysis of total RNA from small intestine revealed quantifiable enteric defensin mRNA in five samples from 19 24 weeks of gestation at levels approximately 40-250-fold less than those observed in the adult, with HD-5 mRNA levels greater than those of HD-6 in all samples. In situ hybridization analysis localized expression of enteric defensin mRNA to Paneth cells at 24 weeks of gestation, as is seen in the newborn term infant and the adult. Consistent with earlier morphological studies, the ratio of Paneth cell number per crypt was reduced in samples at 24 weeks of gestation compared with the adult, and this lower cell number partially accounts for the lower defensin mRNA levels as determined by Northern

  2. Gene Expression Profiling in Pachyonychia Congenita Skin

    PubMed Central

    Cao, Yu-An; Hickerson, Robyn P.; Seegmiller, Brandon L.; Grapov, Dmitry; Gross, Maren M.; Bessette, Marc R.; Phinney, Brett S.; Flores, Manuel A.; Speaker, Tycho J.; Vermeulen, Annaleen; Bravo, Albert A.; Bruckner, Anna L.; Milstone, Leonard M.; Schwartz, Mary E.; Rice, Robert H.; Kaspar, Roger L.

    2015-01-01

    Background Pachyonychia congenita (PC) is a skin disorder resulting from mutations in keratin (K) proteins including K6a, K6b, K16, and K17. One of the major symptoms is painful plantar keratoderma. The pathogenic sequelae resulting from the keratin mutations remain unclear. Objective To better understand PC pathogenesis. Methods RNA profiling was performed on biopsies taken from PC-involved and uninvolved plantar skin of seven genotyped PC patients (two K6a, one K6b, three K16, and one K17) as well as from control volunteers. Protein profiling was generated from tape-stripping samples. Results A comparison of PC-involved skin biopsies to adjacent uninvolved plantar skin identified 112 differentially-expressed mRNAs common to patient groups harboring K6 (i.e., both K6a and K6b) and K16 mutations. Among these mRNAs, 25 encode structural proteins including keratins, small proline-rich and late cornified envelope proteins, 20 are related to metabolism and 16 encode proteases, peptidases, and their inhibitors including kallikrein-related peptidases (KLKs), and serine protease inhibitors (SERPINs). mRNAs were also identified to be differentially expressed only in K6 (81) or K16 (141) patient samples. Furthermore, 13 mRNAs were identified that may be involved in pain including nociception and neuropathy. Protein profiling, comparing three K6a plantar tape-stripping samples to non-PC controls, showed changes in the PC corneocytes similar, but not identical, to the mRNA analysis. Conclusion Many differentially-expressed genes identified in PC-involved skin encode components critical for skin barrier homeostasis including keratinocyte proliferation, differentiation, cornification, and desquamation. The profiling data provide a foundation for unraveling the pathogenesis of PC and identifying targets for developing effective PC therapeutics. PMID:25656049

  3. Genetic basis of differential opsin gene expression in cichlid fishes.

    PubMed

    Carleton, K L; Hofmann, C M; Klisz, C; Patel, Z; Chircus, L M; Simenauer, L H; Soodoo, N; Albertson, R C; Ser, J R

    2010-04-01

    Visual sensitivity can be tuned by differential expression of opsin genes. Among African cichlid fishes, seven cone opsin genes are expressed in different combinations to produce diverse visual sensitivities. To determine the genetic architecture controlling these adaptive differences, we analysed genetic crosses between species expressing different complements of opsin genes. Quantitative genetic analyses suggest that expression is controlled by only a few loci with correlations among some genes. Genetic mapping identifies clear evidence of trans-acting factors in two chromosomal regions that contribute to differences in opsin expression as well as one cis-regulatory region. Therefore, both cis and trans regulation are important. The simple genetic architecture suggested by these results may explain why opsin gene expression is evolutionarily labile, and why similar patterns of expression have evolved repeatedly in different lineages.

  4. Carcinogen-induced trans activation of gene expression

    SciTech Connect

    Kleinberger, T.; Flint, Y.B.; Blank, M.; Etkin, S.; Lavi, S.

    1988-03-01

    The authors report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later.

  5. Carcinogen-induced trans activation of gene expression.

    PubMed Central

    Kleinberger, T; Flint, Y B; Blank, M; Etkin, S; Lavi, S

    1988-01-01

    We report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later. Images PMID:2835673

  6. Learning-based Segmentation Framework for Tissue Images Containing Gene Expression Data

    SciTech Connect

    Bello, Musodiq; Ju, Tao; Carson, James P.; Warren, Joe; Chiu, Wah; Kakadiaris, Ioannis

    2007-05-01

    Abstract Associating specific gene activity with functional locations in the brain results in a greater understanding of the role of the gene. To perform such an association for the over 20,000 genes in the mammalian genome, reliable automated methods that characterize the distribution of gene expression in relation to a standard anatomical model are required. In this paper, we propose a new automatic method that results in the segmentation of gene expression images into distinct anatomical regions in which the expression can be quantified and compared with other images. Our contribution is a novel hybrid atlas that utilizes a statistical shape model based on a subdivision mesh, texture differentiation at region boundaries, and features of anatomical landmarks to delineate boundaries of anatomical regions in gene expression images. This atlas, which provides a common coordinate system for internal brain data, was trained on 36 images manually annotated by neuroanatomists and tested on 64 images. Our framework has achieved a mean overlap ratio of up to 91 § 7% in this challenging dataset. This tool for large-scale annotation will help scientists interpret gene expression patterns more efficiently.

  7. Validation of suitable reference genes for gene expression analysis in the halophyte Salicornia europaea by real-time quantitative PCR

    PubMed Central

    Xiao, Xinlong; Ma, Jinbiao; Wang, Junru; Wu, Xiaomeng; Li, Pengbo; Yao, Yinan

    2015-01-01

    Real-time quantitative polymerase chain reaction (RT-qPCR), a reliable technique for quantifying gene expression, requires stable reference genes to normalize its data. Salicornia europaea, a stem succulent halophyte with remarkable salt resistance and high capacity for ion accumulation, has not been investigated with regards to the selection of appropriate reference genes for RT-qPCR. In this study, the expression of 11 candidate reference genes, GAPDH (Glyceraldehyde 3-phosphate dehydrogenase), Actin, α-Tub (α-tubulin), β-Tub (β-tubulin), EF1-α (Elongation factor 1-α), UBC (Ubiquitin-conjugating enzyme), UBQ (Polyubiquitin), CYP (Cyclophilin), TIP41 (TIP41-like protein), CAC (Clathrin adaptor complexes), and DNAJ (DnaJ-like protein), was analyzed in S. europaea samples, which were classified into groups according to various abiotic stresses (NaCl, nitrogen, drought, cold and heat), tissues and ages. Three commonly used software programs (geNorm, NormFinder and BestKeeper) were applied to evaluate the stability of gene expression, and comprehensive ranks of stability were generated by aggregate analysis. The results show that the relatively stable genes for each group are the following: (1) CAC and UBC for whole samples; (2) CAC and UBC for NaCl stress; (3) Actin and α-Tub for nitrogen treatment; (4) Actin and GAPDH for drought stress; (5) α-Tub and UBC for cold stress; (6) TIP41 and DNAJ for heat stress; (7) UBC and UBQ for different tissues; and (8) UBC and Actin for various developmental stages. These genes were validated by comparing transcriptome profiles. Using two stable reference genes was recommended in the normalization of RT-qPCR data. This study identifies optimal reference genes for RT-qPCR in S. europaea, which will benefit gene expression analysis under these conditions. PMID:25653658

  8. Epigenetic regulation of pyruvate carboxylase gene expression in the postpartum liver.

    PubMed

    Walker, C G; Crookenden, M A; Henty, K M; Handley, R R; Kuhn-Sherlock, B; White, H M; Donkin, S S; Snell, R G; Meier, S; Heiser, A; Loor, J J; Mitchell, M D; Roche, J R

    2016-07-01

    Hepatic gluconeogenesis is essential for maintenance of whole body glucose homeostasis and glucose supply for mammary lactose synthesis in the dairy cow. Upregulation of the gluconeogenic enzyme pyruvate carboxylase (PC) during the transition period is vital in the adaptation to the greater glucose demands associated with peripartum lactogenesis. The objective of this study was to determine if PC transcription in hepatocytes is regulated by DNA methylation and if treatment with a nonsteroidal anti-inflammatory drug (NSAID) alters methylation of an upstream DNA sequence defined as promoter 1. Dairy cows were left untreated (n=20), or treated with a NSAID during the first 5 d postcalving (n=20). Liver was biopsied at d 7 precalving and d 7, 14, and 28 postcalving. Total PC and transcript specific gene expression was quantified using quantitative PCR and DNA methylation of promoter 1 was quantified using bisulfite Sanger sequencing. Expression of PC changed over the transition period, with increased expression postcalving occurring concurrently with increased circulating concentration of nonesterified fatty acids. The DNA methylation percentage was variable at all sites quantified and ranged from 21 to 54% across the 15 CpG dinucleotides within promoter 1. The DNA methylation at wk 1 postcalving, however, was not correlated with gene expression of promoter 1-regulated transcripts and we did not detect an effect of NSAID treatment on DNA methylation or PC gene expression. Our results do not support a role for DNA methylation in regulating promoter 1-driven gene expression of PC at wk 1 postcalving. Further research is required to determine the mechanisms regulating increased PC expression over the transition period. PMID:27085418

  9. Association of tissue lineage and gene expression: conservatively and differentially expressed genes define common and special functions of tissues

    PubMed Central

    2010-01-01

    Background Embryogenesis is the process by which the embryo is formed, develops, and establishes developmental hierarchies of tissues. The recent advance in microarray technology made it possible to investigate the tissue specific patterns of gene expression and their relationship with tissue lineages. This study is focused on how tissue specific functions, tissue lineage, and cell differentiation are correlated, which is essential to understand embryonic development and organism complexity. Results We performed individual gene and gene set based analysis on multiple tissue expression data, in association with the classic topology of mammalian fate maps of embryogenesis. For each sub-group of tissues on the fate map, conservatively, differentially and correlatively expressed genes or gene sets were identified. Tissue distance was found to correlate with gene expression divergence. Tissues of the ectoderm or mesoderm origins from the same segments on the fate map shared more similar expression pattern than those from different origins. Conservatively expressed genes or gene sets define common functions in a tissue group and are related to tissue specific diseases, which is supported by results from Gene Ontology and KEGG pathway analysis. Gene expression divergence is larger in certain human tissues than in the mouse homologous tissues. Conclusion The results from tissue lineage and gene expression analysis indicate that common function features of neighbor tissue groups were defined by the conservatively expressed genes and were related to tissue specific diseases, and differentially expressed genes contribute to the functional divergence of tissues. The difference of gene expression divergence in human and mouse homologous tissues reflected the organism complexity, i.e. distinct neural development levels and different body sizes. PMID:21172044

  10. Global gene expression profiles in developing soybean seeds.

    PubMed

    Asakura, Tomiko; Tamura, Tomoko; Terauchi, Kaede; Narikawa, Tomoyo; Yagasaki, Kazuhiro; Ishimaru, Yoshiro; Abe, Keiko

    2012-03-01

    The gene expression profiles in soybean (Glycine max L.) seeds at 4 stages of development, namely, pod, 2-mm bean, 5-mm bean, and full-size bean, were examined by DNA microarray analysis. The total genes of each sample were classified into 4 clusters based on stage of development. Gene expression was strictly controlled by seed size, which coincides with the development stage. First, stage specific gene expression was examined. Many transcription factors were expressed in pod, 2-mm bean and 5-mm bean. In contrast, storage proteins were mainly expressed in full-size bean. Next, we extracted the genes that are differentially expressed genes (DEGs) that were extracted using the Rank products method of the Bioconductor software package. These DEGs were sorted into 8 groups using the hclust function according to gene expression patterns. Three of the groups across which the expression levels progressively increased included 100 genes, while 3 groups across which the levels decreased contained 47 genes. Storage proteins, seed-maturation proteins, some protease inhibitors, and the allergen Gly m Bd 28K were classified into the former groups. Lipoxygenase (LOX) family members were present in both the groups, indicating the multi-functionality with different expression patterns. PMID:22245912

  11. Faster-X evolution of gene expression in Drosophila.

    PubMed

    Meisel, Richard P; Malone, John H; Clark, Andrew G

    2012-01-01

    DNA sequences on X chromosomes often have a faster rate of evolution when compared to similar loci on the autosomes, and well articulated models provide reasons why the X-linked mode of inheritance may be responsible for the faster evolution of X-linked genes. We analyzed microarray and RNA-seq data collected from females and males of six Drosophila species and found that the expression levels of X-linked genes also diverge faster than autosomal gene expression, similar to the "faster-X" effect often observed in DNA sequence evolution. Faster-X evolution of gene expression was recently described in mammals, but it was limited to the evolutionary lineages shortly following the creation of the therian X chromosome. In contrast, we detect a faster-X effect along both deep lineages and those on the tips of the Drosophila phylogeny. In Drosophila males, the dosage compensation complex (DCC) binds the X chromosome, creating a unique chromatin environment that promotes the hyper-expression of X-linked genes. We find that DCC binding, chromatin environment, and breadth of expression are all predictive of the rate of gene expression evolution. In addition, estimates of the intraspecific genetic polymorphism underlying gene expression variation suggest that X-linked expression levels are not under relaxed selective constraints. We therefore hypothesize that the faster-X evolution of gene expression is the result of the adaptive fixation of beneficial mutations at X-linked loci that change expression level in cis. This adaptive faster-X evolution of gene expression is limited to genes that are narrowly expressed in a single tissue, suggesting that relaxed pleiotropic constraints permit a faster response to selection. Finally, we present a conceptional framework to explain faster-X expression evolution, and we use this framework to examine differences in the faster-X effect between Drosophila and mammals.

  12. Variation in Gene Expression Patterns in Human Gastric Cancers

    PubMed Central

    Chen, Xin; Leung, Suet Y.; Yuen, Siu T.; Chu, Kent-Man; Ji, Jiafu; Li, Rui; Chan, Annie S.Y.; Law, Simon; Troyanskaya, Olga G.; Wong, John; So, Samuel; Botstein, David; Brown, Patrick O.

    2003-01-01

    Gastric cancer is the world's second most common cause of cancer death. We analyzed gene expression patterns in 90 primary gastric cancers, 14 metastatic gastric cancers, and 22 nonneoplastic gastric tissues, using cDNA microarrays representing ∼30,300 genes. Gastric cancers were distinguished from nonneoplastic gastric tissues by characteristic differences in their gene expression patterns. We found a diversity of gene expression patterns in gastric cancer, reflecting variation in intrinsic properties of tumor and normal cells and variation in the cellular composition of these complex tissues. We identified several genes whose expression levels were significantly correlated with patient survival. The variations in gene expression patterns among cancers in different patients suggest differences in pathogenetic pathways and potential therapeutic strategies. PMID:12925757

  13. Ordered expression of virulence genes in Salmonella enterica serovar typhimurium.

    PubMed

    Papezova, K; Gregorova, D; Jonuschies, J; Rychlik, I

    2007-01-01

    Using transcriptional promoter fusions, we investigated the expression of selected SPI-1 and SPI-2 genes of Salmonella enterica serovar Typhimurium (S. Typhimurium). Promoters of genes related to the invasion of the epithelial cell (hilA, hilC, hilD, invF, sicA, sopA, sopB and sopE2) were active in Luria-Bertani (LB) medium and LB with butyrate but were suppressed by bile salts and in glucose minimal (M9) medium. Genes related to S. Typhimurium intracellular survival (phoP, ssrA, ssaB, ssaG, sifA, sifB and pipB) were characterized by their expression in stationary phase in LB and M9 medium. Activity of phoP and ssrA promoters indicated that these might be expressed inside the gut. SPI-1 genes were expressed on the transition to stationary phase while SPI-2 genes were expressed in stationary phase. Among SPI-1 genes, those with regulatory functions preceded in expression the effector genes and sop genes were expressed in the order of sopA, sopB and sopE2, showing hierarchy in the expression of S. Typhimurium virulence genes.

  14. Benzoic Acid-Inducible Gene Expression in Mycobacteria

    PubMed Central

    Dragset, Marte S.; Barczak, Amy K.; Kannan, Nisha; Mærk, Mali; Flo, Trude H.; Valla, Svein; Rubin, Eric J.; Steigedal, Magnus

    2015-01-01

    Conditional expression is a powerful tool to investigate the role of bacterial genes. Here, we adapt the Pseudomonas putida-derived positively regulated XylS/Pm expression system to control inducible gene expression in Mycobacterium smegmatis and Mycobacterium tuberculosis, the causative agent of human tuberculosis. By making simple changes to a Gram-negative broad-host-range XylS/Pm-regulated gene expression vector, we prove that it is possible to adapt this well-studied expression system to non-Gram-negative species. With the benzoic acid-derived inducer m-toluate, we achieve a robust, time- and dose-dependent reversible induction of Pm-mediated expression in mycobacteria, with low background expression levels. XylS/Pm is thus an important addition to existing mycobacterial expression tools, especially when low basal expression is of particular importance. PMID:26348349

  15. Genes, environment and gene expression in colon tissue: a pathway approach to determining functionality.

    PubMed

    Slattery, Martha L; Pellatt, Daniel F; Wolff, Roger K; Lundgreen, Abbie

    2016-01-01

    Genetic and environmental factors have been shown to work together to alter cancer risk. In this study we evaluate previously identified gene and lifestyle interactions in a candidate pathway that were associated with colon cancer risk to see if these interactions altered gene expression. We analyzed non-tumor RNA-seq data from 144 colon cancer patients who had genotype, recent cigarette smoking, diet, body mass index (BMI), and recent aspirin/non-steroidal anti-inflammatory use data. Using a false discovery rate of 0.1, we evaluated differential gene expression between high and low levels of lifestyle exposure and genotypes using DESeq2. Thirteen pathway genes and 17 SNPs within those genes were associated with altered expression of other genes in the pathway. BMI, NSAIDs use and dietary components of the oxidative balance score (OBS) also were associated with altered gene expression. SNPs previously identified as interacting with these lifestyle factors, altered expression of pathway genes. NSAIDs interacted with 10 genes (15 SNPs) within those genes to alter expression of 28 pathway genes; recent cigarette smoking interacted with seven genes (nine SNPs) to alter expression of 27 genes. BMI interacted with FLT1, KDR, SEPN1, TERT, TXNRD2, and VEGFA to alter expression of eight genes. Three genes (five SNPs) interacted with OBS to alter expression of 12 genes. These data provide support for previously identified lifestyle and gene interactions associated with colon cancer in that they altered expression of key pathway genes. The need to consider lifestyle factors in conjunction with genetic factors is illustrated.

  16. Quantifying the Relative Importance of Phylogeny and Environmental Preferences As Drivers of Gene Content in Prokaryotic Microorganisms

    PubMed Central

    Tamames, Javier; Sánchez, Pablo D.; Nikel, Pablo I.; Pedrós-Alió, Carlos

    2016-01-01

    Two complementary forces shape microbial genomes: vertical inheritance of genes by phylogenetic descent, and acquisition of new genes related to adaptation to particular habitats and lifestyles. Quantification of the relative importance of each driving force proved difficult. We determined the contribution of each factor, and identified particular genes or biochemical/cellular processes linked to environmental preferences (i.e., propensity of a taxon to live in particular habitats). Three types of data were confronted: (i) complete genomes, which provide gene content of different taxa; (ii) phylogenetic information, via alignment of 16S rRNA sequences, which allowed determination of the distance between taxa, and (iii) distribution of species in environments via 16S rRNA sampling experiments, reflecting environmental preferences of different taxa. The combination of these three datasets made it possible to describe and quantify the relationships among them. We found that, although phylogenetic descent was responsible for shaping most genomes, a discernible part of the latter was correlated to environmental adaptations. Particular families of genes were identified as environmental markers, as supported by direct studies such as metagenomic sequencing. These genes are likely important for adaptation of bacteria to particular conditions or habitats, such as carbohydrate or glycan metabolism genes being linked to host-associated environments. PMID:27065987

  17. Gammaherpesvirus Lytic Gene Expression as Characterized by DNA Array

    PubMed Central

    Ahn, Joo Wook; Powell, Kenneth L.; Kellam, Paul; Alber, Dagmar G.

    2002-01-01

    Gammaherpesviruses are associated with a number of diseases including lymphomas and other malignancies. Murine gammaherpesvirus 68 (MHV-68) constitutes the most amenable animal model for this family of pathogens. However experimental characterization of gammaherpesvirus gene expression, at either the protein or RNA level, lags behind that of other, better-studied alpha- and beta-herpesviruses. We have developed a cDNA array to globally characterize MHV-68 gene expression profiles, thus providing an experimental supplement to a genome that is chiefly annotated by homology. Viral genes started to be transcribed as early as 3 h postinfection (p.i.), and this was followed by a rapid escalation of gene expression that could be seen at 5 h p.i. Individual genes showed their own transcription profiles, and most genes were still being expressed at 18 h p.i. Open reading frames (ORFs) M3 (chemokine-binding protein), 52, and M9 (capsid protein) were particularly noticeable due to their very high levels of expression. Hierarchical cluster analysis of transcription profiles revealed four main groups of genes and allowed functional predictions to be made by comparing expression profiles of uncharacterized genes to those of genes of known function. Each gene was also categorized according to kinetic class by blocking de novo protein synthesis and viral DNA replication in vitro. One gene, ORF 73, was found to be expressed with α-kinetics, 30 genes were found to be expressed with β-kinetics, and 42 genes were found to be expressed with γ-kinetics. This fundamental characterization furthers the development of this model and provides an experimental basis for continued investigation of gammaherpesvirus pathology. PMID:12021358

  18. Large Scale Gene Expression Meta-Analysis Reveals Tissue-Specific, Sex-Biased Gene Expression in Humans

    PubMed Central

    Mayne, Benjamin T.; Bianco-Miotto, Tina; Buckberry, Sam; Breen, James; Clifton, Vicki; Shoubridge, Cheryl; Roberts, Claire T.

    2016-01-01

    The severity and prevalence of many diseases are known to differ between the sexes. Organ specific sex-biased gene expression may underpin these and other sexually dimorphic traits. To further our understanding of sex differences in transcriptional regulation, we performed meta-analyses of sex biased gene expression in multiple human tissues. We analyzed 22 publicly available human gene expression microarray data sets including over 2500 samples from 15 different tissues and 9 different organs. Briefly, by using an inverse-variance method we determined the effect size difference of gene expression between males and females. We found the greatest sex differences in gene expression in the brain, specifically in the anterior cingulate cortex, (1818 genes), followed by the heart (375 genes), kidney (224 genes), colon (218 genes), and thyroid (163 genes). More interestingly, we found different parts of the brain with varying numbers and identity of sex-biased genes, indicating that specific cortical regions may influence sexually dimorphic traits. The majority of sex-biased genes in other tissues such as the bladder, liver, lungs, and pancreas were on the sex chromosomes or involved in sex hormone production. On average in each tissue, 32% of autosomal genes that were expressed in a sex-biased fashion contained androgen or estrogen hormone response elements. Interestingly, across all tissues, we found approximately two-thirds of autosomal genes that were sex-biased were not under direct influence of sex hormones. To our knowledge this is the largest analysis of sex-biased gene expression in human tissues to date. We identified many sex-biased genes that were not under the direct influence of sex chromosome genes or sex hormones. These may provide targets for future development of sex-specific treatments for diseases. PMID:27790248

  19. Gene expression profiling in Ishikawa cells: A fingerprint for estrogen active compounds

    SciTech Connect

    Boehme, Kathleen; Simon, Stephanie

    2009-04-01

    Several anthropogenous and naturally occurring substances, referred to as estrogen active compounds (EACs), are able to interfere with hormone and in particular estrogen receptor signaling. EACs can either cause adverse health effects in humans and wildlife populations or have beneficial effects on estrogen-dependent diseases. The aim of this study was to examine global gene expression profiles in estrogen receptor (ER)-proficient Ishikawa plus and ER-deficient Ishikawa minus endometrial cancer cells treated with selected well-known EACs (Diethylstilbestrol, Genistein, Zearalenone, Resveratrol, Bisphenol A and o,p'-DDT). We also investigated the effect of the pure antiestrogen ICI 182,780 (ICI) on the expression patterns caused by these compounds. Transcript levels were quantified 24 h after compound treatment using Illumina BeadChip Arrays. We identified 87 genes with similar expression changes in response to all EAC treatments in Ishikawa plus. ICI lowered the magnitude or reversed the expression of these genes, indicating ER dependent regulation. Apart from estrogenic gene regulation, Bisphenol A, o,p'-DDT, Zearalenone, Genistein and Resveratrol displayed similarities to ICI in their expression patterns, suggesting mixed estrogenic/antiestrogenic properties. In particular, the predominant antiestrogenic expression response of Resveratrol could be clearly distinguished from the other test compounds, indicating a distinct mechanism of action. Divergent gene expression patterns of the phytoestrogens, as well as weaker estrogenic gene expression regulation determined for the anthropogenous chemicals Bisphenol A and o,p'-DDT, warrants a careful assessment of potential detrimental and/or beneficial effects of EACs. The characteristic expression fingerprints and the identified subset of putative marker genes can be used for screening chemicals with an unknown mode of action and for predicting their potential to exert endocrine disrupting effects.

  20. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies

    PubMed Central

    Kosinová, Lucie; Cahová, Monika; Fábryová, Eva; Týcová, Irena; Koblas, Tomáš; Leontovyč, Ivan; Saudek, František; Kříž, Jan

    2016-01-01

    The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3) in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0–120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48–120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information from 48 hrs

  1. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies.

    PubMed

    Kosinová, Lucie; Cahová, Monika; Fábryová, Eva; Týcová, Irena; Koblas, Tomáš; Leontovyč, Ivan; Saudek, František; Kříž, Jan

    2016-01-01

    The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3) in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0-120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48-120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information from 48 hrs onwards.

  2. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies.

    PubMed

    Kosinová, Lucie; Cahová, Monika; Fábryová, Eva; Týcová, Irena; Koblas, Tomáš; Leontovyč, Ivan; Saudek, František; Kříž, Jan

    2016-01-01

    The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3) in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0-120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48-120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information from 48 hrs onwards

  3. Global variability in gene expression and alternative splicing is modulated by mitochondrial content.

    PubMed

    Guantes, Raul; Rastrojo, Alberto; Neves, Ricardo; Lima, Ana; Aguado, Begoña; Iborra, Francisco J

    2015-05-01

    Noise in gene expression is a main determinant of phenotypic variability. Increasing experimental evidence suggests that genome-wide cellular constraints largely contribute to the heterogeneity observed in gene products. It is still unclear, however, which global factors affect gene expression noise and to what extent. Since eukaryotic gene expression is an energy demanding process, differences in the energy budget of each cell could determine gene expression differences. Here, we quantify the contribution of mitochondrial variability (a natural source of ATP variation) to global variability in gene expression. We find that changes in mitochondrial content can account for ∼50% of the variability observed in protein levels. This is the combined result of the effect of mitochondria dosage on transcription and translation apparatus content and activities. Moreover, we find that mitochondrial levels have a large impact on alternative splicing, thus modulating both the abundance and type of mRNAs. A simple mathematical model in which mitochondrial content simultaneously affects transcription rate and splicing site choice can explain the alternative splicing data. The results of this study show that mitochondrial content (and/or probably function) influences mRNA abundance, translation, and alternative splicing, which ultimately affects cellular phenotype.

  4. Global variability in gene expression and alternative splicing is modulated by mitochondrial content

    PubMed Central

    Guantes, Raul; Rastrojo, Alberto; Neves, Ricardo; Lima, Ana; Aguado, Begoña; Iborra, Francisco J.

    2015-01-01

    Noise in gene expression is a main determinant of phenotypic variability. Increasing experimental evidence suggests that genome-wide cellular constraints largely contribute to the heterogeneity observed in gene products. It is still unclear, however, which global factors affect gene expression noise and to what extent. Since eukaryotic gene expression is an energy demanding process, differences in the energy budget of each cell could determine gene expression differences. Here, we quantify the contribution of mitochondrial variability (a natural source of ATP variation) to global variability in gene expression. We find that changes in mitochondrial content can account for ∼50% of the variability observed in protein levels. This is the combined result of the effect of mitochondria dosage on transcription and translation apparatus content and activities. Moreover, we find that mitochondrial levels have a large impact on alternative splicing, thus modulating both the abundance and type of mRNAs. A simple mathematical model in which mitochondrial content simultaneously affects transcription rate and splicing site choice can explain the alternative splicing data. The results of this study show that mitochondrial content (and/or probably function) influences mRNA abundance, translation, and alternative splicing, which ultimately affects cellular phenotype. PMID:25800673

  5. Global variability in gene expression and alternative splicing is modulated by mitochondrial content.

    PubMed

    Guantes, Raul; Rastrojo, Alberto; Neves, Ricardo; Lima, Ana; Aguado, Begoña; Iborra, Francisco J

    2015-05-01

    Noise in gene expression is a main determinant of phenotypic variability. Increasing experimental evidence suggests that genome-wide cellular constraints largely contribute to the heterogeneity observed in gene products. It is still unclear, however, which global factors affect gene expression noise and to what extent. Since eukaryotic gene expression is an energy demanding process, differences in the energy budget of each cell could determine gene expression differences. Here, we quantify the contribution of mitochondrial variability (a natural source of ATP variation) to global variability in gene expression. We find that changes in mitochondrial content can account for ∼50% of the variability observed in protein levels. This is the combined result of the effect of mitochondria dosage on transcription and translation apparatus content and activities. Moreover, we find that mitochondrial levels have a large impact on alternative splicing, thus modulating both the abundance and type of mRNAs. A simple mathematical model in which mitochondrial content simultaneously affects transcription rate and splicing site choice can explain the alternative splicing data. The results of this study show that mitochondrial content (and/or probably function) influences mRNA abundance, translation, and alternative splicing, which ultimately affects cellular phenotype. PMID:25800673

  6. Cell cycle gene expression under clinorotation

    NASA Astrophysics Data System (ADS)

    Artemenko, Olga

    2016-07-01

    Cyclins and cyclin-dependent kinase (CDK) are main regulators of the cell cycle of eukaryotes. It's assumes a significant change of their level in cells under microgravity conditions and by other physical factors actions. The clinorotation use enables to determine the influence of gravity on simulated events in the cell during the cell cycle - exit from the state of quiet stage and promotion presynthetic phase (G1) and DNA synthesis phase (S) of the cell cycle. For the clinorotation effect study on cell proliferation activity is the necessary studies of molecular mechanisms of cell cycle regulation and development of plants under altered gravity condition. The activity of cyclin D, which is responsible for the events of the cell cycle in presynthetic phase can be controlled by the action of endogenous as well as exogenous factors, but clinorotation is one of the factors that influence on genes expression that regulate the cell cycle.These data can be used as a model for further research of cyclin - CDK complex for study of molecular mechanisms regulation of growth and proliferation. In this investigation we tried to summarize and analyze known literature and own data we obtained relatively the main regulators of the cell cycle in altered gravity condition.

  7. Assembly and Expression of Shark Ig Genes.

    PubMed

    Hsu, Ellen

    2016-05-01

    Sharks are modern descendants of the earliest vertebrates possessing Ig superfamily receptor-based adaptive immunity. They respond to immunogen with Abs that, upon boosting, appear more rapidly and show affinity maturation. Specific Abs and immunological memory imply that Ab diversification and clonal selection exist in cartilaginous fish. Shark Ag receptors are generated through V(D)J recombination, and because it is a mechanism known to generate autoreactive receptors, this implies that shark lymphocytes undergo selection. In the mouse, the ∼2.8-Mb IgH and IgL loci require long-range, differential activation of component parts for V(D)J recombination, allelic exclusion, and receptor editing. These processes, including class switching, evolved with and appear inseparable from the complex locus organization. In contrast, shark Igs are encoded by 100-200 autonomously rearranging miniloci. This review describes how the shark primary Ab repertoire is generated in the absence of structural features considered essential in mammalian Ig gene assembly and expression. PMID:27183649

  8. Redox regulation of photosynthetic gene expression

    PubMed Central

    Queval, Guillaume; Foyer, Christine H.

    2012-01-01

    Redox chemistry and redox regulation are central to the operation of photosynthesis and respiration. However, the roles of different oxidants and antioxidants in the regulation of photosynthetic or respiratory gene expression remain poorly understood. Leaf transcriptome profiles of a range of Arabidopsis thaliana genotypes that are deficient in either hydrogen peroxide processing enzymes or in low molecular weight antioxidant were therefore compared to determine how different antioxidant systems that process hydrogen peroxide influence transcripts encoding proteins targeted to the chloroplasts or mitochondria. Less than 10 per cent overlap was observed in the transcriptome patterns of leaves that are deficient in either photorespiratory (catalase (cat)2) or chloroplastic (thylakoid ascorbate peroxidase (tapx)) hydrogen peroxide processing. Transcripts encoding photosystem II (PSII) repair cycle components were lower in glutathione-deficient leaves, as were the thylakoid NAD(P)H (nicotinamide adenine dinucleotide (phosphate)) dehydrogenases (NDH) mRNAs. Some thylakoid NDH mRNAs were also less abundant in tAPX-deficient and ascorbate-deficient leaves. Transcripts encoding the external and internal respiratory NDHs were increased by low glutathione and low ascorbate. Regulation of transcripts encoding specific components of the photosynthetic and respiratory electron transport chains by hydrogen peroxide, ascorbate and glutathione may serve to balance non-cyclic and cyclic electron flow pathways in relation to oxidant production and reductant availability. PMID:23148274

  9. Gene expression profiling of anticancer immune responses.

    PubMed

    Wang, Ena; Panelli, Monica C; Monsurró, Vladia; Marincola, Francesco M

    2004-06-01

    Anticancer immune responses can be enhanced by immune manipulation, however, the biological mechanism responsible for these immune responses remains largely unexplained. Conventional immunology researchers have extensively studied specific interactions between immune and cancer cells, and additional investigations have identified co-factors that may enhance the effectiveness of such interactions. As the molecular understanding of individual interactions increases, it is becoming apparent that no single mechanism can explain the phenomenon of tumor rejection. The contribution of several components of the innate and adaptive immune response is likely to be required for successful tumor rejection. These components may be variably recruited and activated by molecules with immune modulatory properties being produced by tumor and bystander cells within the tumor micro-environment. Such complexity can only be appreciated and solved by high-throughput tools capable of providing a global view of biological processes as they occur. This review will present selected examples of how high-throughput gene expression profiling may contribute to the understanding of anticancer immune responses. As reviews on technological aspects of the genomic analysis of cancer are already available, this review will provide a speculative discussion about their potential usefulness.

  10. Unpredictable neonatal stress enhances adult anxiety and alters amygdala gene expression related to serotonin and GABA

    PubMed Central

    Sarro, Emma C; Sullivan, Regina M; Barr, Gordon

    2014-01-01

    Anxiety-related disorders are among the most common psychiatric illnesses, thought to have both genetic and environmental causes. Early-life trauma, such as abuse from a caregiver, can be predictable or unpredictable, each resulting in increased prevalence and severity of a unique set of disorders. In this study, we examined the influence of early unpredictable trauma on both the behavioral expression of adult anxiety and gene expression within the amygdala. Neonatal rats were exposed to unpaired odor-shock conditioning for 5 days, which produces deficits in adult behavior and amygdala dysfunction. In adulthood, we used the Light/Dark box test to measure anxiety-related behaviors, measuring the latency to enter the lit area and quantified urination and defecation. The amygdala was then dissected and a microarray analysis was performed to examine changes in gene expression. Animals that had received early unpredictable trauma displayed significantly longer latencies to enter the lit area and more defecation and urination. The microarray analysis revealed over-represented genes related to learning and memory, synaptic transmission and trans-membrane transport. Gene ontology and pathway analysis identified highly represented disease states related to anxiety phenotypes, including social anxiety, obsessive-compulsive disorders, PTSD and bipolar disorder. Addiction related genes were also overrepresented in this analysis. Unpredictable shock during early development increased anxiety-like behaviors in adulthood with concomitant changes in genes related to neurotransmission, resulting in gene expression patterns similar to anxiety-related psychiatric disorders. PMID:24240029

  11. Profiling gene expression in whole blood samples following an in-vitro challenge.

    PubMed

    Spijker, Sabine; van de Leemput, Joyce C H; Hoekstra, Chantal; Boomsma, Dorret I; Smit, August B

    2004-12-01

    Genomics tools (gene- and protein-expression studies) can be used to find possible target genes involved in a quantifiable trait or disease state. However in many instances, cells and tissues directly involved in the trait's expression, for example, brain tissue, are not amenable for gene expression analysis. Whole blood cells share a molecular make-up for cellular communication and gene regulation systems with many other cell types, for example, neuronal cells, and have the advantage of being very accessible for gene profiling. We investigated the feasibility of nationwide blood sample collection for lymphocyte RNA isolation and real-time PCR analysis to quantify genomic responses. We tested several designs for blood collection and storage: blood sampling in PAXgene blood collection tubes and storage at -20 degrees C, blood sampling in heparin tubes and decanting the samples (with or without in-vitro stimulus) into either PAXgene blood collection tubes and storage at -20 degrees C, or polypropylene tubes followed by snap-freezing and storage at -80 degrees C. The latter procedure is the best cost-wise when only small amounts of total RNA are needed for downstream applications. Lymphocyte gene expression studies are most likely hampered by the quality of isolated RNA rather than the sampling method. We show that large-scale nationwide sample collections did not alter RNA quality or gene expression levels when compared to sampling and processing in a more controlled way. To this end, we present an optimized protocol for easy and standardized isolation of high quality RNA using the PAXgene isolation kit. Based on these results, we suggest that whole blood genomic data can be used as a genomic probe in experimental and clinical research.

  12. Expression of nisin genes in cheese--a quantitative real-time polymerase chain reaction approach.

    PubMed

    Trmčić, A; Monnet, C; Rogelj, I; Bogovič Matijašić, B

    2011-01-01

    The role of bacteriocins in different environments has not been thoroughly explained, mainly because of the difficulties related to the detection of their production. Nisin, an antimicrobial peptide produced by Lactococcus lactis has a long history of safe use in food products and has been studied from many aspects of genetics, biosynthesis, immunity, regulation, and mode of action. Still, some aspects concerning the dynamics of nisin gene expression remain unknown, especially in complex media like cheese. The main objective of the present study was to quantify in a cheese-like medium the expression of nisin genes in L. lactis M78, a well-characterized nisin A producer isolated from raw milk. The expression of all 11 genes involved in nisin biosynthesis was evaluated during cheese production by real-time reverse transcription-PCR. Total RNA was extracted from cheeses using a direct extraction method without prior separation of microbial cells. The M78 strain grew well in experimental cheeses, producing detectable amounts of nisin after 4 h of fermentation. The presence of nisin as an activator modified both the expression of nisin genes and the accumulation of active nisin. Four groups could be distinguished based on gene expression as a function of time: nisA, nisFEG, nisRK and nisBTCIP. Based on nisin-producing strain growth, nisin activity, function of nisin genes, and their location, correlations were established that contribute to the explanation of regulation of nisin biosynthesis and immunity. This study is the first in which the evolution of bacteriocin gene transcripts has been quantified rigorously in a cheese-like medium.

  13. Sex-Specific Selection and Sex-Biased Gene Expression in Humans and Flies

    PubMed Central

    Kirkpatrick, Mark

    2016-01-01

    Sexual dimorphism results from sex-biased gene expression, which evolves when selection acts differently on males and females. While there is an intimate connection between sex-biased gene expression and sex-specific selection, few empirical studies have studied this relationship directly. Here we compare the two on a genome-wide scale in humans and flies. We find a distinctive “Twin Peaks” pattern in humans that relates the strength of sex-specific selection, quantified by genetic divergence between male and female adults at autosomal loci, to the degree of sex-biased expression. Genes with intermediate degrees of sex-biased expression show evidence of ongoing sex-specific selection, while genes with either little or completely sex-biased expression do not. This pattern apparently results from differential viability selection in males and females acting in the current generation. The Twin Peaks pattern is also found in Drosophila using a different measure of sex-specific selection acting on fertility. We develop a simple model that successfully recapitulates the Twin Peaks. Our results suggest that many genes with intermediate sex-biased expression experience ongoing sex-specific selection in humans and flies. PMID:27658217

  14. The Role of Multiple Transcription Factors In Archaeal Gene Expression

    SciTech Connect

    Charles J. Daniels

    2008-09-23

    Since the inception of this research program, the project has focused on two central questions: What is the relationship between the 'eukaryal-like' transcription machinery of archaeal cells and its counterparts in eukaryal cells? And, how does the archaeal cell control gene expression using its mosaic of eukaryal core transcription machinery and its bacterial-like transcription regulatory proteins? During the grant period we have addressed these questions using a variety of in vivo approaches and have sought to specifically define the roles of the multiple TATA binding protein (TBP) and TFIIB-like (TFB) proteins in controlling gene expression in Haloferax volcanii. H. volcanii was initially chosen as a model for the Archaea based on the availability of suitable genetic tools; however, later studies showed that all haloarchaea possessed multiple tbp and tfb genes, which led to the proposal that multiple TBP and TFB proteins may function in a manner similar to alternative sigma factors in bacterial cells. In vivo transcription and promoter analysis established a clear relationship between the promoter requirements of haloarchaeal genes and those of the eukaryal RNA polymerase II promoter. Studies on heat shock gene promoters, and the demonstration that specific tfb genes were induced by heat shock, provided the first indication that TFB proteins may direct expression of specific gene families. The construction of strains lacking tbp or tfb genes, coupled with the finding that many of these genes are differentially expressed under varying growth conditions, provided further support for this model. Genetic tools were also developed that led to the construction of insertion and deletion mutants, and a novel gene expression scheme was designed that allowed the controlled expression of these genes in vivo. More recent studies have used a whole genome array to examine the expression of these genes and we have established a linkage between the expression of specific tfb

  15. Arabidopsis gene expression patterns are altered during spaceflight

    NASA Astrophysics Data System (ADS)

    Paul, Anna-Lisa; Popp, Michael P.; Gurley, William B.; Guy, Charles; Norwood, Kelly L.; Ferl, Robert J.

    The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments results in differential gene expression. A 5-day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β-Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on gene expression patterns initially by using the Adh/GUS transgene to address specifically the possibility that spaceflight induces a hypoxic stress response (Paul, A.L., Daugherty, C.J., Bihn, E.A., Chapman, D.K., Norwood, K.L., Ferl, R.J., 2001. Transgene expression patterns indicate that spaceflight affects stress signal perception and transduction in arabidopsis, Plant Physiol. 126, 613-621). As a follow-on to the reporter gene analysis, we report here the evaluation of genome-wide patterns of native gene expression within Arabidopsis shoots utilizing the Agilent DNA array of 21,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes was further characterized with quantitative Real-Time RT PCR (ABI - Taqman®). Comparison of the patterns of expression for arrays probed with RNA isolated from plants exposed to spaceflight compared to RNA isolated from ground control plants revealed 182 genes that were differentially expressed in response to the spaceflight mission by more than 4-fold, and of those only 50 genes were expressed at levels chosen to support a conservative change call. None of the genes that are hallmarks of hypoxic stress were induced to this level. However, genes related to heat shock were dramatically induced - but in a pattern and under growth conditions that are not easily explained by elevated temperatures. These gene expression data are discussed in light of current models for plant responses to the spaceflight environment and with regard to potential future spaceflight experiment

  16. Microdissection of the gene expression codes driving nephrogenesis

    PubMed Central

    Brunskill, Eric W; Patterson, Larry T

    2010-01-01

    The kidney represents an excellent model system for learning the principles of organogenesis. It is intermediate in complexity, and employs many commonly used developmental processes. As such, kidney development has been the subject of intensive study, using a variety of techniques, including in situ hybridization, organ culture and gene targeting, revealing many critical genes and pathways. Nevertheless, proper organogenesis requires precise patterns of cell type specific differential gene expression, involving very large numbers of genes. This review is focused on the use of global profiling technologies to create an atlas of gene expression codes driving development of different mammalian kidney compartments. Such an atlas allows one to select a gene of interest, and to determine its expression level in each element of the developing kidney, or to select a structure of interest, such as the renal vesicle, and to examine its complete gene expression state. Novel component specific molecular markers are identified, and the changing waves of gene expression that drive nephrogenesis are defined. As the tools continue to improve for the purification of specific cell types and expression profiling of even individual cells it is possible to predict an atlas of gene expression during kidney development that extends to single cell resolution. PMID:21220959

  17. Microdissection of the gene expression codes driving nephrogenesis.

    PubMed

    Potter, S Steven; Brunskill, Eric W; Patterson, Larry T

    2010-01-01

    The kidney represents an excellent model system for learning the principles of organogenesis. It is intermediate in complexity, and employs many commonly used developmental processes. As such, kidney development has been the subject of intensive study, using a variety of techniques, including in situ hybridization, organ culture and gene targeting, revealing many critical genes and pathways. Nevertheless, proper organogenesis requires precise patterns of cell type specific differential gene expression, involving very large numbers of genes. This review is focused on the use of global profiling technologies to create an atlas of gene expression codes driving development of different mammalian kidney compartments. Such an atlas allows one to select a gene of interest, and to determine its expression level in each element of the developing kidney, or to select a structure of interest, such as the renal vesicle, and to examine its complete gene expression state. Novel component specific molecular markers are identified, and the changing waves of gene expression that drive nephrogenesis are defined. As the tools continue to improve for the purification of specific cell types and expression profiling of even individual cells it is possible to predict an atlas of gene expression during kidney development that extends to single cell resolution. PMID:21220959

  18. The role of gene expression in ecological speciation

    PubMed Central

    Pavey, Scott A; Collin, Hélène; Nosil, Patrik; Rogers, Sean M

    2010-01-01

    Ecological speciation is the process by which barriers to gene flow between populations evolve due to adaptive divergence via natural selection. A relatively unexplored area in ecological speciation is the role of gene expression. Gene expression may be associated with ecologically important phenotypes not evident from morphology and play a role during colonization of new environments. Here we review two potential roles of gene expression in ecological speciation: (1) its indirect role in facilitating population persistence and (2) its direct role in contributing to genetically based reproductive isolation. We find indirect evidence that gene expression facilitates population persistence, but direct tests are lacking. We also find clear examples of gene expression having effects on phenotypic traits and adaptive genetic divergence, but links to the evolution of reproductive isolation itself remain indirect. Gene expression during adaptive divergence seems to often involve complex genetic architectures controlled by gene networks, regulatory regions, and “eQTL hotspots.” Nonetheless, we review how approaches for isolating the functional mutations contributing to adaptive divergence are proving to be successful. The study of gene expression has promise for increasing our understanding ecological speciation, particularly when integrative approaches are applied. PMID:20860685

  19. Characterizing bacterial gene expression in nitrogen cycle metabolism with RT-qPCR.

    PubMed

    Graham, James E; Wantland, Nicholas B; Campbell, Mark; Klotz, Martin G

    2011-01-01

    Recent advances in DNA sequencing have greatly accelerated our ability to obtain the raw information needed to recognize both known and potential novel modular microbial genomic capacity for nitrogen metabolism. With PCR-based approaches to quantifying microbial mRNA expression now mainstream in most laboratories, researchers can now more efficiently propose and test hypotheses on the contributions of individual microbes to the biological accessibility of nitrogen upon which all other life depends. We review known microbial roles in these key nitrogen transformations, and describe the necessary steps in carrying out relevant gene expression studies. An example experimental design is then provided characterizing Nitrosococcus oceani mRNA expression in cultures responding to ammonia. The approach described, that of assessing microbial genome inventory and testing putative modular gene expression by mRNA quantification, is likely to remain an important tool in understanding individual microbial contributions within microbial community activities that maintain the Earth's nitrogen balance.

  20. An Exercise to Estimate Differential Gene Expression in Human Cells

    ERIC Educational Resources Information Center

    Chaudhry, M. Ahmad

    2006-01-01

    The expression of genes in cells of various tissue types varies considerably and is correlated with the function of a particular organ. The pattern of gene expression changes in diseased tissues, in response to therapy or infection and exposure to environmental mutagens, chemicals, ultraviolet light, and ionizing radiation. To better understand…

  1. MEPD: medaka expression pattern database, genes and more

    PubMed Central

    Alonso-Barba, Juan I.; Rahman, Raza-Ur; Wittbrodt, Joachim; Mateo, Juan L.

    2016-01-01

    The Medaka Expression Pattern Database (MEPD; http://mepd.cos.uni-heidelberg.de/) is designed as a repository of medaka expression data for the scientific community. In this update we present two main improvements. First, we have changed the previous clone-centric view for in situ data to a gene-centric view. This is possible because now we have linked all the data present in MEPD to the medaka gene annotation in ENSEMBL. In addition, we have also connected the medaka genes in MEPD to their corresponding orthologous gene in zebrafish, again using the ENSEMBL database. Based on this, we provide a link to the Zebrafish Model Organism Database (ZFIN) to allow researches to compare expression data between these two fish model organisms. As a second major improvement, we have modified the design of the database to enable it to host regulatory elements, promoters or enhancers, expression patterns in addition to gene expression. The combination of gene expression, by traditional in situ, and regulatory element expression, typically by fluorescence reporter gene, within the same platform assures consistency in terms of annotation. In our opinion, this will allow researchers to uncover new insights between the expression domain of genes and their regulatory landscape. PMID:26450962

  2. MEPD: medaka expression pattern database, genes and more.

    PubMed

    Alonso-Barba, Juan I; Rahman, Raza-Ur; Wittbrodt, Joachim; Mateo, Juan L

    2016-01-01

    The Medaka Expression Pattern Database (MEPD; http://mepd.cos.uni-heidelberg.de/) is designed as a repository of medaka expression data for the scientific community. In this update we present two main improvements. First, we have changed the previous clone-centric view for in situ data to a gene-centric view. This is possible because now we have linked all the data present in MEPD to the medaka gene annotation in ENSEMBL. In addition, we have also connected the medaka genes in MEPD to their corresponding orthologous gene in zebrafish, again using the ENSEMBL database. Based on this, we provide a link to the Zebrafish Model Organism Database (ZFIN) to allow researches to compare expression data between these two fish model organisms. As a second major improvement, we have modified the design of the database to enable it to host regulatory elements, promoters or enhancers, expression patterns in addition to gene expression. The combination of gene expression, by traditional in situ, and regulatory element expression, typically by fluorescence reporter gene, within the same platform assures consistency in terms of annotation. In our opinion, this will allow researchers to uncover new insights between the expression domain of genes and their regulatory landscape. PMID:26450962

  3. Gene Expression Profiling in the Hibernating Primate, Cheirogaleus Medius.

    PubMed

    Faherty, Sheena L; Villanueva-Cañas, José Luis; Klopfer, Peter H; Albà, M Mar; Yoder, Anne D

    2016-01-01

    Hibernation is a complex physiological response that some mammalian species employ to evade energetic demands. Previous work in mammalian hibernators suggests that hibernation is activated not by a set of genes unique to hibernators, but by differential expression of genes that are present in all mammals. This question of universal genetic mechanisms requires further investigation and can only be tested through additional investigations of phylogenetically dispersed species. To explore this question, we use RNA-Seq to investigate gene expression dynamics as they relate to the varying physiological states experienced throughout the year in a group of primate hibernators-Madagascar's dwarf lemurs (genus Cheirogaleus). In a novel experimental approach, we use longitudinal sampling of biological tissues as a method for capturing gene expression profiles from the same individuals throughout their annual hibernation cycle. We identify 90 candidate genes that have variable expression patterns when comparing two active states (Active 1 and Active 2) with a torpor state. These include genes that are involved in metabolic pathways, feeding behavior, and circadian rhythms, as might be expected to correlate with seasonal physiological state changes. The identified genes appear to be critical for maintaining the health of an animal that undergoes prolonged periods of metabolic depression concurrent with the hibernation phenotype. By focusing on these differentially expressed genes in dwarf lemurs, we compare gene expression patterns in previously studied mammalian hibernators. Additionally, by employing evolutionary rate analysis, we find that hibernation-related genes do not evolve under positive selection in hibernating species relative to nonhibernators. PMID:27412611

  4. Gene Expression Measurement Module (GEMM) - a fully automated, miniaturized instrument for measuring gene expression in space

    NASA Astrophysics Data System (ADS)

    Karouia, Fathi; Ricco, Antonio; Pohorille, Andrew; Peyvan, Kianoosh

    2012-07-01

    The capability to measure gene expression on board spacecrafts opens the doors to a large number of experiments on the influence of space environment on biological systems that will profoundly impact our ability to conduct safe and effective space travel, and might also shed light on terrestrial physiology or biological function and human disease and aging processes. Measurements of gene expression will help us to understand adaptation of terrestrial life to conditions beyond the planet of origin, identify deleterious effects of the space environment on a wide range of organisms from microbes to humans, develop effective countermeasures against these effects, determine metabolic basis of microbial pathogenicity and drug resistance, test our ability to sustain and grow in space organisms that can be used for life support and in situ resource utilization during long-duration space exploration, and monitor both the spacecraft environment and crew health. These and other applications hold significant potential for discoveries in space biology, biotechnology and medicine. Accordingly, supported by funding from the NASA Astrobiology Science and Technology Instrument Development Program, we are developing a fully automated, miniaturized, integrated fluidic system for small spacecraft capable of in-situ measuring microbial expression of thousands of genes from multiple samples. The instrument will be capable of (1) lysing bacterial cell walls, (2) extracting and purifying RNA released from cells, (3) hybridizing it on a microarray and (4) providing electrochemical readout, all in a microfluidics cartridge. The prototype under development is suitable for deployment on nanosatellite platforms developed by the NASA Small Spacecraft Office. The first target application is to cultivate and measure gene expression of the photosynthetic bacterium Synechococcus elongatus, i.e. a cyanobacterium known to exhibit remarkable metabolic diversity and resilience to adverse conditions

  5. Efficient shRNA-mediated inhibition of gene expression in zebrafish.

    PubMed

    De Rienzo, Gianluca; Gutzman, Jennifer H; Sive, Hazel

    2012-09-01

    Despite the broad repertoire of loss of function (LOF) tools available for use in the zebrafish, there remains a need for a simple and rapid method that can inhibit expression of genes at later stages. RNAi would fulfill that role, and a previous report (Dong et al. 2009) provided encouraging data. The goal of this study was to further address the ability of expressed shRNAs to inhibit gene expression. This included quantifying RNA knockdown, testing specificity of shRNA effects, and determining whether tissue-specific LOF could be achieved. Using an F0 transgenic approach, this report demonstrates that for two genes, wnt5b and zDisc1, each with described mutant and morphant phenotypes, shRNAs efficiently decrease endogenous RNA levels. Phenotypes elicited by shRNA resemble those of mutants and morphants, and are reversed by expression of cognate RNA, further demonstrating specificity. Tissue-specific expression of zDisc1 shRNAs in F0 transgenics demonstrates that conditional LOF can be readily obtained. These results suggest that shRNA expression presents a viable approach for rapid inhibition of zebrafish gene expression.

  6. Expression of VEGF and Semaphorin Genes Define Subgroups of Triple Negative Breast Cancer

    PubMed Central

    Bender, R. Joseph; Mac Gabhann, Feilim

    2013-01-01

    Triple negative breast cancers (TNBC) are difficult to treat due to a lack of targets and heterogeneity. Inhibition of angiogenesis is a promising therapeutic strategy, but has had limited effectiveness so far in breast cancer. To quantify heterogeneity in angiogenesis-related gene expression in breast cancer, we focused on two families – VEGFs and semaphorins – that compete for neuropilin co-receptors on endothelial cells. We compiled microarray data for over 2,600 patient tumor samples and analyzed the expression of VEGF- and semaphorin-related ligands and receptors. We used principal component analysis to identify patterns of gene expression, and clustering to group samples according to these patterns. We used available survival data to determine whether these clusters had prognostic as well as therapeutic relevance. TNBC was highly associated with dysregulation of VEGF- and semaphorin-related genes; in particular, it appeared that expression of both VEGF and semaphorin genes were altered in a pro-angiogenesis direction. A pattern of high VEGFA expression with low expression of secreted semaphorins was associated with 60% of triple-negative breast tumors. While all TNBC groups demonstrated poor prognosis, this signature also correlated with lower 5-year survival rates in non-TNBC samples. A second TNBC pattern, including high VEGFC expression, was also identified. These pro-angiogenesis signatures may identify cancers that are more susceptible to VEGF inhibition. PMID:23667446

  7. The effect of negative autoregulation on eukaryotic gene expression

    NASA Astrophysics Data System (ADS)

    Nevozhay, Dmitry; Adams, Rhys; Murphy, Kevin; Josic, Kresimir; Balázsi, G. Ábor

    2009-03-01

    Negative autoregulation is a frequent motif in gene regulatory networks, which has been studied extensively in prokaryotes. Nevertheless, some effects of negative feedback on gene expression in eukaryotic transcriptional networks remain unknown. We studied how the strength of negative feedback regulation affects the characteristics of gene expression in yeast cells carrying synthetic transcriptional cascades. We observed a drastic reduction of gene expression noise and a change in the shape of the dose-response curve. We explained these experimentally observed effects by stochastic simulations and a simple set of algebraic equations.

  8. Features of Gene Expression of Bacillus pumilus Metalloendopeptidase.

    PubMed

    Rudakova, N L; Sabirova, A R; Balaban, N P; Tikhonova, A O; Sharipova, M R

    2016-08-01

    Features of gene expression of the secreted Bacillus pumilus metalloendopeptidase belonging to the adamalysin/reprolysin family were investigated. In the regulatory region of the gene, we identified hypothetical binding sites for transcription factors CcpA and TnrA. We found that the expression of the metalloendopeptidase gene is controlled by mechanisms of carbon and nitrogen catabolite repression. In experiments involving nitrogen metabolism regulatory protein mutant strains, we found that the control of the metalloendopeptidase gene expression involves proteins of ammonium transport GlnK and AmtB interacting with the TnrA-regulator. PMID:27677556

  9. Direct Introduction of Genes into Rats and Expression of the Genes

    NASA Astrophysics Data System (ADS)

    Benvenisty, Nissim; Reshef, Lea

    1986-12-01

    A method of introducing actively expressed genes into intact mammals is described. DNA precipitated with calcium phosphate has been injected intraperitoneally into newborn rats. The injected genes have been taken up and expressed by the animal tissues. To examine the generality of the method we have injected newborn rats with the chloramphenicol acetyltransferase prokaryotic gene fused with various viral and cellular gene promoters and the gene for hepatitis B surface antigen, and we observed appearance of chloramphenicol acetyltransferase activity and hepatitis B surface antigen in liver and spleen. In addition, administration of genes coding for hormones (insulin or growth hormone) resulted in their expression.

  10. Correlation between gene expression and GO semantic similarity.

    PubMed

    Sevilla, José L; Segura, Víctor; Podhorski, Adam; Guruceaga, Elizabeth; Mato, José M; Martínez-Cruz, Luis A; Corrales, Fernando J; Rubio, Angel

    2005-01-01

    This research analyzes some aspects of the relationship between gene expression, gene function, and gene annotation. Many recent studies are implicitly based on the assumption that gene products that are biologically and functionally related would maintain this similarity both in their expression profiles as well as in their Gene Ontology (GO) annotation. We analyze how accurate this assumption proves to be using real publicly available data. We also aim to validate a measure of semantic similarity for GO annotation. We use the Pearson correlation coefficient and its absolute value as a measure of similarity between expression profiles of gene products. We explore a number of semantic similarity measures (Resnik, Jiang, and Lin) and compute the similarity between gene products annotated using the GO. Finally, we compute correlation coefficients to compare gene expression similarity against GO semantic similarity. Our results suggest that the Resnik similarity measure outperforms the others and seems better suited for use in Gene Ontology. We also deduce that there seems to be correlation between semantic similarity in the GO annotation and gene expression for the three GO ontologies. We show that this correlation is negligible up to a certain semantic similarity value; then, for higher similarity values, the relationship trend becomes almost linear. These results can be used to augment the knowledge provided by clustering algorithms and in the development of bioinformatic tools for finding and characterizing gene products.

  11. Stable Somatic Gene Expression in Mouse Lungs Following Electroporation-mediated Tol2 Transposon Delivery.

    PubMed

    Muliawan, Hary Sakti; Nakayama, Kazuhiko; Yagi, Keiko; Ikeda, Koji; Yagita, Kazuhiro; Hirata, Ken-ichi; Emoto, Noriaki

    2015-10-07

    Gene delivery to the lung has rapidly progressed as an important method for studying various chronic lung diseases. Viral vectors, albeit highly efficient, are limited by the host immune response. Electroporation, a well-known non-viral method, can efficiently deliver genes to the lung, but is unable to induce stable gene expression. The Tol2 transposon is another non-viral method that can induce stable gene expression by reinserting its genes into the host genome. In this study, we combined electroporation and Tol2 transposons to obtain stable, high-level gene expression in the mouse lung. Tol2 transposon plasmids (pT2A-EGFP; Tol2, pCAGGS-TP; transposase) were optimized in vitro, and the electroporation procedure (pCAG-EGFP) was optimized in mouse lungs. After optimization, a combination of electroporation plus the Tol2 transposon was used in a comparative analysis with electroporation plus pCAG-EGFP. GFP expression levels were quantified and visualized on days 4 and 7 post-electroporation. We successfully reproduced the Tol2 transposon system in vitro and the electroporation procedure in vivo. We observed sustainable GFP expression using electroporation plus the Tol2 transposon on days 4 and 7, while electroporation plus pCAG-EGFP resulted in decreased GFP expression on day 7. We were able to induce high-level, stable gene expression in mouse lungs using a combination of electroporation and the Tol2 transposon. This represents a safer method for lung gene delivery that can be used as an alternative to viral vectors.

  12. Distribution of population-averaged observables in stochastic gene expression.

    PubMed

    Bhattacharyya, Bhaswati; Kalay, Ziya

    2014-01-01

    Observation of phenotypic diversity in a population of genetically identical cells is often linked to the stochastic nature of chemical reactions involved in gene regulatory networks. We investigate the distribution of population-averaged gene expression levels as a function of population, or sample, size for several stochastic gene expression models to find out to what extent population-averaged quantities reflect the underlying mechanism of gene expression. We consider three basic gene regulation networks corresponding to transcription with and without gene state switching and translation. Using analytical expressions for the probability generating function of observables and large deviation theory, we calculate the distribution and first two moments of the population-averaged mRNA and protein levels as a function of model parameters, population size, and number of measurements contained in a data set. We validate our results using stochastic simulations also report exact results on the asymptotic properties of population averages which show qualitative differences among different models. PMID:24580265

  13. Fundamental principles of energy consumption for gene expression.

    PubMed

    Huang, Lifang; Yuan, Zhanjiang; Yu, Jianshe; Zhou, Tianshou

    2015-12-01

    How energy is consumed in gene expression is largely unknown mainly due to complexity of non-equilibrium mechanisms affecting expression levels. Here, by analyzing a representative gene model that considers complexity of gene expression, we show that negative feedback increases energy consumption but positive feedback has an opposite effect; promoter leakage always reduces energy consumption; generating more bursts needs to consume more energy; and the speed of promoter switching is at the cost of energy consumption. We also find that the relationship between energy consumption and expression noise is multi-mode, depending on both the type of feedback and the speed of promoter switching. Altogether, these results constitute fundamental principles of energy consumption for gene expression, which lay a foundation for designing biologically reasonable gene modules. In addition, we discuss possible biological implications of these principles by combining experimental facts.

  14. An Mpeg (macrophage expressed gene) from the Pacific oyster Crassostrea gigas: molecular characterization and gene expression.

    PubMed

    He, Xiaocui; Zhang, Yang; Yu, Ziniu

    2011-03-01

    Mpegs (macrophage expressed genes) encode members of the MACPF (membrane-attack complex/perforin) protein superfamily that play essential roles in innate immunity. In the present study, a homolog of Mpeg1 was identified in Crassostrea gigas and designed Cg-Mpeg1. The complete cDNA of Cg-Mpeg1 is 2781 bp in length, containing an ORF of 2226 bp, which encodes a putative protein of 742 amino acids with a predicted 19-aa hydrophobic signal peptide, an MACPF domain, and a transmembrane domain. Phylogenetic analysis shows that Cg-Mpeg1 is similar to other mollusk MACPF proteins and might originate in an ancient ancestor gene before the divergence of protostomes and deuterostomes. Localization study revealed that Cg-Mpeg1 protein is found primarily in late endosomes. The MACPF domain from Cg-Mpeg1 exhibits significant antibacterial activity to both Gram-negative and positive bacteria. Furthermore, Real-time Quantitative PCR analysis showed that Cg-Mpeg1 is expressed in all tissues detected with highest expression in gill and gonads. Moreover, Mpeg1 mRNA levels are significantly up-regulated following infection with Vibrio alginolyticus. These results highlight that Cg-Mpeg1 plays an essential role in host defense and elimination of pathogens in C. gigas.

  15. Correlation of protein and gene expression profiles of inflammatory proteins after endotoxin challenge in human subjects.

    PubMed

    Prabhakar, Uma; Conway, Theresa M; Murdock, Paul; Mooney, Jeff L; Clark, Steve; Hedge, Priti; Bond, Brian C; Jazwinska, Elizabeth C; Barnes, Michael R; Tobin, Frank; Damian-Iordachi, Valeriu; Greller, Larry; Hurle, Mark; Stubbs, Andrew P; Li, Zhong; Valoret, Elizabeth I; Erickson-Miller, Connie; Cass, Lisa; Levitt, Blanche; Davis, Hugh M; Jorkasky, Diane K; Williams, William V

    2005-07-01

    Administration of endotoxin (LPS) in humans results in profound physiological responses, including activation of peripheral blood mononuclear cells and the release of inflammatory factors. The time course of the response of selected inflammatory proteins was examined in healthy subjects (n = 6) administered a single intravenous dose of the purified derivative of endotoxin (3.0 ng/kg). Microarray analysis demonstrated changes in the expression of a number of genes, which were confirmed in separate in vitro endotoxin stimulation experiments. Subsequent TaqMan analysis of genes of interest indicated time-dependent changes in the expression of many of these genes. This included pre-B cell enhancing factor, which was identified on microarray analysis as being markedly upregulated following endotoxin stimulation. Protein expression of the genes examined by TaqMan analysis was measured and demonstrated the appearance of tumor necrosis factor (TNF)-alpha and sTNF-R proteins in the plasma beginning within 1 h after dosing, followed by other cytokines/ inflammatory markers (e.g., IL-1ra, G-CSF, IL-6, IL-8, and IL-10) and suppressors of cytokine signaling (SOCS-1 and SOCS-3). In general, cytokine protein expression correlated well with gene expression; however, the temporal profile of expression of some genes did not correlate well with the protein data. For many of these proteins, the lack of correlation was attributable to alternate tissue sources, which were demonstrated on TaqMan analysis. Principal component analysis indicated that cytokines could be grouped according to their temporal pattern of response, with most transcript levels returning to baseline 24 h following endotoxin administration. The combination of cDNA microarray and TaqMan analysis to identify and quantify changes in gene expression, along with the analysis of protein expression, can be useful in investigating inflammatory and other diseases.

  16. Distinguishing the Time Course of Lexical and Discourse Processes through Context, Coreference, and Quantified Expressions

    ERIC Educational Resources Information Center

    Huang, Yi Ting; Gordon, Peter C.

    2011-01-01

    How does prior context influence lexical and discourse-level processing during real-time language comprehension? Experiment 1 examined whether the referential ambiguity introduced by a repeated, anaphoric expression had an immediate or delayed effect on lexical and discourse processing, using an eye-tracking-while-reading task. Eye movements…

  17. Dimensionality of Data Matrices with Applications to Gene Expression Profiles

    ERIC Educational Resources Information Center

    Feng, Xingdong

    2009-01-01

    Probe-level microarray data are usually stored in matrices. Take a given probe set (gene), for example, each row of the matrix corresponds to an array, and each column corresponds to a probe. Often, people summarize each array by the gene expression level. Is one number sufficient to summarize a whole probe set for a specific gene in an array?…

  18. Identifying gene expression modules that define human cell fates.

    PubMed

    Germanguz, I; Listgarten, J; Cinkornpumin, J; Solomon, A; Gaeta, X; Lowry, W E

    2016-05-01

    Using a compendium of cell-state-specific gene expression data, we identified genes that uniquely define cell states, including those thought to represent various developmental stages. Our analysis sheds light on human cell fate through the identification of core genes that are altered over several developmental milestones, and across regional specification. Here we present cell-type specific gene expression data for 17 distinct cell states and demonstrate that these modules of genes can in fact define cell fate. Lastly, we introduce a web-based database to disseminate the results.

  19. Expression of homeobox genes in human erythroleukemia cells.

    PubMed

    Shen, W F; Largman, C; Lowney, P; Hack, F M; Lawrence, H J

    1989-01-01

    Because homeobox-containing genes play a major role in embryogenesis and tissue identity in Drosophila and because similar genes encode tissue-specific transcription factors in mammalian cells, we hypothesized that homeobox genes might plan a role in hematopoietic differentiation and lineage commitment. We therefore surveyed a number of human leukemic cell lines for expression of homeobox-containing genes by Northern gel analysis with probes from the Hox 2 cluster of homeobox genes on chromosome 17. We observed transcripts for Hox 2.1, 2.2, 2.3 and 2.6 in the erythroid line HEL and for Hox 2.3 and 2.6 in the erythroid line K562. Using homeobox-specific probes we confirmed that the transcripts visualized contained the homeodomains for each gene as well as the flanking sequences. The myeloid lines HL60, KG1 and U937 did not express specific transcripts for any of the 4 genes studied. However, all these cell lines demonstrated bands when probed at low stringency with certain Hox 2 probes, indicating the expression of other homologous but as yet unidentified homeobox genes. Expression of Hox 2.3 and 2.6 was seen in some T and B lymphoid cell lines. Induction of differentiation in HEL cells resulted in complex modulation of expression of the Hox 2 genes. We have therefore observed erythroid-restricted expression of certain Hox 2 homeobox containing genes in human erythroid cell lines and modulation of that expression with differentiation, suggesting a role for these genes in the regulation of hematopoiesis. Different homeobox genes appear to be expressed in non-erythroid leukemic cell lines.

  20. Peripheral blood gene expression profiles in COPD subjects.

    PubMed

    Bhattacharya, Soumyaroop; Tyagi, Shivraj; Srisuma, Sorachai; Demeo, Dawn L; Shapiro, Steven D; Bueno, Raphael; Silverman, Edwin K; Reilly, John J; Mariani, Thomas J

    2011-01-01

    To identify non-invasive gene expression markers for chronic obstructive pulmonary disease (COPD), we performed genome-wide expression profiling of peripheral blood samples from 12 subjects with significant airflow obstruction and an equal number of non-obstructed controls. RNA was isolated from Peripheral Blood Mononuclear Cells (PBMCs) and gene expression was assessed using Affymetrix U133 Plus 2.0 arrays.Tests for gene expression changes that discriminate between COPD cases (FEV1< 70% predicted, FEV1/FVC < 0.7) and controls (FEV1> 80% predicted, FEV1/FVC > 0.7) were performed using Significance Analysis of Microarrays (SAM) and Bayesian Analysis of Differential Gene Expression (BADGE). Using either test at high stringency (SAM median FDR = 0 or BADGE p < 0.01) we identified differential expression for 45 known genes. Correlation of gene expression with lung function measurements (FEV1 & FEV1/FVC), using both Pearson and Spearman correlation coefficients (p < 0.05), identified a set of 86 genes. A total of 16 markers showed evidence of significant correlation (p < 0.05) with quantitative traits and differential expression between cases and controls. We further compared our peripheral gene expression markers with those we previously identified from lung tissue of the same cohort. Two genes, RP9and NAPE-PLD, were identified as decreased in COPD cases compared to controls in both lung tissue and blood. These results contribute to our understanding of gene expression changes in the peripheral blood of patients with COPD and may provide insight into potential mechanisms involved in the disease. PMID:21884629

  1. Transcript length mediates developmental timing of gene expression across Drosophila.

    PubMed

    Artieri, Carlo G; Fraser, Hunter B

    2014-11-01

    The time required to transcribe genes with long primary transcripts may limit their ability to be expressed in cells with short mitotic cycles, a phenomenon termed intron delay. As such short cycles are a hallmark of the earliest stages of insect development, we tested the impact of intron delay on the Drosophila developmental transcriptome. We find that long zygotically expressed genes show substantial delay in expression relative to their shorter counterparts, which is not observed for maternally deposited transcripts. Patterns of RNA-seq coverage along transcripts show that this delay is consistent with their inability to completely transcribe long transcripts, but not with transcriptional initiation-based regulatory control. We further show that highly expressed zygotic genes maintain compact transcribed regions across the Drosophila phylogeny, allowing conservation of embryonic expression patterns. We propose that the physical constraints of intron delay affect patterns of expression and the evolution of gene structure of a substantial portion of the Drosophila transcriptome.

  2. Chamber Specific Gene Expression Landscape of the Zebrafish Heart

    PubMed Central

    Singh, Angom Ramcharan; Sivadas, Ambily; Sabharwal, Ankit; Vellarikal, Shamsudheen Karuthedath; Jayarajan, Rijith; Verma, Ankit; Kapoor, Shruti; Joshi, Adita; Scaria, Vinod; Sivasubbu, Sridhar

    2016-01-01

    The organization of structure and function of cardiac chambers in vertebrates is defined by chamber-specific distinct gene expression. This peculiarity and uniqueness of the genetic signatures demonstrates functional resolution attributed to the different chambers of the heart. Altered expression of the cardiac chamber genes can lead to individual chamber related dysfunctions and disease patho-physiologies. Information on transcriptional repertoire of cardiac compartments is important to understand the spectrum of chamber specific anomalies. We have carried out a genome wide transcriptome profiling study of the three cardiac chambers in the zebrafish heart using RNA sequencing. We have captured the gene expression patterns of 13,396 protein coding genes in the three cardiac chambers—atrium, ventricle and bulbus arteriosus. Of these, 7,260 known protein coding genes are highly expressed (≥10 FPKM) in the zebrafish heart. Thus, this study represents nearly an all-inclusive information on the zebrafish cardiac transcriptome. In this study, a total of 96 differentially expressed genes across the three cardiac chambers in zebrafish were identified. The atrium, ventricle and bulbus arteriosus displayed 20, 32 and 44 uniquely expressing genes respectively. We validated the expression of predicted chamber-restricted genes using independent semi-quantitative and qualitative experimental techniques. In addition, we identified 23 putative novel protein coding genes that are specifically restricted to the ventricle and not in the atrium or bulbus arteriosus. In our knowledge, these 23 novel genes have either not been investigated in detail or are sparsely studied. The transcriptome identified in this study includes 68 differentially expressing zebrafish cardiac chamber genes that have a human ortholog. We also carried out spatiotemporal gene expression profiling of the 96 differentially expressed genes throughout the three cardiac chambers in 11 developmental stages and 6

  3. Differential Bacterial Gene Expression During Experimental Pneumococcal Endophthalmitis

    PubMed Central

    Thornton, Justin A.; Tullos, Nathan A.; Sanders, Melissa E.; Ridout, Granger; Wang, Yong-Dong; Taylor, Sidney D.; McDaniel, Larry S.; Marquart, Mary E.

    2015-01-01

    Streptococcus pneumoniae (pneumococcus) is a potential cause of bacterial endophthalmitis in humans that can result in ocular morbidity. We sought to identify pneumococcal genes that are differentially expressed during growth in the vitreous humor of the eye in an experimental endophthalmitis model. Microarray analysis was used to identify genes that were differentially expressed when pneumococci replicated in the vitreous of rabbit eyes as compared with bacteria grown in vitro in Todd Hewitt medium. Array results were verified by quantitative real-time PCR analysis of representative genes. Select genes potentially playing a role in virulence during endophthalmitis were deleted and mutants were tested for reduced eye pathogenesis and altered adhesion to host cells. Array analysis identified 134 genes that were differentially expressed during endophthalmitis. 112 genes demonstrated increased expression during growth in the eye whereas 22 were down-regulated. Real-time analysis verified increased expression of neuraminidase A (SP1693), neuraminidase B (SP1687), and serine protease (SP1954), and decreased expression of RlrA (SP0461) and choline transporter (SP1861). Mutation of neuraminidases A and B had no major effect on pathogenesis. Loss of SP1954 led to increased adherence to host cells. S. pneumoniae enhances and represses expression of a variety of genes during endophthalmitis. While some of these genes reflect changes in metabolic requirements, some appear to play a role in immune evasion and pathogenesis in the eye. PMID:25791614

  4. With Reference to Reference Genes: A Systematic Review of Endogenous Controls in Gene Expression Studies

    PubMed Central

    Chapman, Joanne R.; Waldenström, Jonas

    2015-01-01

    The choice of reference genes that are stably expressed amongst treatment groups is a crucial step in real-time quantitative PCR gene expression studies. Recent guidelines have specified that a minimum of two validated reference genes should be used for normalisation. However, a quantitative review of the literature showed that the average number of reference genes used across all studies was 1.2. Thus, the vast majority of studies continue to use a single gene, with β-actin (ACTB) and/or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) being commonly selected in studies of vertebrate gene expression. Few studies (15%) tested a panel of potential reference genes for stability of expression before using them to normalise data. Amongst studies specifically testing reference gene stability, few found ACTB or GAPDH to be optimal, whereby these genes were significantly less likely to be chosen when larger panels of potential reference genes were screened. Fewer reference genes were tested for stability in non-model organisms, presumably owing to a dearth of available primers in less well characterised species. Furthermore, the experimental conditions under which real-time quantitative PCR analyses were conducted had a large influence on the choice of reference genes, whereby different studies of rat brain tissue showed different reference genes to be the most stable. These results highlight the importance of validating the choice of normalising reference genes before conducting gene expression studies. PMID:26555275

  5. With Reference to Reference Genes: A Systematic Review of Endogenous Controls in Gene Expression Studies.

    PubMed

    Chapman, Joanne R; Waldenström, Jonas

    2015-01-01

    The choice of reference genes that are stably expressed amongst treatment groups is a crucial step in real-time quantitative PCR gene expression studies. Recent guidelines have specified that a minimum of two validated reference genes should be used for normalisation. However, a quantitative review of the literature showed that the average number of reference genes used across all studies was 1.2. Thus, the vast majority of studies continue to use a single gene, with β-actin (ACTB) and/or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) being commonly selected in studies of vertebrate gene expression. Few studies (15%) tested a panel of potential reference genes for stability of expression before using them to normalise data. Amongst studies specifically testing reference gene stability, few found ACTB or GAPDH to be optimal, whereby these genes were significantly less likely to be chosen when larger panels of potential reference genes were screened. Fewer reference genes were tested for stability in non-model organisms, presumably owing to a dearth of available primers in less well characterised species. Furthermore, the experimental conditions under which real-time quantitative PCR analyses were conducted had a large influence on the choice of reference genes, whereby different studies of rat brain tissue showed different reference genes to be the most stable. These results highlight the importance of validating the choice of normalising reference genes before conducting gene expression studies.

  6. Gene Expression Atlas at the European Bioinformatics Institute

    PubMed Central

    Kapushesky, Misha; Emam, Ibrahim; Holloway, Ele; Kurnosov, Pavel; Zorin, Andrey; Malone, James; Rustici, Gabriella; Williams, Eleanor; Parkinson, Helen; Brazma, Alvis

    2010-01-01

    The Gene Expression Atlas (http://www.ebi.ac.uk/gxa) is an added-value database providing information about gene expression in different cell types, organism parts, developmental stages, disease states, sample treatments and other biological/experimental conditions. The content of this database derives from curation, re-annotation and statistical analysis of selected data from the ArrayExpress Archive of Functional Genomics Data. A simple interface allows the user to query for differential gene expression either (i) by gene names or attributes such as Gene Ontology terms, or (ii) by biological conditions, e.g. diseases, organism parts or cell types. The gene queries return the conditions where expression has been reported, while condition queries return which genes are reported to be expressed in these conditions. A combination of both query types is possible. The query results are ranked using various statistical measures and by how many independent studies in the database show the particular gene-condition association. Currently, the database contains information about more than 200 000 genes from nine species and almost 4500 biological conditions studied in over 30 000 assays from over 1000 independent studies. PMID:19906730

  7. Magnetic field-controlled gene expression in encapsulated cells

    PubMed Central

    Ortner, Viktoria; Kaspar, Cornelius; Halter, Christian; Töllner, Lars; Mykhaylyk, Olga; Walzer, Johann; Günzburg, Walter H.; Dangerfield, John A.; Hohenadl, Christine; Czerny, Thomas

    2012-01-01

    Cell and gene therapies have an enormous range of potential applications, but as for most other therapies, dosing is a critical issue, which makes regulated gene expression a prerequisite for advanced strategies. Several inducible expression systems have been established, which mainly rely on small molecules as inducers, such as hormones or antibiotics. The application of these inducers is difficult to control and the effects on gene regulation are slow. Here we describe a novel system for induction of gene expression in encapsulated cells. This involves the modification of cells to express potential therapeutic genes under the control of a heat inducible promoter and the co-encapsulation of these cells with magnetic nanoparticles. These nanoparticles produce heat when subjected to an alternating magnetic field; the elevated temperatures in the capsules then induce gene expression. In the present study we define the parameters of such systems and provide proof-of-principle using reporter gene constructs. The fine-tuned heating of nanoparticles in the magnetic field allows regulation of gene expression from the outside over a broad range and within short time. Such a system has great potential for advancement of cell and gene therapy approaches. PMID:22197778

  8. Protamine stimulates bone sialoprotein gene expression.

    PubMed

    Zhou, Liming; Matsumura, Hiroyoshi; Mezawa, Masaru; Takai, Hideki; Nakayama, Yohei; Mitarai, Makoto; Ogata, Yorimasa

    2013-03-10

    Protamine is a small, arginine-rich, nuclear protein that replaces histone late in the haploid phase of spermatogenesis and is believed to be essential for sperm head condensation and DNA stabilization. Protamine has many biological activities and has roles in hematopoiesis, immune responses, the nervous system and bone metabolism. Bone sialoprotein (BSP) is a mineralized connective tissue-specific protein expressed in differentiated osteoblasts that appears to function in the initial mineralization of bone. Protamine (71.35 ng/ml) increased BSP mRNA levels by 6h in osteoblast-like ROS 17/2.8 cells. In a transient transfection assay, protamine (71.35 ng/ml) increased luciferase activity of the construct (-116 to +60) in ROS 17/2.8 cells and rat bone marrow stromal cells. Luciferase activities induced by protamine were blocked by protein kinase A, tyrosine kinase and ERK1/2 inhibitors. Introduction of 2 bp mutations to the luciferase constructs showed that the effects of protamine were mediated by a cAMP response element (CRE), a fibroblast growth factor 2 response element (FRE) and a homeodomain protein-binding site (HOX). Gel shift analyses showed that protamine (71.35 ng/ml) increased the nuclear protein binding to CRE, FRE and HOX. CREB, phospho-CREB, c-Fos, c-Jun, JunD and Fra2 antibodies disrupted the formation of CRE-protein complexes. Dlx5, Msx2, Runx2 and Smad1 antibodies disrupted FRE- and HOX-protein complex formations. These studies demonstrate that protamine induces BSP transcription by targeting CRE, FRE and HOX sites in the proximal promoter of the rat BSP gene. Moreover, phospho-CREB, c-Fos, c-Jun, JunD, Fra2, Dlx5, Msx2, Runx2 and Smadl transcription factors appear to be key regulators of protamine effects on BSP transcription.

  9. Skin aging, gene expression and calcium.

    PubMed

    Rinnerthaler, Mark; Streubel, Maria Karolin; Bischof, Johannes; Richter, Klaus

    2015-08-01

    The human epidermis provides a very effective barrier function against chemical, physical and microbial insults from the environment. This is only possible as the epidermis renews itself constantly. Stem cells located at the basal lamina which forms the dermoepidermal junction provide an almost inexhaustible source of keratinocytes which differentiate and die during their journey to the surface where they are shed off as scales. Despite the continuous renewal of the epidermis it nevertheless succumbs to aging as the turnover rate of the keratinocytes is slowing down dramatically. Aging is associated with such hallmarks as thinning of the epidermis, elastosis, loss of melanocytes associated with an increased paleness and lucency of the skin and a decreased barrier function. As the differentiation of keratinocytes is strictly calcium dependent, calcium also plays an important role in the aging epidermis. Just recently it was shown that the epidermal calcium gradient in the skin that facilitates the proliferation of keratinocytes in the stratum basale and enables differentiation in the stratum granulosum is lost in the process of skin aging. In the course of this review we try to explain how this calcium gradient is built up on the one hand and is lost during aging on the other hand. How this disturbed calcium homeostasis is affecting the gene expression in aged skin and is leading to dramatic changes in the composition of the cornified envelope will also be discussed. This loss of the epidermal calcium gradient is not only specific for skin aging but can also be found in skin diseases such as Darier disease, Hailey-Hailey disease, psoriasis and atopic dermatitis, which might be very helpful to get a deeper insight in skin aging. PMID:25262846

  10. Targeted expression of suicide gene by tissue-specific promoter and microRNA regulation for cancer gene therapy.

    PubMed

    Danda, Ravikanth; Krishnan, Gopinath; Ganapathy, Kalaivani; Krishnan, Uma Maheswari; Vikas, Khetan; Elchuri, Sailaja; Chatterjee, Nivedita; Krishnakumar, Subramanian

    2013-01-01

    In order to realise the full potential of cancer suicide gene therapy that allows the precise expression of suicide gene in cancer cells, we used a tissue specific Epithelial cell adhesion molecule (EpCAM) promoter (EGP-2) that directs transgene Herpes simplex virus-thymidine kinase (HSV-TK) expression preferentially in EpCAM over expressing cancer cells. EpCAM levels are considerably higher in retinoblastoma (RB), a childhood eye cancer with limited expression in normal cells. Use of miRNA regulation, adjacent to the use of the tissue-specific promoter, would provide the second layer of control to the transgene expression only in the tumor cells while sparing the normal cells. To test this hypothesis we cloned let-7b miRNA targets in the 3'UTR region of HSV-TK suicide gene driven by EpCAM promoter because let-7 family miRNAs, including let-7b, were found to be down regulated in the RB tumors and cell lines. We used EpCAM over expressing and let-7 down regulated RB cell lines Y79, WERI-Rb1 (EpCAM (+ve)/let-7b(down-regulated)), EpCAM down regulated, let-7 over expressing normal retinal Müller glial cell line MIO-M1(EpCAM (-ve)/let-7b(up-regulated)), and EpCAM up regulated, let-7b up-regulated normal thyroid cell line N-Thy-Ori-3.1(EpCAM (+ve)/let-7b(up-regulated)) in the study. The cell proliferation was measured by MTT assay, apoptosis was measured by probing cleaved Caspase3, EpCAM and TK expression were quantified by Western blot. Our results showed that the EGP2-promoter HSV-TK (EGP2-TK) construct with 2 or 4 copies of let-7b miRNA targets expressed TK gene only in Y79, WERI-Rb-1, while the TK gene did not express in MIO-M1. In summary, we have developed a tissue-specific, miRNA-regulated dual control vector, which selectively expresses the suicide gene in EpCAM over expressing cells. PMID:24391761

  11. Adaptive gene expression divergence inferred from population genomics.

    PubMed

    Holloway, Alisha K; Lawniczak, Mara K N; Mezey, Jason G; Begun, David J; Jones, Corbin D

    2007-10-01

    Detailed studies of individual genes have shown that gene expression divergence often results from adaptive evolution of regulatory sequence. Genome-wide analyses, however, have yet to unite patterns of gene expression with polymorphism and divergence to infer population genetic mechanisms underlying expression evolution. Here, we combined genomic expression data--analyzed in a phylogenetic context--with whole genome light-shotgun sequence data from six Drosophila simulans lines and reference sequences from D. melanogaster and D. yakuba. These data allowed us to use molecular population genetics to test for neutral versus adaptive gene expression divergence on a genomic scale. We identified recent and recurrent adaptive evolution along the D. simulans lineage by contrasting sequence polymorphism within D. simulans to divergence from D. melanogaster and D. yakuba. Genes that evolved higher levels of expression in D. simulans have experienced adaptive evolution of the associated 3' flanking and amino acid sequence. Concomitantly, these genes are also decelerating in their rates of protein evolution, which is in agreement with the finding that highly expressed genes evolve slowly. Interestingly, adaptive evolution in 5' cis-regulatory regions did not correspond strongly with expression evolution. Our results provide a genomic view of the intimate link between selection acting on a phenotype and associated genic evolution.

  12. Gene Expression by Mouse Inner Ear Hair Cells during Development

    PubMed Central

    Scheffer, Déborah I.; Shen, Jun

    2015-01-01

    Hair cells of the inner ear are essential for hearing and balance. As a consequence, pathogenic variants in genes specifically expressed in hair cells often cause hereditary deafness. Hair cells are few in number and not easily isolated from the adjacent supporting cells, so the biochemistry and molecular biology of hair cells can be difficult to study. To study gene expression in hair cells, we developed a protocol for hair cell isolation by FACS. With nearly pure hair cells and surrounding cells, from cochlea and utricle and from E16 to P7, we performed a comprehensive cell type-specific RNA-Seq study of gene expression during mouse inner ear development. Expression profiling revealed new hair cell genes with distinct expression patterns: some are specific for vestibular hair cells, others for cochlear hair cells, and some are expressed just before or after maturation of mechanosensitivity. We found that many of the known hereditary deafness genes are much more highly expressed in hair cells than surrounding cells, suggesting that genes preferentially expressed in hair cells are good candidates for unknown deafness genes. PMID:25904789

  13. Reference genes for the normalization of gene expression in eucalyptus species.

    PubMed

    de Oliveira, Luisa Abruzzi; Breton, Michèle Claire; Bastolla, Fernanda Macedo; Camargo, Sandro da Silva; Margis, Rogério; Frazzon, Jeverson; Pasquali, Giancarlo

    2012-02-01

    Gene expression analysis is increasingly important in biological research, with reverse transcription-quantitative PCR (RT-qPCR) becoming the method of choice for high-throughput and accurate expression profiling of selected genes. Considering the increased sensitivity, reproducibility and large dynamic range of this method, the requirements for proper internal reference gene(s) for relative expression normalization have become much more stringent. Given the increasing interest in the functional genomics of Eucalyptus, we sought to identify and experimentally verify suitable reference genes for the normalization of gene expression associated with the flower, leaf and xylem of six species of the genus. We selected 50 genes that exhibited the least variation in microarrays of E. grandis leaves and xylem, and E. globulus xylem. We further performed the experimental analysis using RT-qPCR for six Eucalyptus species and three different organs/tissues. Employing algorithms geNorm and NormFinder, we assessed the gene expression stability of eight candidate new reference genes. Classic housekeeping genes were also included in the analysis. The stability profiles of candidate genes were in very good agreement. PCR results proved that the expression of novel Eucons04, Eucons08 and Eucons21 genes was the most stable in all Eucalyptus organs/tissues and species studied. We showed that the combination of these genes as references when measuring the expression of a test gene results in more reliable patterns of expression than traditional housekeeping genes. Hence, novel Eucons04, Eucons08 and Eucons21 genes are the best suitable references for the normalization of expression studies in the Eucalyptus genus. PMID:22197885

  14. Identification and validation of reference genes for gene expression studies in water buffalo.

    PubMed

    Terzi, V; Morcia, C; Spini, M; Tudisco, R; Cutrignelli, M I; Infascelli, F; Stanca, A M; Faccioli, P

    2010-06-01

    In gene expression analysis, a key step to obtain informative data from reverse transcription quantitative PCR (RT qPCR) assay is normalization, that is usually achieved by ratio to correct the abundance of the gene of interest against that of an endogenous reference gene. The finding of such reference genes, ideally expressed in a stable way in multiple tissue samples and in different experimental conditions, is a non-trivial problem. In this work, a set of genes potentially useful as reference for gene expression studies in water buffalo has been identified and evaluated. In the first step, a publicly available Bos taurus expressed sequence tags database has been downloaded from the TIGR Gene Index and mined by some simple frequency algorithms to find out which tentative consensuses are present in a remarkable number of different cDNA libraries and, consequently, are more suitable to be included in a starter set of candidate reference genes. To validate the potential of such candidates for their use as normalizers in buffalo gene expression analysis, an RT qPCR analysis has been carried out, in which the expression stability of these genes has been evaluated on a panel of buffalo tissues and organs. Our results indicate that ribosomal proteins L4 and L5 and Gek protein encoding genes can be useful as normalizers to compare gene expression levels across tissues and organs in buffalo.

  15. Gene Expression Profile Analysis of Type 2 Diabetic Mouse Liver

    PubMed Central

    Zhang, Fang; Xu, Xiang; Zhang, Yi; Zhou, Ben; He, Zhishui; Zhai, Qiwei

    2013-01-01

    Liver plays a key role in glucose metabolism and homeostasis, and impaired hepatic glucose metabolism contributes to the development of type 2 diabetes. However, the precise gene expression profile of diabetic liver and its association with diabetes and related diseases are yet to be further elucidated. In this study, we detected the gene expression profile by high-throughput sequencing in 9-week-old normal and type 2 diabetic db/db mouse liver. Totally 12132 genes were detected, and 2627 genes were significantly changed in diabetic mouse liver. Biological process analysis showed that the upregulated genes in diabetic mouse liver were mainly enriched in metabolic processes. Surprisingly, the downregulated genes in diabetic mouse liver were mainly enriched in immune-related processes, although all the altered genes were still mainly enriched in metabolic processes. Similarly, KEGG pathway analysis showed that metabolic pathways were the major pathways altered in diabetic mouse liver, and downregulated genes were enriched in immune and cancer pathways. Analysis of the key enzyme genes in fatty acid and glucose metabolism showed that some key enzyme genes were significantly increased and none of the detected key enzyme genes were decreased. In addition, FunDo analysis showed that liver cancer and hepatitis were most likely to be associated with diabetes. Taken together, this study provides the digital gene expression profile of diabetic mouse liver, and demonstrates the main diabetes-associated hepatic biological processes, pathways, key enzyme genes in fatty acid and glucose metabolism and potential hepatic diseases. PMID:23469233

  16. Eucalypt MADS-Box Genes Expressed in Developing Flowers

    PubMed Central

    Southerton, Simon G.; Marshall, Heidi; Mouradov, Aidyn; Teasdale, Robert D.

    1998-01-01

    Three MADS-box genes were identified from a cDNA library derived from young flowers of Eucalyptus grandis W. Hill ex Maiden. The three egm genes are single-copy genes and are expressed almost exclusively in flowers. The egm1 and egm3 genes shared strongest homology with other plant MADS-box genes, which mediate between the floral meristem and the organ-identity genes. The egm3 gene was also expressed strongly in the receptacle or floral tube, which surrounds the carpels in the eucalypt flower and bears the sepals, petals, and numerous stamens. There appeared to be a group of genes in eucalypts with strong homology with the 3′ region of the egm1 gene. The egm2 gene was expressed in eucalypt petals and stamens and was most homologous to MADS-box genes, which belong to the globosa group of genes, which regulate organogenesis of the second and third floral whorls. The possible role of these three genes in eucalypt floral development is discussed. PMID:9765522

  17. Expression of HOX C homeobox genes in lymphoid cells.

    PubMed

    Lawrence, H J; Stage, K M; Mathews, C H; Detmer, K; Scibienski, R; MacKenzie, M; Migliaccio, E; Boncinelli, E; Largman, C

    1993-08-01

    The class I homeobox genes located in four clusters in mammalian genomes (HOX A, HOX B, HOX C, and HOX D) appear to play a major role in fetal development. Previous surveys of homeobox gene expression in human leukemic cell lines have shown that certain HOX A genes are expressed only in myeloid cell lines, whereas HOX B gene expression is largely restricted to cells with erythroid potential. We now report a survey of the expression patterns of 9 homeobox genes from the HOX C locus in a panel of 24 human and 7 murine leukemic cell lines. The most striking observation is the lymphoid-specific pattern of expression of HOX C4, located at the 3' end of the locus. A major transcript of 1.9 kilobases is observed in both T-cell and B-cell lines. HOX C4 expression is also detected in normal human marrow and peripheral blood lymphocytes, but not in mature granulocytes or monocytes. HOX C8 is also expressed in human lymphoid cells but is expressed in other blood cell types as well. However, the HOX C8 transcript pattern is lineage specific. These data, in conjunction with earlier findings, suggest that homeobox gene expression influences lineage determination during hematopoiesis.

  18. Exact protein distributions for stochastic models of gene expression using partitioning of Poisson processes

    NASA Astrophysics Data System (ADS)

    Pendar, Hodjat; Platini, Thierry; Kulkarni, Rahul V.

    2013-04-01

    Stochasticity in gene expression gives rise to fluctuations in protein levels across a population of genetically identical cells. Such fluctuations can lead to phenotypic variation in clonal populations; hence, there is considerable interest in quantifying noise in gene expression using stochastic models. However, obtaining exact analytical results for protein distributions has been an intractable task for all but the simplest models. Here, we invoke the partitioning property of Poisson processes to develop a mapping that significantly simplifies the analysis of stochastic models of gene expression. The mapping leads to exact protein distributions using results for mRNA distributions in models with promoter-based regulation. Using this approach, we derive exact analytical results for steady-state and time-dependent distributions for the basic two-stage model of gene expression. Furthermore, we show how the mapping leads to exact protein distributions for extensions of the basic model that include the effects of posttranscriptional and posttranslational regulation. The approach developed in this work is widely applicable and can contribute to a quantitative understanding of stochasticity in gene expression and its regulation.

  19. Exact protein distributions for stochastic models of gene expression using partitioning of Poisson processes.

    PubMed

    Pendar, Hodjat; Platini, Thierry; Kulkarni, Rahul V

    2013-04-01

    Stochasticity in gene expression gives rise to fluctuations in protein levels across a population of genetically identical cells. Such fluctuations can lead to phenotypic variation in clonal populations; hence, there is considerable interest in quantifying noise in gene expression using stochastic models. However, obtaining exact analytical results for protein distributions has been an intractable task for all but the simplest models. Here, we invoke the partitioning property of Poisson processes to develop a mapping that significantly simplifies the analysis of stochastic models of gene expression. The mapping leads to exact protein distributions using results for mRNA distributions in models with promoter-based regulation. Using this approach, we derive exact analytical results for steady-state and time-dependent distributions for the basic two-stage model of gene expression. Furthermore, we show how the mapping leads to exact protein distributions for extensions of the basic model that include the effects of posttranscriptional and posttranslational regulation. The approach developed in this work is widely applicable and can contribute to a quantitative understanding of stochasticity in gene expression and its regulation.

  20. Organization, structure and expression of murine interferon alpha genes.

    PubMed

    Zwarthoff, E C; Mooren, A T; Trapman, J

    1985-02-11

    Using a human interferon-alpha probe we have isolated recombinant phages containing murine interferon-alpha (Mu IFN-alpha) genes from a genomic library. One of these phages contained two complete Mu IFN-alpha genes and part of a third gene. The insert of a second phage held two IFN genes. This indicates that the Mu IFN-alpha genes are clustered in the genome as is the case for the analogous human genes. The nucleotide sequences of these 5 genes were determined. They show that the genes are all different, albeit highly homologous. The deduced amino acid sequences show that four of the five genes contain a putative glycosylation site. Three genes were transiently expressed in COS cells and they gave rise to protein products showing antiviral properties. The expression of the five Mu IFN-alpha genes and the Mu IFN-beta gene was studied in virus-induced mouse L cells. The individual mRNAs were visualized in a nuclease S1 experiment, using a specific probe for each gene. In RNA preparations from induced cells mRNAs for each of the five alpha genes and the beta gene were present. However, substantial differences in the amounts of the individual mRNAs were observed.

  1. A tool kit for quantifying eukaryotic rRNA gene sequences from human microbiome samples.

    PubMed

    Dollive, Serena; Peterfreund, Gregory L; Sherrill-Mix, Scott; Bittinger, Kyle; Sinha, Rohini; Hoffmann, Christian; Nabel, Christopher S; Hill, David A; Artis, David; Bachman, Michael A; Custers-Allen, Rebecca; Grunberg, Stephanie; Wu, Gary D; Lewis, James D; Bushman, Frederic D

    2012-07-03

    Eukaryotic microorganisms are important but understudied components of the human microbiome. Here we present a pipeline for analysis of deep sequencing data on single cell eukaryotes. We designed a new 18S rRNA gene-specific PCR primer set and compared a published rRNA gene internal transcribed spacer (ITS) gene primer set. Amplicons were tested against 24 specimens from defined eukaryotes and eight well-characterized human stool samples. A software pipeline https://sourceforge.net/projects/brocc/ was developed for taxonomic attribution, validated against simulated data, and tested on pyrosequence data. This study provides a well-characterized tool kit for sequence-based enumeration of eukaryotic organisms in human microbiome samples.

  2. Evaluation of Quantitative PCR Reference Genes for Gene Expression Studies in Tribolium castaneum After Fungal Challenge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To investigate gene expression in Tribolium castaneum exposed to Beauveria bassiana, reference genes for qPCR were evaluated. Of these, the widely used genes for ß-actin, a-tubulin, and RPS6 were not stable. The most stable were ribosomal protein genes, RPS3, RPS18, and RPL13a. Syntaxin1, syntaxin6...

  3. Expression of heat shock protein genes in insect stress responses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The heat shock proteins (HSPs) that are abundantly expressed in insects are important modulators of insect survival. Expression of HSP genes in insects is not only developmentally regulated, but also induced by various stressors in order to confer protection against such stressors. The expression o...

  4. IDENTIFICATION OF BIOLOGICALLY RELEVANT GENES USING A DATABASE OF RAT LIVER AND KIDNEY BASELINE GENE EXPRESSION

    EPA Science Inventory

    Microarray data from independent labs and studies can be compared to potentially identify toxicologically and biologically relevant genes. The Baseline Animal Database working group of HESI was formed to assess baseline gene expression from microarray data derived from control or...

  5. Gene Body Methylation can alter Gene Expression and is a Therapeutic Target in Cancer

    PubMed Central

    Yang, Xiaojing; Han, Han; De Carvalho, Daniel D.; Lay, Fides D.; Jones, Peter A.; Liang, Gangning

    2014-01-01

    SUMMARY DNA methylation in promoters is well known to silence genes and is the presumed therapeutic target of methylation inhibitors. Gene body methylation is positively correlated with expression yet its function is unknown. We show that 5-aza-2'-deoxycytidine treatment not only reactivates genes but decreases the over-expression of genes, many of which are involved in metabolic processes regulated by c-MYC. Down-regulation is caused by DNA demethylation of the gene bodies and restoration of high levels of expression requires remethylation by DNMT3B. Gene body methylation may therefore be an unexpected therapeutic target for DNA methylation inhibitors, resulting in the normalization of gene over-expression induced during carcinogenesis. Our results provide direct evidence for a causal relationship between gene body methylation and transcription. PMID:25263941

  6. Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression.

    EPA Science Inventory

    Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression Exposure to many drugs and environmentally-relevant chemicals can cause adverse outcomes. These adverse outcomes, such as cancer, have been linked to mol...

  7. Noise in gene expression is coupled to growth rate.

    PubMed

    Keren, Leeat; van Dijk, David; Weingarten-Gabbay, Shira; Davidi, Dan; Jona, Ghil; Weinberger, Adina; Milo, Ron; Segal, Eran

    2015-12-01

    Genetically identical cells exposed to the same environment display variability in gene expression (noise), with important consequences for the fidelity of cellular regulation and biological function. Although population average gene expression is tightly coupled to growth rate, the effects of changes in environmental conditions on expression variability are not known. Here, we measure the single-cell expression distributions of approximately 900 Saccharomyces cerevisiae promoters across four environmental conditions using flow cytometry, and find that gene expression noise is tightly coupled to the environment and is generally higher at lower growth rates. Nutrient-poor conditions, which support lower growth rates, display elevated levels of noise for most promoters, regardless of their specific expression values. We present a simple model of noise in expression that results from having an asynchronous population, with cells at different cell-cycle stages, and with different partitioning of the cells between the stages at different growth rates. This model predicts non-monotonic global changes in noise at different growth rates as well as overall higher variability in expression for cell-cycle-regulated genes in all conditions. The consistency between this model and our data, as well as with noise measurements of cells growing in a chemostat at well-defined growth rates, suggests that cell-cycle heterogeneity is a major contributor to gene expression noise. Finally, we identify gene and promoter features that play a role in gene expression noise across conditions. Our results show the existence of growth-related global changes in gene expression noise and suggest their potential phenotypic implications. PMID:26355006

  8. Noise in gene expression is coupled to growth rate

    PubMed Central

    Keren, Leeat; van Dijk, David; Weingarten-Gabbay, Shira; Davidi, Dan; Jona, Ghil; Weinberger, Adina; Milo, Ron; Segal, Eran

    2015-01-01

    Genetically identical cells exposed to the same environment display variability in gene expression (noise), with important consequences for the fidelity of cellular regulation and biological function. Although population average gene expression is tightly coupled to growth rate, the effects of changes in environmental conditions on expression variability are not known. Here, we measure the single-cell expression distributions of approximately 900 Saccharomyces cerevisiae promoters across four environmental conditions using flow cytometry, and find that gene expression noise is tightly coupled to the environment and is generally higher at lower growth rates. Nutrient-poor conditions, which support lower growth rates, display elevated levels of noise for most promoters, regardless of their specific expression values. We present a simple model of noise in expression that results from having an asynchronous population, with cells at different cell-cycle stages, and with different partitioning of the cells between the stages at different growth rates. This model predicts non-monotonic global changes in noise at different growth rates as well as overall higher variability in expression for cell-cycle–regulated genes in all conditions. The consistency between this model and our data, as well as with noise measurements of cells growing in a chemostat at well-defined growth rates, suggests that cell-cycle heterogeneity is a major contributor to gene expression noise. Finally, we identify gene and promoter features that play a role in gene expression noise across conditions. Our results show the existence of growth-related global changes in gene expression noise and suggest their potential phenotypic implications. PMID:26355006

  9. Chromatin mechanisms in the developmental control of imprinted gene expression.

    PubMed

    Sanli, Ildem; Feil, Robert

    2015-10-01

    Hundreds of protein-coding genes and regulatory non-coding RNAs (ncRNAs) are subject to genomic imprinting. The mono-allelic DNA methylation marks that control imprinted gene expression are somatically maintained throughout development, and this process is linked to specific chromatin features. Yet, at many imprinted genes, the mono-allelic expression is lineage or tissue-specific. Recent studies provide mechanistic insights into the developmentally-restricted action of the 'imprinting control regions' (ICRs). At several imprinted domains, the ICR expresses a long ncRNA that mediates chromatin repression in cis (and probably in trans as well). ICRs at other imprinted domains mediate higher-order chromatin structuration that enhances, or prevents, transcription of close-by genes. Here, we present how chromatin and ncRNAs contribute to developmental control of imprinted gene expression and discuss implications for disease. This article is part of a Directed Issue entitled: Epigenetics dynamics in development and disease.

  10. Microarray Analysis of Pneumococcal Gene Expression during Invasive Disease

    PubMed Central

    Orihuela, Carlos J.; Radin, Jana N.; Sublett, Jack E.; Gao, Geli; Kaushal, Deepak; Tuomanen, Elaine I.

    2004-01-01

    Streptococcus pneumoniae is a leading cause of invasive bacterial disease. This is the first study to examine the expression of S. pneumoniae genes in vivo by using whole-genome microarrays available from The Institute for Genomic Research. Total RNA was collected from pneumococci isolated from infected blood, infected cerebrospinal fluid, and bacteria attached to a pharyngeal epithelial cell line in vitro. Microarray analysis of pneumococcal genes expressed in these models identified body site-specific patterns of expression for virulence factors, transporters, transcription factors, translation-associated proteins, metabolism, and genes with unknown function. Contributions to virulence predicted for several unknown genes with enhanced expression in vivo were confirmed by insertion duplication mutagenesis and challenge of mice with the mutants. Finally, we cross-referenced our results with previous studies that used signature-tagged mutagenesis and differential fluorescence induction to identify genes that are potentially required by a broad range of pneumococcal strains for invasive disease. PMID:15385455

  11. Computational gene expression profiling under salt stress reveals patterns of co-expression

    PubMed Central

    Sanchita; Sharma, Ashok

    2016-01-01

    Plants respond differently to environmental conditions. Among various abiotic stresses, salt stress is a condition where excess salt in soil causes inhibition of plant growth. To understand the response of plants to the stress conditions, identification of the responsible genes is required. Clustering is a data mining technique used to group the genes with similar expression. The genes of a cluster show similar expression and function. We applied clustering algorithms on gene expression data of Solanum tuberosum showing differential expression in Capsicum annuum under salt stress. The clusters, which were common in multiple algorithms were taken further for analysis. Principal component analysis (PCA) further validated the findings of other cluster algorithms by visualizing their clusters in three-dimensional space. Functional annotation results revealed that most of the genes were involved in stress related responses. Our findings suggest that these algorithms may be helpful in the prediction of the function of co-expressed genes. PMID:26981411

  12. Gene expression profiles of Nitrosomonas europaea, an obligate chemolitotroph

    SciTech Connect

    Daniel J. Arp

    2005-05-25

    Nitrosomonas europaea is an aerobic lithoautotrophic bacterium that uses ammonia (NH3) as its energy source. As a nitrifier, it is an important participant in the nitrogen cycle, which can also influence the carbon cycle. The focus of this work was to explore the genetic structure and mechanisms underlying the lithoautotrophic growth style of N. europaea. Whole genome gene expression: The gene expression profile of cells in exponential growth and during starvation was analyzed using microarrays. During growth, 98% of the genes increased in expression at least two fold compared to starvation conditions. In growing cells, approximately 30% of the genes were expressed eight fold higher, Approximately 10% were expressed more than 15 fold higher. Approximately 3% (91 genes) were expressed to more than 20 fold of their levels in starved cells. Carbon fixation gene expression: N. europaea fixes carbon via the Calvin-Benson-Bassham (CBB) cycle via a type I ribulose bisphosphate carboxylase/oxygenase (RubisCO). This study showed that transcription of cbb genes was up-regulated when the carbon source was limited, while amo, hao and other energy harvesting related genes were down-regulated. Iron related gene expression: Because N. europaea has a relatively high content of hemes, sufficient Fe must be available in the medium for it to grow. The genome revealed that approximately 5% of the coding genes in N. europaea are dedicated to Fe transport and assimilation. Nonetheless, with the exception of citrate biosynthesis genes, N. europaea lacks genes for siderophore production. The Fe requirements for growth and the expression of the putative membrane siderophore receptors were determined. The N. europaea genome has over 100 putative genes ({approx}5% of the coding genes) related to Fe uptake and its siderophore receptors could be grouped phylogenetically in four clusters. Fe related genes, such as a number of TonB-dependent Fe-siderophore receptors for ferrichrome and

  13. Gene expression profiles of Nitrosomonas europaea, an obligate chemolitotroph

    SciTech Connect

    Daniel J Arp

    2005-06-15

    Nitrosomonas europaea is an aerobic lithoautotrophic bacterium that uses ammonia (NH3) as its energy source. As a nitrifier, it is an important participant in the nitrogen cycle, which can also influence the carbon cycle. The focus of this work was to explore the genetic structure and mechanisms underlying the lithoautotrophic growth style of N. europaea. Whole genome gene expression. The gene expression profile of cells in exponential growth and during starvation was analyzed using microarrays. During growth, 98% of the genes increased in expression at least two fold compared to starvation conditions. In growing cells, approximately 30% of the genes were expressed eight fold higher, Approximately 10% were expressed more than 15 fold higher. Approximately 3% (91 genes) were expressed to more than 20 fold of their levels in starved cells. Carbon fixation gene expression. N. europaea fixes carbon via the Calvin-Benson-Bassham (CBB) cycle via a type I ribulose bisphosphate carboxylase/oxygenase (RubisCO). This study showed that transcription of cbb genes was up-regulated when the carbon source was limited, while amo, hao and other energy harvesting related genes were down-regulated. Iron related gene expression. Because N. europaea has a relatively high content of hemes, sufficient Fe must be available in the medium for it to grow. The genome revealed that approximately 5% of the coding genes in N. europaea are dedicated to Fe transport and assimilation. Nonetheless, with the exception of citrate biosynthesis genes, N. europaea lacks genes for siderophore production. The Fe requirements for growth and the expression of the putative membrane siderophore receptors were determined. The N. europaea genome has over 100 putative genes ({approx}5% of the coding genes) related to Fe uptake and its siderophore receptors could be grouped phylogenetically in four clusters. Fe related genes, such as a number of TonB-dependent Fe-siderophore receptors for ferrichrome and

  14. Rapid evolution of male-biased gene expression in Drosophila.

    PubMed

    Meiklejohn, Colin D; Parsch, John; Ranz, José M; Hartl, Daniel L

    2003-08-19

    A number of genes associated with sexual traits and reproduction evolve at the sequence level faster than the majority of genes coding for non-sex-related traits. Whole genome analyses allow this observation to be extended beyond the limited set of genes that have been studied thus far. We use cDNA microarrays to demonstrate that this pattern holds in Drosophila for the phenotype of gene expression as well, but in one sex only. Genes that are male-biased in their expression show more variation in relative expression levels between conspecific populations and two closely related species than do female-biased genes or genes with sexually monomorphic expression patterns. Additionally, elevated ratios of interspecific expression divergence to intraspecific expression variation among male-biased genes suggest that differences in rates of evolution may be due in part to natural selection. This finding has implications for our understanding of the importance of sexual dimorphism for speciation and rates of phenotypic evolution.

  15. Prediction of gene expression in embryonic structures of Drosophila melanogaster.

    PubMed

    Samsonova, Anastasia A; Niranjan, Mahesan; Russell, Steven; Brazma, Alvis

    2007-07-01

    Understanding how sets of genes are coordinately regulated in space and time to generate the diversity of cell types that characterise complex metazoans is a major challenge in modern biology. The use of high-throughput approaches, such as large-scale in situ hybridisation and genome-wide expression profiling via DNA microarrays, is beginning to provide insights into the complexities of development. However, in many organisms the collection and annotation of comprehensive in situ localisation data is a difficult and time-consuming task. Here, we present a widely applicable computational approach, integrating developmental time-course microarray data with annotated in situ hybridisation studies, that facilitates the de novo prediction of tissue-specific expression for genes that have no in vivo gene expression localisation data available. Using a classification approach, trained with data from microarray and in situ hybridisation studies of gene expression during Drosophila embryonic development, we made a set of predictions on the tissue-specific expression of Drosophila genes that have not been systematically characterised by in situ hybridisation experiments. The reliability of our predictions is confirmed by literature-derived annotations in FlyBase, by overrepresentation of Gene Ontology biological process annotations, and, in a selected set, by detailed gene-specific studies from the literature. Our novel organism-independent method will be of considerable utility in enriching the annotation of gene function and expression in complex multicellular organisms.

  16. Identification of reference genes in human myelomonocytic cells for gene expression studies in altered gravity.

    PubMed

    Thiel, Cora S; Hauschild, Swantje; Tauber, Svantje; Paulsen, Katrin; Raig, Christiane; Raem, Arnold; Biskup, Josefine; Gutewort, Annett; Hürlimann, Eva; Unverdorben, Felix; Buttron, Isabell; Lauber, Beatrice; Philpot, Claudia; Lier, Hartwin; Engelmann, Frank; Layer, Liliana E; Ullrich, Oliver

    2015-01-01

    Gene expression studies are indispensable for investigation and elucidation of molecular mechanisms. For the process of normalization, reference genes ("housekeeping genes") are essential to verify gene expression analysis. Thus, it is assumed that these reference genes demonstrate similar expression levels over all experimental conditions. However, common recommendations about reference genes were established during 1 g conditions and therefore their applicability in studies with altered gravity has not been demonstrated yet. The microarray technology is frequently used to generate expression profiles under defined conditions and to determine the relative difference in expression levels between two or more different states. In our study, we searched for potential reference genes with stable expression during different gravitational conditions (microgravity, normogravity, and hypergravity) which are additionally not altered in different hardware systems. We were able to identify eight genes (ALB, B4GALT6, GAPDH, HMBS, YWHAZ, ABCA5, ABCA9, and ABCC1) which demonstrated no altered gene expression levels in all tested conditions and therefore represent good candidates for the standardization of gene expression studies in altered gravity. PMID:25654098

  17. Identification of reference genes in human myelomonocytic cells for gene expression studies in altered gravity.

    PubMed

    Thiel, Cora S; Hauschild, Swantje; Tauber, Svantje; Paulsen, Katrin; Raig, Christiane; Raem, Arnold; Biskup, Josefine; Gutewort, Annett; Hürlimann, Eva; Unverdorben, Felix; Buttron, Isabell; Lauber, Beatrice; Philpot, Claudia; Lier, Hartwin; Engelmann, Frank; Layer, Liliana E; Ullrich, Oliver

    2015-01-01

    Gene expression studies are indispensable for investigation and elucidation of molecular mechanisms. For the process of normalization, reference genes ("housekeeping genes") are essential to verify gene expression analysis. Thus, it is assumed that these reference genes demonstrate similar expression levels over all experimental conditions. However, common recommendations about reference genes were established during 1 g conditions and therefore their applicability in studies with altered gravity has not been demonstrated yet. The microarray technology is frequently used to generate expression profiles under defined conditions and to determine the relative difference in expression levels between two or more different states. In our study, we searched for potential reference genes with stable expression during different gravitational conditions (microgravity, normogravity, and hypergravity) which are additionally not altered in different hardware systems. We were able to identify eight genes (ALB, B4GALT6, GAPDH, HMBS, YWHAZ, ABCA5, ABCA9, and ABCC1) which demonstrated no altered gene expression levels in all tested conditions and therefore represent good candidates for the standardization of gene expression studies in altered gravity.

  18. Adult mouse brain gene expression patterns bear an embryologic imprint.

    PubMed

    Zapala, Matthew A; Hovatta, Iiris; Ellison, Julie A; Wodicka, Lisa; Del Rio, Jo A; Tennant, Richard; Tynan, Wendy; Broide, Ron S; Helton, Rob; Stoveken, Barbara S; Winrow, Christopher; Lockhart, Daniel J; Reilly, John F; Young, Warren G; Bloom, Floyd E; Lockhart, David J; Barlow, Carrolee

    2005-07-19

    The current model to explain the organization of the mammalian nervous system is based on studies of anatomy, embryology, and evolution. To further investigate the molecular organization of the adult mammalian brain, we have built a gene expression-based brain map. We measured gene expression patterns for 24 neural tissues covering the mouse central nervous system and found, surprisingly, that the adult brain bears a transcriptional "imprint" consistent with both embryological origins and classic evolutionary relationships. Embryonic cellular position along the anterior-posterior axis of the neural tube was shown to be closely associated with, and possibly a determinant of, the gene expression patterns in adult structures. We also observed a significant number of embryonic patterning and homeobox genes with region-specific expression in the adult nervous system. The relationships between global expression patterns for different anatomical regions and the nature of the observed region-specific genes suggest that the adult brain retains a degree of overall gene expression established during embryogenesis that is important for regional specificity and the functional relationships between regions in the adult. The complete collection of extensively annotated gene expression data along with data mining and visualization tools have been made available on a publicly accessible web site (www.barlow-lockhart-brainmapnimhgrant.org).

  19. Genome-wide quantification of homeolog expression ratio revealed nonstochastic gene regulation in synthetic allopolyploid Arabidopsis.

    PubMed

    Akama, Satoru; Shimizu-Inatsugi, Rie; Shimizu, Kentaro K; Sese, Jun

    2014-04-01

    Genome duplication with hybridization, or allopolyploidization, occurs commonly in plants, and is considered to be a strong force for generating new species. However, genome-wide quantification of homeolog expression ratios was technically hindered because of the high homology between homeologous gene pairs. To quantify the homeolog expression ratio using RNA-seq obtained from polyploids, a new method named HomeoRoq was developed, in which the genomic origin of sequencing reads was estimated using mismatches between the read and each parental genome. To verify this method, we first assembled the two diploid parental genomes of Arabidopsis halleri subsp. gemmifera and Arabidopsis lyrata subsp. petraea (Arabidopsis petraea subsp. umbrosa), then generated a synthetic allotetraploid, mimicking the natural allopolyploid Arabidopsis kamchatica. The quantified ratios corresponded well to those obtained by Pyrosequencing. We found that the ratios of homeologs before and after cold stress treatment were highly correlated (r = 0.870). This highlights the presence of nonstochastic polyploid gene regulation despite previous research identifying stochastic variation in expression. Moreover, our new statistical test incorporating overdispersion identified 226 homeologs (1.11% of 20 369 expressed homeologs) with significant ratio changes, many of which were related to stress responses. HomeoRoq would contribute to the study of the genes responsible for polyploid-specific environmental responses. PMID:24423873

  20. Analysis of bHLH coding genes using gene co-expression network approach.

    PubMed

    Srivastava, Swati; Sanchita; Singh, Garima; Singh, Noopur; Srivastava, Gaurava; Sharma, Ashok

    2016-07-01

    Network analysis provides a powerful framework for the interpretation of data. It uses novel reference network-based metrices for module evolution. These could be used to identify module of highly connected genes showing variation in co-expression network. In this study, a co-expression network-based approach was used for analyzing the genes from microarray data. Our approach consists of a simple but robust rank-based network construction. The publicly available gene expression data of Solanum tuberosum under cold and heat stresses were considered to create and analyze a gene co-expression network. The analysis provide highly co-expressed module of bHLH coding genes based on correlation values. Our approach was to analyze the variation of genes expression, according to the time period of stress through co-expression network approach. As the result, the seed genes were identified showing multiple connections with other genes in the same cluster. Seed genes were found to be vary in different time periods of stress. These analyzed seed genes may be utilized further as marker genes for developing the stress tolerant plant species.

  1. The choice of reference genes for assessing gene expression in sugarcane under salinity and drought stresses

    PubMed Central

    Guo, Jinlong; Ling, Hui; Wu, Qibin; Xu, Liping; Que, Youxiong

    2014-01-01

    Sugarcane (Saccharum spp. hybrids) is a world-wide cash crop for sugar and biofuel in tropical and subtropical regions and suffers serious losses in cane yield and sugar content under salinity and drought stresses. Although real-time quantitative PCR has a numerous advantage in the expression quantification of stress-related genes for the elaboration of the corresponding molecular mechanism in sugarcane, the variation happened across the process of gene expression quantification should be normalized and monitored by introducing one or several reference genes. To validate suitable reference genes or gene sets for sugarcane gene expression normalization, 13 candidate reference genes have been tested across 12 NaCl- and PEG-treated sugarcane samples for four sugarcane genotypes using four commonly used systematic statistical algorithms termed geNorm, BestKeeper, NormFinder and the deltaCt method. The results demonstrated that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and eukaryotic elongation factor 1-alpha (eEF-1a) were identified as suitable reference genes for gene expression normalization under salinity/drought-treatment in sugarcane. Moreover, the expression analyses of SuSK and 6PGDH further validated that a combination of clathrin adaptor complex (CAC) and cullin (CUL) as reference should be better for gene expression normalization. These results can facilitate the future research on gene expression in sugarcane under salinity and drought stresses. PMID:25391499

  2. The choice of reference genes for assessing gene expression in sugarcane under salinity and drought stresses.

    PubMed

    Guo, Jinlong; Ling, Hui; Wu, Qibin; Xu, Liping; Que, Youxiong

    2014-01-01

    Sugarcane (Saccharum spp. hybrids) is a world-wide cash crop for sugar and biofuel in tropical and subtropical regions and suffers serious losses in cane yield and sugar content under salinity and drought stresses. Although real-time quantitative PCR has a numerous advantage in the expression quantification of stress-related genes for the elaboration of the corresponding molecular mechanism in sugarcane, the variation happened across the process of gene expression quantification should be normalized and monitored by introducing one or several reference genes. To validate suitable reference genes or gene sets for sugarcane gene expression normalization, 13 candidate reference genes have been tested across 12 NaCl- and PEG-treated sugarcane samples for four sugarcane genotypes using four commonly used systematic statistical algorithms termed geNorm, BestKeeper, NormFinder and the deltaCt method. The results demonstrated that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and eukaryotic elongation factor 1-alpha (eEF-1a) were identified as suitable reference genes for gene expression normalization under salinity/drought-treatment in sugarcane. Moreover, the expression analyses of SuSK and 6PGDH further validated that a combination of clathrin adaptor complex (CAC) and cullin (CUL) as reference should be better for gene expression normalization. These results can facilitate the future research on gene expression in sugarcane under salinity and drought stresses. PMID:25391499

  3. The choice of reference genes for assessing gene expression in sugarcane under salinity and drought stresses.

    PubMed

    Guo, Jinlong; Ling, Hui; Wu, Qibin; Xu, Liping; Que, Youxiong

    2014-11-13

    Sugarcane (Saccharum spp. hybrids) is a world-wide cash crop for sugar and biofuel in tropical and subtropical regions and suffers serious losses in cane yield and sugar content under salinity and drought stresses. Although real-time quantitative PCR has a numerous advantage in the expression quantification of stress-related genes for the elaboration of the corresponding molecular mechanism in sugarcane, the variation happened across the process of gene expression quantification should be normalized and monitored by introducing one or several reference genes. To validate suitable reference genes or gene sets for sugarcane gene expression normalization, 13 candidate reference genes have been tested across 12 NaCl- and PEG-treated sugarcane samples for four sugarcane genotypes using four commonly used systematic statistical algorithms termed geNorm, BestKeeper, NormFinder and the deltaCt method. The results demonstrated that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and eukaryotic elongation factor 1-alpha (eEF-1a) were identified as suitable reference genes for gene expression normalization under salinity/drought-treatment in sugarcane. Moreover, the expression analyses of SuSK and 6PGDH further validated that a combination of clathrin adaptor complex (CAC) and cullin (CUL) as reference should be better for gene expression normalization. These results can facilitate the future research on gene expression in sugarcane under salinity and drought stresses.

  4. The gsdf gene locus harbors evolutionary conserved and clustered genes preferentially expressed in fish previtellogenic oocytes.

    PubMed

    Gautier, Aude; Le Gac, Florence; Lareyre, Jean-Jacques

    2011-02-01

    The gonadal soma-derived factor (GSDF) belongs to the transforming growth factor-β superfamily and is conserved in teleostean fish species. Gsdf is specifically expressed in the gonads, and gene expression is restricted to the granulosa and Sertoli cells in trout and medaka. The gsdf gene expression is correlated to early testis differentiation in medaka and was shown to stimulate primordial germ cell and spermatogonia proliferation in trout. In the present study, we show that the gsdf gene localizes to a syntenic chromosomal fragment conserved among vertebrates although no gsdf-related gene is detected on the corresponding genomic region in tetrapods. We demonstrate using quantitative RT-PCR that most of the genes localized in the synteny are specifically expressed in medaka gonads. Gsdf is the only gene of the synteny with a much higher expression in the testis compared to the ovary. In contrast, gene expression pattern analysis of the gsdf surrounding genes (nup54, aff1, klhl8, sdad1, and ptpn13) indicates that these genes are preferentially expressed in the female gonads. The tissue distribution of these genes is highly similar in medaka and zebrafish, two teleostean species that have diverged more than 110 million years ago. The cellular localization of these genes was determined in medaka gonads using the whole-mount in situ hybridization technique. We confirm that gsdf gene expression is restricted to Sertoli and granulosa cells in contact with the premeiotic and meiotic cells. The nup54 gene is expressed in spermatocytes and previtellogenic oocytes. Transcripts corresponding to the ovary-specific genes (aff1, klhl8, and sdad1) are detected only in previtellogenic oocytes. No expression was detected in the gonocytes in 10 dpf embryos. In conclusion, we show that the gsdf gene localizes to a syntenic chromosomal fragment harboring evolutionary conserved genes in vertebrates. These genes are preferentially expressed in previtelloogenic oocytes, and thus, they

  5. Novel redox nanomedicine improves gene expression of polyion complex vector

    NASA Astrophysics Data System (ADS)

    Toh, Kazuko; Yoshitomi, Toru; Ikeda, Yutaka; Nagasaki, Yukio

    2011-12-01

    Gene therapy has generated worldwide attention as a new medical technology. While non-viral gene vectors are promising candidates as gene carriers, they have several issues such as toxicity and low transfection efficiency. We have hypothesized that the generation of reactive oxygen species (ROS) affects gene expression in polyplex supported gene delivery systems. The effect of ROS on the gene expression of polyplex was evaluated using a nitroxide radical-containing nanoparticle (RNP) as an ROS scavenger. When polyethyleneimine (PEI)/pGL3 or PEI alone was added to the HeLa cells, ROS levels increased significantly. In contrast, when (PEI)/pGL3 or PEI was added with RNP, the ROS levels were suppressed. The luciferase expression was increased by the treatment with RNP in a dose-dependent manner and the cellular uptake of pDNA was also increased. Inflammatory cytokines play an important role in ROS generation in vivo. In particular, tumor necrosis factor (TNF)-α caused intracellular ROS generation in HeLa cells and decreased gene expression. RNP treatment suppressed ROS production even in the presence of TNF-α and increased gene expression. This anti-inflammatory property of RNP suggests that it may be used as an effective adjuvant for non-viral gene delivery systems.

  6. Anterior-posterior regionalized gene expression in the Ciona notochord

    PubMed Central

    Veeman, Michael

    2014-01-01

    Background In the simple ascidian chordate Ciona the signaling pathways and gene regulatory networks giving rise to initial notochord induction are largely understood and the mechanisms of notochord morphogenesis are being systematically elucidated. The notochord has generally been thought of as a non-compartmentalized or regionalized organ that is not finely patterned at the level of gene expression. Quantitative imaging methods have recently shown, however, that notochord cell size, shape and behavior vary consistently along the anterior-posterior (AP) axis. Results Here we screen candidate genes by whole mount in situ hybridization for potential AP asymmetry. We identify 4 genes that show non-uniform expression in the notochord. Ezrin/radixin/moesin (ERM) is expressed more strongly in the secondary notochord lineage than the primary. CTGF is expressed stochastically in a subset of notochord cells. A novel calmodulin-like gene (BCamL) is expressed more strongly at both the anterior and posterior tips of the notochord. A TGF-β ortholog is expressed in a gradient from posterior to anterior. The asymmetries in ERM, BCamL and TGF-β expression are evident even before the notochord cells have intercalated into a single-file column. Conclusions We conclude that the Ciona notochord is not a homogeneous tissue but instead shows distinct patterns of regionalized gene expression. PMID:24288133

  7. Validation of housekeeping genes for gene expression studies in an ice alga Chlamydomonas during freezing acclimation.

    PubMed

    Liu, Chenlin; Wu, Guangting; Huang, Xiaohang; Liu, Shenghao; Cong, Bailin

    2012-05-01

    Antarctic ice alga Chlamydomonas sp. ICE-L can endure extreme low temperature and high salinity stress under freezing conditions. To elucidate the molecular acclimation mechanisms using gene expression analysis, the expression stabilities of ten housekeeping genes of Chlamydomonas sp. ICE-L during freezing stress were analyzed. Some discrepancies were detected in the ranking of the candidate reference genes between geNorm and NormFinder programs, but there was substantial agreement between the groups of genes with the most and the least stable expression. RPL19 was ranked as the best candidate reference genes. Pairwise variation (V) analysis indicated the combination of two reference genes was sufficient for qRT-PCR data normalization under the experimental conditions. Considering the co-regulation between RPL19 and RPL32 (the most stable gene pairs given by geNorm program), we propose that the mean data rendered by RPL19 and GAPDH (the most stable gene pairs given by NormFinder program) be used to normalize gene expression values in Chlamydomonas sp. ICE-L more accurately. The example of FAD3 gene expression calculation demonstrated the importance of selecting an appropriate category and number of reference genes to achieve an accurate and reliable normalization of gene expression during freeze acclimation in Chlamydomonas sp. ICE-L.

  8. Macromolecular crowding creates heterogeneous environments of gene expression in picolitre droplets

    NASA Astrophysics Data System (ADS)

    Hansen, Maike M. K.; Meijer, Lenny H. H.; Spruijt, Evan; Maas, Roel J. M.; Rosquelles, Marta Ventosa; Groen, Joost; Heus, Hans A.; Huck, Wilhelm T. S.

    2016-02-01

    Understanding the dynamics of complex enzymatic reactions in highly crowded small volumes is crucial for the development of synthetic minimal cells. Compartmentalized biochemical reactions in cell-sized containers exhibit a degree of randomness due to the small number of molecules involved. However, it is unknown how the physical environment contributes to the stochastic nature of multistep enzymatic processes. Here, we present a robust method to quantify gene expression noise in vitro using droplet microfluidics. We study the changes in stochasticity in the cell-free gene expression of two genes compartmentalized within droplets as a function of DNA copy number and macromolecular crowding. We find that decreased diffusion caused by a crowded environment leads to the spontaneous formation of heterogeneous microenvironments of mRNA as local production rates exceed the diffusion rates of macromolecules. This heterogeneity leads to a higher probability of the molecular machinery staying in the same microenvironment, directly increasing the system's stochasticity.

  9. Measuring single-cell gene expression dynamics in bacteria using fluorescence time-lapse microscopy

    PubMed Central

    Young, Jonathan W; Locke, James C W; Altinok, Alphan; Rosenfeld, Nitzan; Bacarian, Tigran; Swain, Peter S; Mjolsness, Eric; Elowitz, Michael B

    2014-01-01

    Quantitative single-cell time-lapse microscopy is a powerful method for analyzing gene circuit dynamics and heterogeneous cell behavior. We describe the application of this method to imaging bacteria by using an automated microscopy system. This protocol has been used to analyze sporulation and competence differentiation in Bacillus subtilis, and to quantify gene regulation and its fluctuations in individual Escherichia coli cells. The protocol involves seeding and growing bacteria on small agarose pads and imaging the resulting microcolonies. Images are then reviewed and analyzed using our laboratory's custom MATLAB analysis code, which segments and tracks cells in a frame-to-frame method. This process yields quantitative expression data on cell lineages, which can illustrate dynamic expression profiles and facilitate mathematical models of gene circuits. With fast-growing bacteria, such as E. coli or B. subtilis, image acquisition can be completed in 1 d, with an additional 1–2 d for progressing through the analysis procedure. PMID:22179594

  10. Macromolecular crowding develops heterogeneous environments of gene expression in picoliter droplets

    PubMed Central

    Hansen, Maike M. K.; Meijer, Lenny H. H.; Spruijt, Evan; Maas, Roel J. M.; Roquelles, Marta Ventosa; Groen, Joost; Heus, Hans A.; Huck, Wilhelm T. S.

    2015-01-01

    Understanding the dynamics of complex enzymatic reactions in highly crowded small volumes is crucial for the development of synthetic minimal cells. Compartmentalised biochemical reactions in cell-sized containers exhibit a degree of randomness due to the small number of molecules involved. However, it is unknown how the physical environment contributes to the stochastic nature of multistep enzymatic processes. Here, we present a robust method to quantify gene expression noise in vitro using droplet microfluidics. We study the changes in stochasticity in cell-free gene expression of two genes compartmentalised within droplets as a function of DNA copy number and macromolecular crowding. We find that decreased diffusion caused by a crowded environment leads to the spontaneous formation of heterogeneous micro-environments of mRNA as local production rates exceed diffusion rates of macromolecules. This heterogeneity leads to a higher probability of the molecular machinery to stay in the same microenvironment, directly increasing the system’s stochasticity. PMID:26501750

  11. Standardization of Gene Expression Quantification by Absolute Real-Time qRT-PCR System Using a Single Standard for Marker and Reference Genes.

    PubMed

    Zhou, Yi-Hong; Raj, Vinay R; Siegel, Eric; Yu, Liping

    2010-08-16

    In the last decade, genome-wide gene expression data has been collected from a large number of cancer specimens. In many studies utilizing either microarray-based or knowledge-based gene expression profiling, both the validation of candidate genes and the identification and inclusion of biomarkers in prognosis-modeling has employed real-time quantitative PCR on reverse transcribed mRNA (qRT-PCR) because of its inherent sensitivity and quantitative nature. In qRT-PCR data analysis, an internal reference gene is used to normalize the variation in input sample quantity. The relative quantification method used in current real-time qRT-PCR analysis fails to ensure data comparability pivotal in identification of prognostic biomarkers. By employing an absolute qRT-PCR system that uses a single standard for marker and reference genes (SSMR) to achieve absolute quantification, we showed that the normalized gene expression data is comparable and independent of variations in the quantities of sample as well as the standard used for generating standard curves. We compared two sets of normalized gene expression data with same histological diagnosis of brain tumor from two labs using relative and absolute real-time qRT-PCR. Base-10 logarithms of the gene expression ratio relative to ACTB were evaluated for statistical equivalence between tumors processed by two different labs. The results showed an approximate comparability for normalized gene expression quantified using a SSMR-based qRT-PCR. Incomparable results were seen for the gene expression data using relative real-time qRT-PCR, due to inequality in molar concentration of two standards for marker and reference genes. Overall results show that SSMR-based real-time qRT-PCR ensures comparability of gene expression data much needed in establishment of prognostic/predictive models for cancer patients-a process that requires large sample sizes by combining independent sets of data.

  12. Gene Expression Profiling of Breast Cancer Brain Metastasis

    PubMed Central

    Lee, Ji Yun; Park, Kyunghee; Lee, Eunjin; Ahn, TaeJin; Jung, Hae Hyun; Lim, Sung Hee; Hong, Mineui; Do, In-Gu; Cho, Eun Yoon; Kim, Duk-Hwan; Kim, Ji-Yeon; Ahn, Jin Seok; Im, Young-Hyuck; Park, Yeon Hee

    2016-01-01

    The biology of breast cancer brain metastasis (BCBM) is poorly understood. We aimed to explore genes that are implicated in the process of brain metastasis of primary breast cancer (BC). NanoString nCounter Analysis covering 252 target genes was used for comparison of gene expression levels between 20 primary BCs that relapsed to brain and 41 BCBM samples. PAM50-based intrinsic subtypes such as HER2-enriched and basal-like were clearly over-represented in BCBM. A panel of 22 genes was found to be significantly differentially expressed between primary BC and BCBM. Five of these genes, CXCL12, MMP2, MMP11, VCAM1, and MME, which have previously been associated with tumor progression, angiogenesis, and metastasis, clearly discriminated between primary BC and BCBM. Notably, the five genes were significantly upregulated in primary BC compared to BCBM. Conversely, SOX2 and OLIG2 genes were upregulated in BCBM. These genes may participate in metastatic colonization but not in primary tumor development. Among patient-matched paired samples (n = 17), a PAM50 molecular subtype conversion was observed in eight cases (47.1%), with a trend toward unfavorable subtypes in patients with the distinct gene expression. Our findings, although not conclusive, reveal differentially expressed genes that might mediate the brain metastasis process. PMID:27340107

  13. Differential network analysis from cross-platform gene expression data

    PubMed Central

    Zhang, Xiao-Fei; Ou-Yang, Le; Zhao, Xing-Ming; Yan, Hong

    2016-01-01

    Understanding how the structure of gene dependency network changes between two patient-specific groups is an important task for genomic research. Although many computational approaches have been proposed to undertake this task, most of them estimate correlation networks from group-specific gene expression data independently without considering the common structure shared between different groups. In addition, with the development of high-throughput technologies, we can collect gene expression profiles of same patients from multiple platforms. Therefore, inferring differential networks by considering cross-platform gene expression profiles will improve the reliability of network inference. We introduce a two dimensional joint graphical lasso (TDJGL) model to simultaneously estimate group-specific gene dependency networks from gene expression profiles collected from different platforms and infer differential networks. TDJGL can borrow strength across different patient groups and data platforms to improve the accuracy of estimated networks. Simulation studies demonstrate that TDJGL provides more accurate estimates of gene networks and differential networks than previous competing approaches. We apply TDJGL to the PI3K/AKT/mTOR pathway in ovarian tumors to build differential networks associated with platinum resistance. The hub genes of our inferred differential networks are significantly enriched with known platinum resistance-related genes and include potential platinum resistance-related genes. PMID:27677586

  14. Differential network analysis from cross-platform gene expression data

    NASA Astrophysics Data System (ADS)

    Zhang, Xiao-Fei; Ou-Yang, Le; Zhao, Xing-Ming; Yan, Hong

    2016-09-01

    Understanding how the structure of gene dependency network changes between two patient-specific groups is an important task for genomic research. Although many computational approaches have been proposed to undertake this task, most of them estimate correlation networks from group-specific gene expression data independently without considering the common structure shared between different groups. In addition, with the development of high-throughput technologies, we can collect gene expression profiles of same patients from multiple platforms. Therefore, inferring differential networks by considering cross-platform gene expression profiles will improve the reliability of network inference. We introduce a two dimensional joint graphical lasso (TDJGL) model to simultaneously estimate group-specific gene dependency networks from gene expression profiles collected from different platforms and infer differential networks. TDJGL can borrow strength across different patient groups and data platforms to improve the accuracy of estimated networks. Simulation studies demonstrate that TDJGL provides more accurate estimates of gene networks and differential networks than previous competing approaches. We apply TDJGL to the PI3K/AKT/mTOR pathway in ovarian tumors to build differential networks associated with platinum resistance. The hub genes of our inferred differential networks are significantly enriched with known platinum resistance-related genes and include potential platinum resistance-related genes.

  15. Gene Expressions for Signal Transduction under Acidic Conditions

    PubMed Central

    Fukamachi, Toshihiko; Ikeda, Syunsuke; Wang, Xin; Saito, Hiromi; Tagawa, Masatoshi; Kobayashi, Hiroshi

    2013-01-01

    Although it is now well known that some diseased areas, such as cancer nests, inflammation loci, and infarction areas, are acidified, little is known about cellular signal transduction, gene expression, and cellular functions under acidic conditions. Our group showed that different signal proteins were activated under acidic conditions compared with those observed in a typical medium of around pH 7.4 that has been used until now. Investigations of gene expression under acidic conditions may be crucial to our understanding of signal transduction in acidic diseased areas. In this study, we investigated gene expression in mesothelioma cells cultured at an acidic pH using a DNA microarray technique. After 24 h culture at pH 6.7, expressions of 379 genes were increased more than twofold compared with those in cells cultured at pH 7.5. Genes encoding receptors, signal proteins including transcription factors, and cytokines including growth factors numbered 35, 32, and 17 among the 379 genes, respectively. Since the functions of 78 genes are unknown, it can be argued that cells may have other genes for signaling under acidic conditions. The expressions of 37 of the 379 genes were observed to increase after as little as 2 h. After 24 h culture at pH 6.7, expressions of 412 genes were repressed more than twofold compared with those in cells cultured at pH 7.5, and the 412 genes contained 35, 76, and 7 genes encoding receptors, signal proteins including transcription factors, and cytokines including growth factors, respectively. These results suggest that the signal pathways in acidic diseased areas are different, at least in part, from those examined with cells cultured at a pH of around 7.4. PMID:24705103

  16. Quantifying the effects of co-expressing EGFR and HER2 on HER activation and trafficking

    SciTech Connect

    Shankaran, Harish; Zhang, Yi; Opresko, Lee; Resat, Haluk

    2008-06-27

    The integration of experimental measurements and computational predictions is a powerful means to understand the molecular mechanisms of complex biological systems. The human epidermal growth factor receptor (HER) system is an intricately regulated system that plays critical roles in development and tumorigenesis. Here, we apply integrated experimentation and modeling to analyze HER receptor activation in a panel of cell lines expressing different levels of HER1 and HER2. A mathematical model that includes the fundamental biochemical/biophysical processes involved in receptor activation was used to fit the experimental data, and 19 independent parameters including receptor dimerization affinities, trafficking rates and relative phosphorylation levels were estimated. The parameter values quantitatively support existing ideas on the effect of HER2 on HER1 phosphorylation dynamics, and enable us to predict receptor dimerization patterns in the cell lines. The integrated approach described here can be applied to obtain a predictive understanding of other biomolecular systems.

  17. Membrane channel gene expression in human costal and articular chondrocytes.

    PubMed

    Asmar, A; Barrett-Jolley, R; Werner, A; Kelly, R; Stacey, M

    2016-04-01

    Chondrocytes are the uniquely resident cells found in all types of cartilage and key to their function is the ability to respond to mechanical loads with changes of metabolic activity. This mechanotransduction property is, in part, mediated through the activity of a range of expressed transmembrane channels; ion channels, gap junction proteins, and porins. Appropriate expression of ion channels has been shown essential for production of extracellular matrix and differential expression of transmembrane channels is correlated to musculoskeletal diseases such as osteoarthritis and Albers-Schönberg. In this study we analyzed the consistency of gene expression between channelomes of chondrocytes from human articular and costal (teenage and fetal origin) cartilages. Notably, we found 14 ion channel genes commonly expressed between articular and both types of costal cartilage chondrocytes. There were several other ion channel genes expressed only in articular (6 genes) or costal chondrocytes (5 genes). Significant differences in expression of BEST1 and KCNJ2 (Kir2.1) were observed between fetal and teenage costal cartilage. Interestingly, the large Ca(2+) activated potassium channel (BKα, or KCNMA1) was very highly expressed in all chondrocytes examined. Expression of the gap junction genes for Panx1, GJA1 (Cx43) and GJC1 (Cx45) was also observed in chondrocytes from all cartilage samples. Together, this data highlights similarities between chondrocyte membrane channel gene expressions in cells derived from different anatomical sites, and may imply that common electrophysiological signaling pathways underlie cellular control. The high expression of a range of mechanically and metabolically sensitive membrane channels suggest that chondrocyte mechanotransduction may be more complex than previously thought. PMID:27116676

  18. Membrane channel gene expression in human costal and articular chondrocytes

    PubMed Central

    Asmar, A.; Barrett-Jolley, R.; Werner, A.; Kelly, R.; Stacey, M.

    2016-01-01

    ABSTRACT Chondrocytes are the uniquely resident cells found in all types of cartilage and key to their function is the ability to respond to mechanical loads with changes of metabolic activity. This mechanotransduction property is, in part, mediated through the activity of a range of expressed transmembrane channels; ion channels, gap junction proteins, and porins. Appropriate expression of ion channels has been shown essential for production of extracellular matrix and differential expression of transmembrane channels is correlated to musculoskeletal diseases such as osteoarthritis and Albers-Schönberg. In this study we analyzed the consistency of gene expression between channelomes of chondrocytes from human articular and costal (teenage and fetal origin) cartilages. Notably, we found 14 ion channel genes commonly expressed between articular and both types of costal cartilage chondrocytes. There were several other ion channel genes expressed only in articular (6 genes) or costal chondrocytes (5 genes). Significant differences in expression of BEST1 and KCNJ2 (Kir2.1) were observed between fetal and teenage costal cartilage. Interestingly, the large Ca2+ activated potassium channel (BKα, or KCNMA1) was very highly expressed in all chondrocytes examined. Expression of the gap junction genes for Panx1, GJA1 (Cx43) and GJC1 (Cx45) was also observed in chondrocytes from all cartilage samples. Together, this data highlights similarities between chondrocyte membrane channel gene expressions in cells derived from different anatomical sites, and may imply that common electrophysiological signaling pathways underlie cellular control. The high expression of a range of mechanically and metabolically sensitive membrane channels suggest that chondrocyte mechanotransduction may be more complex than previously thought. PMID:27116676

  19. [Expression of the genes involved in anthocyanin biosynthesis of 'Tsuda' turnip].

    PubMed

    Xu, Zhi-Ru; Li, Yu-Hua

    2006-10-01

    'Tsuda' turnip (Brassica campestris L. ssp. rapa), in which roots anthocyanin pigmentation is light-sensitive, was used as the material. 'Tsuda' plants were held in darkness or irradiated with sun light and constant light for different time. Anthocyanins in root peel of 'Tsuda' turnip exposed to constant light were identified and quantified with a UV-visual spectrophotometer. The results demonstrated that the anthocyanins accumulation in 'Tsuda' was related with light-exposure time (Fig.1 and Table 1). Fragments of genes selected from the subtraction library of 'Tsuda' turnip involved in anthocyanin biosynthesis were used as probes. The Northern blotting results showed that the expression of PAL, CHS, F3H, DFR and ANS could be induced by irradiation with light and the expression of these genes was related with light exposure time. The expression of MYB was basically the same whether in darkness or in light (Figs.2,3).

  20. [Expression of the genes involved in anthocyanin biosynthesis of 'Tsuda' turnip].

    PubMed

    Xu, Zhi-Ru; Li, Yu-Hua

    2006-10-01

    'Tsuda' turnip (Brassica campestris L. ssp. rapa), in which roots anthocyanin pigmentation is light-sensitive, was used as the material. 'Tsuda' plants were held in darkness or irradiated with sun light and constant light for different time. Anthocyanins in root peel of 'Tsuda' turnip exposed to constant light were identified and quantified with a UV-visual spectrophotometer. The results demonstrated that the anthocyanins accumulation in 'Tsuda' was related with light-exposure time (Fig.1 and Table 1). Fragments of genes selected from the subtraction library of 'Tsuda' turnip involved in anthocyanin biosynthesis were used as probes. The Northern blotting results showed that the expression of PAL, CHS, F3H, DFR and ANS could be induced by irradiation with light and the expression of these genes was related with light exposure time. The expression of MYB was basically the same whether in darkness or in light (Figs.2,3). PMID:17075183

  1. Control of alphavirus-based gene expression using engineered riboswitches.

    PubMed

    Bell, Christie L; Yu, Dong; Smolke, Christina D; Geall, Andrew J; Beard, Clayton W; Mason, Peter W

    2015-09-01

    Alphavirus-based replicons are a promising nucleic acid vaccine platform characterized by robust gene expression and immune responses. To further explore their use in vaccination, replicons were engineered to allow conditional control over their gene expression. Riboswitches, comprising a ribozyme actuator and RNA aptamer sensor, were engineered into the replicon 3' UTR. Binding of ligand to aptamer modulates ribozyme activity and, therefore, gene expression. Expression from DNA-launched and VRP-packaged replicons containing riboswitches was successfully regulated, achieving a 47-fold change in expression and modulation of the resulting type I interferon response. Moreover, we developed a novel control architecture where riboswitches were integrated into the 3' and 5' UTR of the subgenomic RNA region of the TC-83 virus, leading to an 1160-fold regulation of viral replication. Our studies demonstrate that the use of riboswitches for control of RNA replicon expression and viral replication holds promise for development of novel and safer vaccination strategies.

  2. Control of alphavirus-based gene expression using engineered riboswitches.

    PubMed

    Bell, Christie L; Yu, Dong; Smolke, Christina D; Geall, Andrew J; Beard, Clayton W; Mason, Peter W

    2015-09-01

    Alphavirus-based replicons are a promising nucleic acid vaccine platform characterized by robust gene expression and immune responses. To further explore their use in vaccination, replicons were engineered to allow conditional control over their gene expression. Riboswitches, comprising a ribozyme actuator and RNA aptamer sensor, were engineered into the replicon 3' UTR. Binding of ligand to aptamer modulates ribozyme activity and, therefore, gene expression. Expression from DNA-launched and VRP-packaged replicons containing riboswitches was successfully regulated, achieving a 47-fold change in expression and modulation of the resulting type I interferon response. Moreover, we developed a novel control architecture where riboswitches were integrated into the 3' and 5' UTR of the subgenomic RNA region of the TC-83 virus, leading to an 1160-fold regulation of viral replication. Our studies demonstrate that the use of riboswitches for control of RNA replicon expression and viral replication holds promise for development of novel and safer vaccination strategies. PMID:26005949

  3. Gene Expression Network Reconstruction by LEP Method Using Microarray Data

    PubMed Central

    You, Na; Mou, Peng; Qiu, Ting; Kou, Qiang; Zhu, Huaijin; Chen, Yuexi; Wang, Xueqin

    2012-01-01

    Gene expression network reconstruction using microarray data is widely studied aiming to investigate the behavior of a gene cluster simultaneously. Under the Gaussian assumption, the conditional dependence between genes in the network is fully described by the partial correlation coefficient matrix. Due to the high dimensionality and sparsity, we utilize the LEP method to estimate it in this paper. Compared to the existing methods, the LEP reaches the highest PPV with the sensitivity controlled at the satisfactory level. A set of gene expression data from the HapMap project is analyzed for illustration. PMID:23365528

  4. The Role of Nuclear Bodies in Gene Expression and Disease

    PubMed Central

    Morimoto, Marie; Boerkoel, Cornelius F.

    2013-01-01

    This review summarizes the current understanding of the role of nuclear bodies in regulating gene expression. The compartmentalization of cellular processes, such as ribosome biogenesis, RNA processing, cellular response to stress, transcription, modification and assembly of spliceosomal snRNPs, histone gene synthesis and nuclear RNA retention, has significant implications for gene regulation. These functional nuclear domains include the nucleolus, nuclear speckle, nuclear stress body, transcription factory, Cajal body, Gemini of Cajal body, histone locus body and paraspeckle. We herein review the roles of nuclear bodies in regulating gene expression and their relation to human health and disease. PMID:24040563

  5. Scaling of Gene Expression with Transcription-Factor Fugacity

    PubMed Central

    Weinert, Franz M.; Brewster, Robert C.; Rydenfelt, Mattias; Phillips, Rob; Kegel, Willem K.

    2015-01-01

    The proteins associated with gene regulation are often shared between multiple pathways simultaneously. By way of contrast, models in regulatory biology often assume these pathways act independently. We demonstrate a framework for calculating the change in gene expression for the interacting case by decoupling repressor occupancy across the cell from the gene of interest by way of a chemical potential. The details of the interacting regulatory architecture are encompassed in an effective concentration, and thus, a single scaling function describes a collection of gene expression data from diverse regulatory situations and collapses it onto a single master curve. PMID:25554908

  6. Integration of biological networks and gene expression data using Cytoscape.

    PubMed

    Cline, Melissa S; Smoot, Michael; Cerami, Ethan; Kuchinsky, Allan; Landys, Nerius; Workman, Chris; Christmas, Rowan; Avila-Campilo, Iliana; Creech, Michael; Gross, Benjamin; Hanspers, Kristina; Isserlin, Ruth; Kelley, Ryan; Killcoyne, Sarah; Lotia, Samad; Maere, Steven; Morris, John; Ono, Keiichiro; Pavlovic, Vuk; Pico, Alexander R; Vailaya, Aditya; Wang, Peng-Liang; Adler, Annette; Conklin, Bruce R; Hood, Leroy; Kuiper, Martin; Sander, Chris; Schmulevich, Ilya; Schwikowski, Benno; Warner, Guy J; Ideker, Trey; Bader, Gary D

    2007-01-01

    Cytoscape is a free software package for visualizing, modeling and analyzing molecular and genetic interaction networks. This protocol explains how to use Cytoscape to analyze the results of mRNA expression profiling, and other functional genomics and proteomics experiments, in the context of an interaction network obtained for genes of interest. Five major steps are described: (i) obtaining a gene or protein network, (ii) displaying the network using layout algorithms, (iii) integrating with gene expression and other functional attributes, (iv) identifying putative complexes and functional modules and (v) identifying enriched Gene Ontology annotations in the network. These steps provide a broad sample of the types of analyses performed by Cytoscape.

  7. Structure and expression of canary myc family genes.

    PubMed Central

    Collum, R G; Clayton, D F; Alt, F W

    1991-01-01

    We found that the canary N-myc gene is highly related to mammalian N-myc genes in both the protein-coding region and the long 3' untranslated region. Examined coding regions of the canary c-myc gene were also highly related to their mammalian counterparts, but in contrast to N-myc, the canary and mammalian c-myc genes were quite divergent in their 3' untranslated regions. We readily detected N-myc and c-myc expression in the adult canary brain and found N-myc expression both at sites of proliferating neuronal precursors and in mature neurons. Images PMID:1996121

  8. Reference gene validation for quantification of gene expression during final oocyte maturation induced by diethylstilbestrol and di-(2-ethylhexyl)-phthalate in common carp.

    PubMed

    Shi, Yanyan; Lu, Jie; Wang, Yilei; Wang, Shuhong

    2016-08-01

    Final oocyte maturation is the key step to successful spawning and fertilization. Quantitative real-time PCR (qPCR) is the technique of election to quantify the abundance of functional genes in such study. Reference gene is essential for correct interpretation of qPCR data. However, an ideal universal reference gene that is stable under all experimental circumstances has not been described. Researchers should validate their reference genes while performing qPCR analysis. The expression of 6 candidate reference genes: 18s rRNA, 28s rRNA, Cathepsin Z, Elongation factor 1-α, Glyceraldehyde-3-phosphate dehydrogenase and β-actin were investigated during final oocyte maturation induced by different compounds (DES and DEHP) in common carp (Cyprinus carpio). Four softwares (Bestkeeper, geNorm, NormFinder and RefFinder) were used to screen the most stable gene in order to evaluate their expression stability. The results revealed that EF1α was highly stable expressed when final oocyte maturation was induced by DES, while gapdh was the most stable gene when final oocyte maturation was induced by DEHP. Stable expressed reference gene selection is critical for all qPCR analysis to get accurate target gene mRNA expression information. PMID:27521935

  9. Genetic alteration and gene expression modulation during cancer progression

    PubMed Central

    Garnis, Cathie; Buys, Timon PH; Lam, Wan L

    2004-01-01

    Cancer progresses through a series of histopathological stages. Progression is thought to be driven by the accumulation of genetic alterations and consequently gene expression pattern changes. The identification of genes and pathways involved will not only enhance our understanding of the biology of this process, it will also provide new targets for early diagnosis and facilitate treatment design. Genomic approaches have proven to be effective in detecting chromosomal alterations and identifying genes disrupted in cancer. Gene expression profiling has led to the subclassification of tumors. In this article, we will describe the current technologies used in cancer gene discovery, the model systems used to validate the significance of the genes and pathways, and some of the genes and pathways implicated in the progression of preneoplastic and early stage cancer. PMID:15035667

  10. A model for gene deregulation detection using expression data

    PubMed Central

    2015-01-01

    In tumoral cells, gene regulation mechanisms are severely altered. Genes that do not react normally to their regulators' activity can provide explanations for the tumoral behavior, and be characteristic of cancer subtypes. We thus propose a statistical methodology to identify the misregulated genes given a reference network and gene expression data. Our model is based on a regulatory process in which all genes are allowed to be deregulated. We derive an EM algorithm where the hidden variables correspond to the status (under/over/normally expressed) of the genes and where the E-step is solved thanks to a message passing algorithm. Our procedure provides posterior probabilities of deregulation in a given sample for each gene. We assess the performance of our method by numerical experiments on simulations and on a bladder cancer data set. PMID:26679516

  11. Evolution of gene expression and expression plasticity in long-term experimental populations of Drosophila melanogaster maintained under constant and variable ethanol stress.

    PubMed

    Yampolsky, Lev Y; Glazko, Galina V; Fry, James D

    2012-09-01

    Gene expression responds to the environment and can also evolve rapidly in response to altered selection regimes. Little is known, however, about the extent to which evolutionary adaptation to a particular type of stress involves changes in the within-generation ('plastic') responses of gene expression to the stress. We used microarrays to quantify gene expression plasticity in response to ethanol in laboratory populations of Drosophila melanogaster differing in their history of ethanol exposure. Two populations ('R' populations) were maintained on regular medium, two ('E') were maintained on medium supplemented with ethanol, and two ('M') were maintained in a mixed regime in which half of the population was reared on one medium type, and half on the other, each generation. After more than 300 generations, embryos from each population were collected and exposed to either ethanol or water as a control, and RNA was extracted from the larvae shortly after hatching. Nearly 2000 transcripts showed significant within-generation responses to ethanol exposure. Evolutionary history also affected gene expression: the E and M populations were largely indistinguishable in expression, but differed significantly in expression from the R populations for over 100 transcripts, the majority of which did not show plastic responses. Notably, in no case was the interaction between selection regime and ethanol exposure significant after controlling for multiple comparisons, indicating that adaptation to ethanol in the E and M populations did not involve substantial changes in gene expression plasticity. The results give evidence that expression plasticity evolves considerably more slowly than mean expression.

  12. Gene expression changes during retinal development and rod specification

    PubMed Central

    Carrigan, Matthew; Hokamp, Karsten; Farrar, G. Jane

    2015-01-01

    Purpose Retinitis pigmentosa (RP) typically results from individual mutations in any one of >70 genes that cause rod photoreceptor cells to degenerate prematurely, eventually resulting in blindness. Gene therapies targeting individual RP genes have shown efficacy at clinical trial; however, these therapies require the surviving photoreceptor cells to be viable and functional, and may be economically feasible for only the more commonly mutated genes. An alternative potential treatment strategy, particularly for late stage disease, may involve stem cell transplants into the photoreceptor layer of the retina. Rod progenitors from postnatal mouse retinas can be transplanted and can form photoreceptors in recipient adult retinas; optimal numbers of transplantable cells are obtained from postnatal day 3–5 (P3–5) retinas. These cells can also be expanded in culture; however, this results in the loss of photoreceptor potential. Gene expression differences between postnatal retinas, cultured retinal progenitor cells (RPCs), and rod photoreceptor precursors were investigated to identify gene expression patterns involved in the specification of rod photoreceptors. Methods Microarrays were used to investigate differences in gene expression between cultured RPCs that have lost photoreceptor potential, P1 retinas, and fresh P5 retinas that contain significant numbers of transplantable photoreceptors. Additionally, fluorescence-activated cell sorting (FACS) sorted Rho-eGFP-expressing rod photoreceptor precursors were compared with Rho-eGFP-negative cells from the same P5 retinas. Differential expression was confirmed with quantitative polymerase chain reaction (q-PCR). Results Analysis of the microarray data sets, including the use of t-distributed stochastic neighbor embedding (t-SNE) to identify expression pattern neighbors of key photoreceptor specific genes, resulted in the identification of 636 genes differentially regulated during rod specification. Forty-four of these

  13. Identification of Reference Genes in Human Myelomonocytic Cells for Gene Expression Studies in Altered Gravity

    PubMed Central

    Thiel, Cora S.; Hauschild, Swantje; Tauber, Svantje; Paulsen, Katrin; Raig, Christiane; Raem, Arnold; Biskup, Josefine; Gutewort, Annett; Hürlimann, Eva; Philpot, Claudia; Lier, Hartwin; Engelmann, Frank; Layer, Liliana E.

    2015-01-01

    Gene expression studies are indispensable for investigation and elucidation of molecular mechanisms. For the process of normalization, reference genes (“housekeeping genes”) are essential to verify gene expression analysis. Thus, it is assumed that these reference genes demonstrate similar expression levels over all experimental conditions. However, common recommendations about reference genes were established during 1 g conditions and therefore their applicability in studies with altered gravity has not been demonstrated yet. The microarray technology is frequently used to generate expression profiles under defined conditions and to determine the relative difference in expression levels between two or more different states. In our study, we searched for potential reference genes with stable expression during different gravitational conditions (microgravity, normogravity, and hypergravity) which are additionally not altered in different hardware systems. We were able to identify eight genes (ALB, B4GALT6, GAPDH, HMBS, YWHAZ, ABCA5, ABCA9, and ABCC1) which demonstrated no altered gene expression levels in all tested conditions and therefore represent good candidates for the standardization of gene expression studies in altered gravity. PMID:25654098

  14. VH gene expression and regulation in the mutant Alicia rabbit. Rescue of VHa2 allotype expression.

    PubMed

    Chen, H T; Alexander, C B; Young-Cooper, G O; Mage, R G

    1993-04-01

    Rabbits of the Alicia strain, derived from rabbits expressing the VHa2 allotype, have a mutation in the H chain locus that has a cis effect upon the expression of VHa2 and VHa- genes. A small deletion at the most J-proximal (3') end of the VH locus leads to low expression of all the genes on the entire chromosome in heterozygous ali mutants and altered relative expression of VH genes in homozygotes. To study VH gene expression and regulation, we used the polymerase chain reaction to amplify the VH genes expressed in spleens of young and adult wild-type and mutant Alicia rabbits. The cDNA from reverse transcription of splenic mRNA was amplified and polymerase chain reaction libraries were constructed and screened with oligonucleotides from framework regions 1 and 3, as well as JH. Thirty-three VH-positive clones were sequenced and analyzed. We found that in mutant Alicia rabbits, products of the first functional VH gene (VH4a2), (or VH4a2-like genes) were expressed in 2- to 8-wk-olds. Expression of both the VHx and VHy types of VHa- genes was also elevated but the relative proportions of VHx and VHy, especially VHx, decreased whereas the relative levels of expression of VH4a2 or VH4a2-like genes increased with age. Our results suggest that the appearance of sequences resembling that of the VH1a2, which is deleted in the mutant ali rabbits, could be caused by alterations of the sequences of the rearranged VH4a2 genes by gene conversions and/or rearrangement of upstream VH1a2-like genes later in development.

  15. Imputing Gene Expression in Uncollected Tissues Within and Beyond GTEx

    PubMed Central

    Wang, Jiebiao; Gamazon, Eric R.; Pierce, Brandon L.; Stranger, Barbara E.; Im, Hae Kyung; Gibbons, Robert D.; Cox, Nancy J.; Nicolae, Dan L.; Chen, Lin S.

    2016-01-01

    Gene expression and its regulation can vary substantially across tissue types. In order to generate knowledge about gene expression in human tissues, the Genotype-Tissue Expression (GTEx) program has collected transcriptome data in a wide variety of tissue types from post-mortem donors. However, many tissue types are difficult to access and are not collected in every GTEx individual. Furthermore, in non-GTEx studies, the accessibility of certain tissue types greatly limits the feasibility and scale of studies of multi-tissue expression. In this work, we developed multi-tissue imputation methods to impute gene expression in uncollected or inaccessible tissues. Via simulation studies, we showed that the proposed methods outperform existing imputation methods in multi-tissue expression imputation and that incorporating imputed expression data can improve power to detect phenotype-expression correlations. By analyzing data from nine selected tissue types in the GTEx pilot project, we demonstrated that harnessing expression quantitative trait loci (eQTLs) and tissue-tissue expression-level correlations can aid imputation of transcriptome data from uncollected GTEx tissues. More importantly, we showed that by using GTEx data as a reference, one can impute expression levels in inaccessible tissues in non-GTEx expression studies. PMID:27040689

  16. Imputing Gene Expression in Uncollected Tissues Within and Beyond GTEx.

    PubMed

    Wang, Jiebiao; Gamazon, Eric R; Pierce, Brandon L; Stranger, Barbara E; Im, Hae Kyung; Gibbons, Robert D; Cox, Nancy J; Nicolae, Dan L; Chen, Lin S

    2016-04-01

    Gene expression and its regulation can vary substantially across tissue types. In order to generate knowledge about gene expression in human tissues, the Genotype-Tissue Expression (GTEx) program has collected transcriptome data in a wide variety of tissue types from post-mortem donors. However, many tissue types are difficult to access and are not collected in every GTEx individual. Furthermore, in non-GTEx studies, the accessibility of certain tissue types greatly limits the feasibility and scale of studies of multi-tissue expression. In this work, we developed multi-tissue imputation methods to impute gene expression in uncollected or inaccessible tissues. Via simulation studies, we showed that the proposed methods outperform existing imputation methods in multi-tissue expression imputation and that incorporating imputed expression data can improve power to detect phenotype-expression correlations. By analyzing data from nine selected tissue types in the GTEx pilot project, we demonstrated that harnessing expression quantitative trait loci (eQTLs) and tissue-tissue expression-level correlations can aid imputation of transcriptome data from uncollected GTEx tissues. More importantly, we showed that by using GTEx data as a reference, one can impute expression levels in inaccessible tissues in non-GTEx expression studies.

  17. Profiling of chicken adipose tissue gene expression by genome array

    PubMed Central

    Wang, Hong-Bao; Li, Hui; Wang, Qi-Gui; Zhang, Xin-Yu; Wang, Shou-Zhi; Wang, Yu-Xiang; Wang, Xiu-Ping

    2007-01-01

    Background Excessive accumulation of lipids in the adipose tissue is a major problem in the present-day broiler industry. However, few studies have analyzed the expression of adipose tissue genes that are involved in pathways and mechanisms leading to adiposity in chickens. Gene expression profiling of chicken adipose tissue could provide key information about the ontogenesis of fatness and clarify the molecular mechanisms underlying obesity. In this study, Chicken Genome Arrays were used to construct an adipose tissue gene expression profile of 7-week-old broilers, and to screen adipose tissue genes that are differentially expressed in lean and fat lines divergently selected over eight generations for high and low abdominal fat weight. Results The gene expression profiles detected 13,234–16,858 probe sets in chicken adipose tissue at 7 weeks, and genes involved in lipid metabolism and immunity such as fatty acid binding protein (FABP), thyroid hormone-responsive protein (Spot14), lipoprotein lipase(LPL), insulin-like growth factor binding protein 7(IGFBP7) and major histocompatibility complex (MHC), were highly expressed. In contrast, some genes related to lipogenesis, such as leptin receptor, sterol regulatory element binding proteins1 (SREBP1), apolipoprotein B(ApoB) and insulin-like growth factor 2(IGF2), were not detected. Moreover, 230 genes that were differentially expressed between the two lines were screened out; these were mainly involved in lipid metabolism, signal transduction, energy metabolism, tumorigenesis and immunity. Subsequently, real-time RT-PCR was performed to validate fifteen differentially expressed genes screened out by the microarray approach and high consistency was observed between the two methods. Conclusion Our results establish the groundwork for further studies of the basic genetic control of growth and development of chicken adipose tissue, and will be beneficial in clarifying the molecular mechanism of obesity in chickens. PMID

  18. Carbon Catabolite Repression Regulates Glyoxylate Cycle Gene Expression in Cucumber.

    PubMed Central

    Graham, I. A.; Denby, K. J.; Leaver, C. J.

    1994-01-01

    We have previously proposed that metabolic status is important in the regulation of cucumber malate synthase (MS) and isocitrate lyase (ICL) gene expression during plant development. In this article, we used a cell culture system to demonstrate that intracellular metabolic status does influence expression of both of these genes. Starvation of cucumber cell cultures resulted in the coordinate induction of the expression of MS and ICL genes, and this effect was reversed when sucrose was returned to the culture media. The induction of gene expression was closely correlated with a drop in intracellular sucrose, glucose, and fructose below threshold concentrations, but it was not correlated with a decrease in respiration rate. Glucose, fructose, or raffinose in the culture media also resulted in repression of MS and ICL. Both 2-deoxyglucose and mannose, which are phosphorylated by hexokinase but not further metabolized, specifically repressed MS and ICL gene expression relative to a third glyoxylate cycle gene, malate dehydrogenase. However, the addition of 3-methylglucose, an analog of glucose that is not phosphorylated, did not result in repression of either MS or ICL. It is proposed that the signal giving rise to a change in gene expression originates from the intracellular concentration of hexose sugars or the flux of hexose sugars into glycolysis. PMID:12244257

  19. All-optical regulation of gene expression in targeted cells

    NASA Astrophysics Data System (ADS)

    Wang, Yisen; He, Hao; Li, Shiyang; Liu, Dayong; Lan, Bei; Hu, Minglie; Cao, Youjia; Wang, Chingyue

    2014-06-01

    Controllable gene expression is always a challenge and of great significance to biomedical research and clinical applications. Recently, various approaches based on extra-engineered light-sensitive proteins have been developed to provide optogenetic actuators for gene expression. Complicated biomedical techniques including exogenous genes engineering, transfection, and material delivery are needed. Here we present an all-optical method to regulate gene expression in targeted cells. Intrinsic or exogenous genes can be activated by a Ca2+-sensitive transcription factor nuclear factor of activated T cells (NFAT) driven by a short flash of femtosecond-laser irradiation. When applied to mesenchymal stem cells, expression of a differentiation regulator Osterix can be activated by this method to potentially induce differentiation of them. A laser-induced ``Ca2+-comb'' (LiCCo) by multi-time laser exposure is further developed to enhance gene expression efficiency. This noninvasive method hence provides an encouraging advance of gene expression regulation, with promising potential of applying in cell biology and stem-cell science.

  20. [Expression of bioinformatically identified genes in skin of psoriasis patients].

    PubMed

    2013-10-01

    Gene expression analysis for EPHA2 (EPH receptor A2), EPHB2 (EPH receptor B2), S100A9 (S100 calcium binding protein A9), PBEF(nicotinamide phosphoribosyltransferase), LILRB2 (leukocyte immunoglobulin-like receptor, subfamily B (with TM and ITIM domains), member 2), PLAUR (plasminogen activator, urokinase receptor), LTB (lymphotoxin beta (TNF superfamily, member 3)), WNT5A (wingless-type MMTV integration site family, member 5A) has been conducted using real-time polymerase chain reaction in specimens affected by psoriasis versus visually intact skin in 18 patients. It was revealed that the expression of the nine examined genes was upregulated in the affected by psoriasis compared to visually intact skin specimens. The highest expression was observed for S100A9, S100AS, PBEF, WNT5A2, and EPHB2 genes. S100A9 and S100A8 gene expression in the affected by psoriasis skin was 100-fold higher versus visually intact skin while PBEF, WNT5A, and EPHB2 gene expression was upregulated more than five-fold. We suggested that the high expression of these genes might be associated with the state of the pathological process in psoriasis. Moreover, the transcriptional activity of these genes might serve a molecular indicator of the efficacy of treatment in psoriasis. PMID:25508677

  1. [Expression of bioinformatically identified genes in skin of psoriasis patients].

    PubMed

    Sobolev, V V; Nikol'skaia, T A; Zolotarenko, A D; Piruzian, E S; Bruskin, S A

    2013-10-01

    Gene expression analysis for EPHA2 (EPH receptor A2), EPHB2 (EPH receptor B2), S100A9 (S100 calcium binding protein A9), PBEF(nicotinamide phosphoribosyltransferase), LILRB2 (leukocyte immunoglobulin-like receptor, subfamily B (with TM and ITIM domains), member 2), PLAUR (plasminogen activator, urokinase receptor), LTB (lymphotoxin beta (TNF superfamily, member 3)), WNT5A (wingless-type MMTV integration site family, member 5A) has been conducted using real-time polymerase chain reaction in specimens affected by psoriasis versus visually intact skin in 18 patients. It was revealed that the expression of the nine examined genes was upregulated in the affected by psoriasis compared to visually intact skin specimens. The highest expression was observed for S100A9, S100AS, PBEF, WNT5A2, and EPHB2 genes. S100A9 and S100A8 gene expression in the affected by psoriasis skin was 100-fold higher versus visually intact skin while PBEF, WNT5A, and EPHB2 gene expression was upregulated more than five-fold. We suggested that the high expression of these genes might be associated with the state of the pathological process in psoriasis. Moreover, the transcriptional activity of these genes might serve a molecular indicator of the efficacy of treatment in psoriasis. PMID:25474898

  2. Influence of mitochondria on gene expression in a citrus cybrid.

    PubMed

    Bassene, Jean-Baptiste; Froelicher, Yann; Navarro, Luis; Ollitrault, Patrick; Ancillo, Gema

    2011-06-01

    The production of cybrids, combining nucleus of a species with alien cytoplasmic organelles, is a valuable method used for improvement of various crops. Several citrus cybrids have been created by somatic hybridization. These genotypes are interesting models to analyze the impact of cytoplasmic genome change on nuclear genome expression. Herein, we report genome-wide gene expression analysis in leaves of a citrus cybrid between C. reticulata cv 'Willowleaf mandarin' and C. limon cv 'Eureka lemon' compared with its lemon parent, using a Citrus 20K cDNA microarray. Molecular analysis showed that this cybrid possesses nuclear and chloroplast genomes of Eureka lemon plus mitochondria from Willowleaf mandarin and, therefore, can be considered as a lemon bearing foreign mitochondria. Mandarin mitochondria influenced the expression of a large set of lemon nuclear genes causing an over-expression of 480 of them and repression of 39 genes. Quantitative real-time RT-PCR further confirmed the credibility of microarray data. Genes over-expressed in cybrid leaves are predominantly attributed to the functional category "cellular protein metabolism" whereas in the down-regulated none functional category was enriched. Overall, mitochondria replacement affected different nuclear genes including particularly genes predicted to be involved in mitochondrial retrograde signaling. Mitochondria regulate all cell structures even chloroplast status. These results suggest that nuclear gene expression is modulated with respect to new information received from the foreign organelle, with the final objective to suit specific needs to ensure better cell physiological balance.

  3. Global Gene Expression Analysis of Murine Limb Development

    PubMed Central

    Taher, Leila; Collette, Nicole M.; Murugesh, Deepa; Maxwell, Evan; Ovcharenko, Ivan; Loots, Gabriela G.

    2011-01-01

    Detailed information about stage-specific changes in gene expression is crucial for understanding the gene regulatory networks underlying development and the various signal transduction pathways contributing to morphogenesis. Here we describe the global gene expression dynamics during early murine limb development, when cartilage, tendons, muscle, joints, vasculature and nerves are specified and the musculoskeletal system of limbs is established. We used whole-genome microarrays to identify genes with differential expression at 5 stages of limb development (E9.5 to 13.5), during fore- and hind-limb patterning. We found that the onset of limb formation is characterized by an up-regulation of transcription factors, which is followed by a massive activation of genes during E10.5 and E11.5 which levels off at later time points. Among the 3520 genes identified as significantly up-regulated in the limb, we find ∼30% to be novel, dramatically expanding the repertoire of candidate genes likely to function in the limb. Hierarchical and stage-specific clustering identified expression profiles that are likely to correlate with functional programs during limb development and further characterization of these transcripts will provide new insights into specific tissue patterning processes. Here, we provide for the first time a comprehensive analysis of developmentally regulated genes during murine limb development, and provide some novel insights into the expression dynamics governing limb morphogenesis. PMID:22174793

  4. Global gene expression analysis of murine limb development.

    PubMed

    Taher, Leila; Collette, Nicole M; Murugesh, Deepa; Maxwell, Evan; Ovcharenko, Ivan; Loots, Gabriela G

    2011-01-01

    Detailed information about stage-specific changes in gene expression is crucial for understanding the gene regulatory networks underlying development and the various signal transduction pathways contributing to morphogenesis. Here we describe the global gene expression dynamics during early murine limb development, when cartilage, tendons, muscle, joints, vasculature and nerves are specified and the musculoskeletal system of limbs is established. We used whole-genome microarrays to identify genes with differential expression at 5 stages of limb development (E9.5 to 13.5), during fore- and hind-limb patterning. We found that the onset of limb formation is characterized by an up-regulation of transcription factors, which is followed by a massive activation of genes during E10.5 and E11.5 which levels off at later time points. Among the 3520 genes identified as significantly up-regulated in the limb, we find ~30% to be novel, dramatically expanding the repertoire of candidate genes likely to function in the limb. Hierarchical and stage-specific clustering identified expression profiles that are likely to correlate with functional programs during limb development and further characterization of these transcripts will provide new insights into specific tissue patterning processes. Here, we provide for the first time a comprehensive analysis of developmentally regulated genes during murine limb development, and provide some novel insights into the expression dynamics governing limb morphogenesis.

  5. Ion channel gene expression predicts survival in glioma patients.

    PubMed

    Wang, Rong; Gurguis, Christopher I; Gu, Wanjun; Ko, Eun A; Lim, Inja; Bang, Hyoweon; Zhou, Tong; Ko, Jae-Hong

    2015-08-03

    Ion channels are important regulators in cell proliferation, migration, and apoptosis. The malfunction and/or aberrant expression of ion channels may disrupt these important biological processes and influence cancer progression. In this study, we investigate the expression pattern of ion channel genes in glioma. We designate 18 ion channel genes that are differentially expressed in high-grade glioma as a prognostic molecular signature. This ion channel gene expression based signature predicts glioma outcome in three independent validation cohorts. Interestingly, 16 of these 18 genes were down-regulated in high-grade glioma. This signature is independent of traditional clinical, molecular, and histological factors. Resampling tests indicate that the prognostic power of the signature outperforms random gene sets selected from human genome in all the validation cohorts. More importantly, this signature performs better than the random gene signatures selected from glioma-associated genes in two out of three validation datasets. This study implicates ion channels in brain cancer, thus expanding on knowledge of their roles in other cancers. Individualized profiling of ion channel gene expression serves as a superior and independent prognostic tool for glioma patients.

  6. Imprinted gene expression in fetal growth and development.

    PubMed

    Lambertini, L; Marsit, C J; Sharma, P; Maccani, M; Ma, Y; Hu, J; Chen, J

    2012-06-01

    Experimental studies showed that genomic imprinting is fundamental in fetoplacental development by timely regulating the expression of the imprinted genes to overlook a set of events determining placenta implantation, growth and embryogenesis. We examined the expression profile of 22 imprinted genes which have been linked to pregnancy abnormalities that may ultimately influence childhood development. The study was conducted in a subset of 106 placenta samples, overrepresented with small and large for gestational age cases, from the Rhode Island Child Health Study. We investigated associations between imprinted gene expression and three fetal development parameters: newborn head circumference, birth weight, and size for gestational age. Results from our investigation show that the maternally imprinted/paternally expressed gene ZNF331 inversely associates with each parameter to drive smaller fetal size, while paternally imprinted/maternally expressed gene SLC22A18 directly associates with the newborn head circumference promoting growth. Multidimensional Scaling analysis revealed two clusters within the 22 imprinted genes which are independently associated with fetoplacental development. Our data suggest that cluster 1 genes work by assuring cell growth and tissue development, while cluster 2 genes act by coordinating these processes. Results from this epidemiologic study offer solid support for the key role of imprinting in fetoplacental development.

  7. Differentially Expressed Genes and Signature Pathways of Human Prostate Cancer

    PubMed Central

    Myers, Jennifer S.; von Lersner, Ariana K.; Robbins, Charles J.; Sang, Qing-Xiang Amy

    2015-01-01

    Genomic technologies including microarrays and next-generation sequencing have enabled the generation of molecular signatures of prostate cancer. Lists of differentially expressed genes between malignant and non-malignant states are thought to be fertile sources of putative prostate cancer biomarkers. However such lists of differentially expressed genes can be highly variable for multiple reasons. As such, looking at differential expression in the context of gene sets and pathways has been more robust. Using next-generation genome sequencing data from The Cancer Genome Atlas, differential gene expression between age- and stage- matched human prostate tumors and non-malignant samples was assessed and used to craft a pathway signature of prostate cancer. Up- and down-regulated genes were assigned to pathways composed of curated groups of related genes from multiple databases. The significance of these pathways was then evaluated according to the number of differentially expressed genes found in the pathway and their position within the pathway using Gene Set Enrichment Analysis and Signaling Pathway Impact Analysis. The “transforming growth factor-beta signaling” and “Ran regulation of mitotic spindle formation” pathways were strongly associated with prostate cancer. Several other significant pathways confirm reported findings from microarray data that suggest actin cytoskeleton regulation, cell cycle, mitogen-activated protein kinase signaling, and calcium signaling are also altered in prostate cancer. Thus we have demonstrated feasibility of pathway analysis and identified an underexplored area (Ran) for investigation in prostate cancer pathogenesis. PMID:26683658

  8. Expression of the Arabidopsis Gene Akr Coincides with Chloroplast Development.

    PubMed

    Zhang, H.; Wang, J.; Goodman, H. M.

    1994-12-01

    Reduced expression of a nuclear gene of Arabidopsis thaliana, Akr, results in the formation of chlorotic plants due to a block in the proplastid-to-chloroplast development pathway (H. Zhang, D.C. Scheirer, W. Fowle, H.M. Goodman [1992] Plant Cell 4: 1575-1588). In an effort to discern the function of the Akr gene product in chloroplast development, transgenic plants containing an Akr::[beta]-glucuronidase gene fusion were constructed to monitor the spatial and temporal patterns of Akr expression. Akr is expressed only in chloroplast-containing tissues and maximal expression occurs during the seedling stage, coincident with chloroplast development. This result is consistent with the hypothesis that Akr is required at an early stage of chloroplast development. The effects of an AKR deficiency on the expression of nuclear and plastid genes required for photosynthetic activity were also examined. Within chloroplast-deficient leaves of plants in which Akr expression is limited by the presence of Akr antisense transgenes or truncated Akr sense transgenes, mRNAs for the nuclear genes Cab2, Cab4, RbcS, and GapA are present at wild-type levels; similarly, levels of mRNAs for the plastid genes rbcL and psbA are not affected by the AKR deficiency. Thus, although expression of these photosynthetic genes is tightly coordinated with the development and maintenance of chloroplasts in wild-type plants, their expression is unaffected in AKR-deficient chlorotic leaves. Therefore, we propose that Akr functions in a pathway different from the one controlling the expression and regulation of the photosynthetic genes during chloroplast development, and at a specific developmental stage after the putative plastid factor is made.

  9. Regulated expression of foreign genes in vivo after germline transfer.

    PubMed Central

    Passman, R S; Fishman, G I

    1994-01-01

    Tight transcriptional control of foreign genes introduced into the germline of transgenic mice would be of great experimental value in studies of gene function. To develop a system in which the spatial and temporal expression of candidate genes implicated in cardiac development or function could be tightly controlled in vivo, we have generated transgenic mice expressing a tetracycline-controlled transactivator (tTA) under the control of a rat alpha myosin heavy chain promoter (MHC alpha-tTA mice), as well as mice harboring a candidate target gene implicated in the control of differentiation, Id1 (tet-Id1 mice). No expression of the target transgene was detected in any tissues of hemizygous tet-Id1 mice. Genetic crosses with MHC alpha-tTA mice resulted in transactivation of the Id1 transgene, but expression was restricted to heart, where tTA was expressed. Furthermore, transactivation of the target gene was tightly and reversibly controlled by systemic therapy with tetracycline, both in utero and postnatally. These studies demonstrate the feasibility of such a binary approach for tightly controlling the timing and extent of expression of transgenes in vivo. This approach should be generally useful for the ectopic expression of candidate genes in selected tissues during delineated developmental stages. Images PMID:7989599

  10. Estradiol-induced gene expression in largemouth bass (Micropterus salmoides)

    USGS Publications Warehouse

    Bowman, C.J.; Kroll, K.J.; Gross, T.G.; Denslow, N.D.

    2002-01-01

    Vitellogenin (Vtg) and estrogen receptor (ER) gene expression levels were measured in largemouth bass to evaluate the activation of the ER-mediated pathway by estradiol (E2). Single injections of E2 ranging from 0.0005 to 5 mg/kg up-regulated plasma Vtg in a dose-dependent manner. Vtg and ER mRNAs were measured using partial cDNA sequences corresponding to the C-terminal domain for Vtg and the ligand-binding domain of ER?? sequences. After acute E2-exposures (2 mg/kg), Vtg and ER mRNAs and plasma Vtg levels peaked after 2 days. The rate of ER mRNA accumulation peaked 36-42 h earlier than Vtg mRNA. The expression window for ER defines the primary response to E2 in largemouth bass and that for Vtg a delayed primary response. The specific effect of E2 on other estrogen-regulated genes was tested during these same time windows using differential display RT-PCR. Specific up-regulated genes that are expressed in the same time window as Vtg were ERp72 (a membrane-bound disulfide isomerase) and a gene with homology to an expressed gene identified in zebrafish. Genes that were expressed in a pattern that mimics the ER include the gene for zona radiata protein ZP2, and a gene with homology to an expressed gene found in winter flounder. One gene for fibrinogen ?? was down-regulated and an unidentified gene was transiently up-regulated after 12 h of exposure and returned to basal levels by 48 h. Taken together these studies indicate that the acute molecular response to E2 involves a complex network of responses over time. ?? 2002 Elsevier Science Ireland Ltd. All rights reserved.

  11. Age-associated bidirectional modulation of gene expression in single identified R15 neuron of Aplysia

    PubMed Central

    2013-01-01

    Background Despite the advances in our understanding of aging-associated behavioral decline, relatively little is known about how aging affects neural circuits that regulate specific behaviors, particularly the expression of genes in specific neural circuits during aging. We have addressed this by exploring a peptidergic neuron R15, an identified neuron of the marine snail Aplysia californica. R15 is implicated in reproduction and osmoregulation and responds to neurotransmitters such as acetylcholine, serotonin and glutamate and is characterized by its action potential bursts. Results We examined changes in gene expression in R15 neurons during aging by microarray analyses of RNAs from two different age groups, mature and old animals. Specifically we find that 1083 ESTs are differentially regulated in mature and old R15 neurons. Bioinformatics analyses of these genes have identified specific biological pathways that are up or downregulated in mature and old neurons. Comparison with human signaling networks using pathway analyses have identified three major networks [(1) cell signaling, cell morphology, and skeletal muscular system development (2) cell death and survival, cellular function maintenance and embryonic development and (3) neurological diseases, developmental and hereditary disorders] altered in old R15 neurons. Furthermore, qPCR analysis of single R15 neurons to quantify expression levels of candidate regulators involved in transcription (CREB1) and translation (S6K) showed that aging is associated with a decrease in expression of these regulators, and similar analysis in three other neurons (L7, L11 and R2) showed that gene expression change during aging could be bidirectional. Conclusions We find that aging is associated with bidirectional changes in gene expression. Detailed bioinformatics analyses and human homolog searches have identified specific biological processes and human-relevant signaling pathways in R15 that are affected during aging

  12. Development of soybean gene expression database (SGED)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Large volumes of microarray expression data is a challenge for analysis. To address this problem a web-based database, Soybean Expression Database (SGED) was built, using PERL/CGI, C and an ORACLE database management system. SGED contains three components. The Data Mining component serves as a repos...

  13. Reconstruction of gene co-expression network from microarray data using local expression patterns

    PubMed Central

    2014-01-01

    Background Biological networks connect genes, gene products to one another. A network of co-regulated genes may form gene clusters that can encode proteins and take part in common biological processes. A gene co-expression network describes inter-relationships among genes. Existing techniques generally depend on proximity measures based on global similarity to draw the relationship between genes. It has been observed that expression profiles are sharing local similarity rather than global similarity. We propose an expression pattern based method called GeCON to extract Gene CO-expression Network from microarray data. Pair-wise supports are computed for each pair of genes based on changing tendencies and regulation patterns of the gene expression. Gene pairs showing negative or positive co-regulation under a given number of conditions are used to construct such gene co-expression network. We construct co-expression network with signed edges to reflect up- and down-regulation between pairs of genes. Most existing techniques do not emphasize computational efficiency. We exploit a fast correlogram matrix based technique for capturing the support of each gene pair to construct the network. Results We apply GeCON to both real and synthetic gene expression data. We compare our results using the DREAM (Dialogue for Reverse Engineering Assessments and Methods) Challenge data with three well known algorithms, viz., ARACNE, CLR and MRNET. Our method outperforms other algorithms based on in silico regulatory network reconstruction. Experimental results show that GeCON can extract functionally enriched network modules from real expression data. Conclusions In view of the results over several in-silico and real expression datasets, the proposed GeCON shows satisfactory performance in predicting co-expression network in a computationally inexpensive way. We further establish that a simple expression pattern matching is helpful in finding biologically relevant gene network. In

  14. Gene Expression Measurement Module (GEMM) - A Fully Automated, Miniaturized Instrument for Measuring Gene Expression in Space

    NASA Technical Reports Server (NTRS)

    Pohorille, Andrew; Peyvan, Kia; Karouia, Fathi; Ricco, Antonio

    2012-01-01

    The capability to measure gene expression on board spacecraft opens the door to a large number of high-value experiments on the influence of the space environment on biological systems. For example, measurements of gene expression will help us to understand adaptation of terrestrial life to conditions beyond the planet of origin, identify deleterious effects of the space environment on a wide range of organisms from microbes to humans, develop effective countermeasures against these effects, and determine the metabolic bases of microbial pathogenicity and drug resistance. These and other applications hold significant potential for discoveries in space biology, biotechnology, and medicine. Supported by funding from the NASA Astrobiology Science and Technology Instrument Development Program, we are developing a fully automated, miniaturized, integrated fluidic system for small spacecraft capable of in-situ measurement of expression of several hundreds of microbial genes from multiple samples. The instrument will be capable of (1) lysing cell walls of bacteria sampled from cultures grown in space, (2) extracting and purifying RNA released from cells, (3) hybridizing the RNA on a microarray and (4) providing readout of the microarray signal, all in a single microfluidics cartridge. The device is suitable for deployment on nanosatellite platforms developed by NASA Ames' Small Spacecraft Division. To meet space and other technical constraints imposed by these platforms, a number of technical innovations are being implemented. The integration and end-to-end technological and biological validation of the instrument are carried out using as a model the photosynthetic bacterium Synechococcus elongatus, known for its remarkable metabolic diversity and resilience to adverse conditions. Each step in the measurement process-lysis, nucleic acid extraction, purification, and hybridization to an array-is assessed through comparison of the results obtained using the instrument with

  15. The Longissimus and Semimembranosus Muscles Display Marked Differences in Their Gene Expression Profiles in Pig

    PubMed Central

    Herault, Frederic; Vincent, Annie; Dameron, Olivier; Le Roy, Pascale; Cherel, Pierre; Damon, Marie

    2014-01-01

    Background Meat quality depends on skeletal muscle structure and metabolic properties. While most studies carried on pigs focus on the Longissimus muscle (LM) for fresh meat consumption, Semimembranosus (SM) is also of interest because of its importance for cooked ham production. Even if both muscles are classified as glycolytic muscles, they exhibit dissimilar myofiber composition and metabolic characteristics. The comparison of LM and SM transcriptome profiles undertaken in this study may thus clarify the biological events underlying their phenotypic differences which might influence several meat quality traits. Methodology/Principal Findings Muscular transcriptome analyses were performed using a custom pig muscle microarray: the 15 K Genmascqchip. A total of 3823 genes were differentially expressed between the two muscles (Benjamini-Hochberg adjusted P value ≤0.05), out of which 1690 and 2133 were overrepresented in LM and SM respectively. The microarray data were validated using the expression level of seven differentially expressed genes quantified by real-time RT-PCR. A set of 1047 differentially expressed genes with a muscle fold change ratio above 1.5 was used for functional characterization. Functional annotation emphasized five main clusters associated to transcriptome muscle differences. These five clusters were related to energy metabolism, cell cycle, gene expression, anatomical structure development and signal transduction/immune response. Conclusions/Significance This study revealed strong transcriptome differences between LM and SM. These results suggest that skeletal muscle discrepancies might arise essentially from different post-natal myogenic activities. PMID:24809746

  16. mRNA expression profiles of calmodulin and liver receptor homolog-1 genes in chickens.

    PubMed

    Zhang, Z-C; Xiao, L-H; Wang, Y; Chen, S-Y; Yang, Z-Q; Zhao, X-L; Zhu, Q; Liu, Y-P

    2012-01-01

    Calmodulin (CALM), a calcium-binding protein, is expressed in the hypothalamic-pituitary-gonadal axis; it plays a pivotal role in the reproductive system by regulating gonadotropin-releasing hormone signaling. Downstream of hypothalamic-pituitary-gonadal signaling pathways, liver receptor homolog-1 (LRH-1) is involved in female gonadal hormone synthesis. In the chicken, although the two genes are known to be associated with reproductive traits, the interaction between gonadotropins and gonadal steroids remains unclear. We used quantitative real-time PCR to quantify the tissular (hypothalamus, pituitary, ovary, liver, kidney, oviduct, heart) and ontogenetic (12, 18, 32, and 45 weeks) mRNA expression profiles of CALM and LRH-1 in Erlang Mountainous chickens to determine their roles in the endocrine control of fertility, and compared these profiles with expression in Roman chickens. We found that the relative expressions of CALM and LRH-1 genes had the highest levels in the pituitary and ovary at 32 weeks. The expression level of CALM mRNA in the pituitary of Roman chickens was significantly higher than that in Erlang Mountainous chickens at 32 and 45 weeks, while the LRH-1 transcript level in the ovaries of Roman chickens was significantly lower than that of Erlang Mountainous chickens at 32 and 45 weeks. In summary, the transcript levels of CALM and LRH-1 genes are associated with chicken reproductive traits; in addition, we found that the CALM gene is the key regulator in the hypothalamic-pituitary-gonadal signaling network.

  17. Expression of LTP genes in response to saline stress in rice seedlings.

    PubMed

    Moraes, G P; Benitez, L C; do Amaral, M N; Vighi, I L; Auler, P A; da Maia, L C; Bianchi, V J; Braga, E J B

    2015-01-01

    Saline stress is one of the primary factors limiting increased rice productivity in the southern region of Brazil. Farming can be affected by salinity that is due to both the origin of the soils as well as the irrigation water. Lipid transfer proteins (LTPs) have many physiological functions, including in the response to saline stress. Therefore, the objective of this study was to quantify the relative expression of 11 genetic isoforms that encode LTP1-type proteins in rice genotypes tolerant and sensitive to saline stress in the vegetative period. When the plants reached development stage V4, alternating irrigation was started with nutritive solution and water containing 150 mM NaCl. The LTP7 gene showed an increase in expression by 13.81-fold after 96 h of stress exposure in the saline-tolerant group, whereas the LTP10 gene expression level was increased by 71.10-fold after 96 h in the saline-sensitive group. The LTP26, LTP23, and LTP18 genes showed increased expression in both genotypes; however, the expression levels and response times were different. Thus, LTP7 and LTP10 showed the highest response to salinity. The LTP18, LTP23, and LTP26 genes were negatively correlated with the response to salinity. PMID:26345756

  18. Selection and Validation of Appropriate Reference Genes for Quantitative Real-Time PCR Analysis of Gene Expression in Lycoris aurea

    PubMed Central

    Ma, Rui; Xu, Sheng; Zhao, Yucheng; Xia, Bing; Wang, Ren

    2016-01-01

    Lycoris aurea (L' Hér.) Herb, a perennial grass species, produces a unique variety of pharmacologically active Amaryllidaceae alkaloids. However, the key enzymes and their expression pattern involved in the biosynthesis of Amaryllidaceae alkaloids (especially for galanthamine) are far from being fully understood. Quantitative real-time polymerase chain reaction (qRT-PCR), a commonly used method for quantifying gene expression, requires stable reference genes to normalize its data. In this study, to choose the appropriate reference genes under different experimental conditions, 14 genes including YLS8 (mitosis protein YLS8), CYP2 (Cyclophilin 2), CYP 1 (Cyclophilin 1), TIP41 (TIP41-like protein), EXP2 (Expressed protein 2), PTBP1 (Polypyrimidine tract-binding protein 1), EXP1 (Expressed protein 1), PP2A (Serine/threonine-protein phosphatase 2A), β-TUB (β-tubulin), α-TUB (α-tubulin), EF1-α (Elongation factor 1-α), UBC (Ubiquitin-conjugating enzyme), ACT (Actin) and GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) were selected from the transcriptome datasets of L. aurea. And then, expressions of these genes were assessed by qRT-PCR in various tissues and the roots under different treatments. The expression stability of the 14 candidates was analyzed by three commonly used software programs (geNorm, NormFinder, and BestKeeper), and their results were further integrated into a comprehensive ranking based on the geometric mean. The results show the relatively stable genes for each subset as follows: (1) EXP1 and TIP41 for all samples; (2) UBC and EXP1 for NaCl stress; (3) PTBP1 and EXP1 for heat stress, polyethylene glycol (PEG) stress and ABA treatment; (4) UBC and CYP2 for cold stress; (5) PTBP1 and PP2A for sodium nitroprusside (SNP) treatment; (6) CYP1 and TIP41 for methyl jasmonate (MeJA) treatment; and (7) EXP1 and TIP41 for various tissues. The reliability of these results was further enhanced through comparison between part qRT-PCR result and RNA

  19. Pervasive Effects of Aging on Gene Expression in Wild Wolves.

    PubMed

    Charruau, Pauline; Johnston, Rachel A; Stahler, Daniel R; Lea, Amanda; Snyder-Mackler, Noah; Smith, Douglas W; vonHoldt, Bridgett M; Cole, Steven W; Tung, Jenny; Wayne, Robert K

    2016-08-01

    Gene expression levels change as an individual ages and responds to environmental conditions. With the exception of humans, such patterns have principally been studied under controlled conditions, overlooking the array of developmental and environmental influences that organisms encounter under conditions in which natural selection operates. We used high-throughput RNA sequencing (RNA-Seq) of whole blood to assess the relative impacts of social status, age, disease, and sex on gene expression levels in a natural population of gray wolves (Canis lupus). Our findings suggest that age is broadly associated with gene expression levels, whereas other examined factors have minimal effects on gene expression patterns. Further, our results reveal evolutionarily conserved signatures of senescence, such as immunosenescence and metabolic aging, between wolves and humans despite major differences in life history and environment. The effects of aging on gene expression levels in wolves exhibit conservation with humans, but the more rapid expression differences observed in aging wolves is evolutionarily appropriate given the species' high level of extrinsic mortality due to intraspecific aggression. Some expression changes that occur with age can facilitate physical age-related changes that may enhance fitness in older wolves. However, the expression of these ancestral patterns of aging in descendant modern dogs living in highly modified domestic environments may be maladaptive and cause disease. This work provides evolutionary insight into aging patterns observed in domestic dogs and demonstrates the applicability of studying natural populations to investigate the mechanisms of aging. PMID:27189566

  20. Expression of Th1/Th2/Th3/Th17-related genes in recurrent aphthous ulcers.

    PubMed

    Lewkowicz, Natalia; Kur, Barbara; Kurnatowska, Anna; Tchorzewski, Henryk; Lewkowicz, Przemyslaw

    2011-10-01

    The pathogenesis of recurrent aphthous ulceration (RAU) is unknown, although an abnormal immune reaction appears to be involved. RAU may result from oral epithelium damage caused by T cell-mediated immune response. To improve understanding of the role of T cells in RAU, the present study analyzed. the expression of T cell-related genes in oral ulcers from patients with RAU, as well as in healthy non-keratinized oral mucosa from aphthae-free volunteers. Biopsies from RAU patients and healthy individuals were analyzed using Human Th1-Th2-Th3 RT(2) Profiler PCR Array and qRT-PCR that allowed to quantify the transcript levels of 86 genes related to T cell activation. We found that cells present in aphthous ulcers express a characteristic Th1-like gene profile. The majority of genes up-regulated in aphthous lesions such as IFN-γ, TNF, IL-15, IRF1, STAT-1 and STAT-4 were Th1-associated. Th2-realated genes were not overexpressed in RAU tissues, with the exception for CCR3. Th3- and Th17-related gene expression patterns were not demonstrated in RAU. These findings clearly reveal that aphthous ulcer formation is predominantly dependent on the activation of the Th1-type immune response.

  1. Saponin determination, expression analysis and functional characterization of saponin biosynthetic genes in Chenopodium quinoa leaves.

    PubMed

    Fiallos-Jurado, Jennifer; Pollier, Jacob; Moses, Tessa; Arendt, Philipp; Barriga-Medina, Noelia; Morillo, Eduardo; Arahana, Venancio; de Lourdes Torres, Maria; Goossens, Alain; Leon-Reyes, Antonio

    2016-09-01

    Quinoa (Chenopodium quinoa Willd.) is a highly nutritious pseudocereal with an outstanding protein, vitamin, mineral and nutraceutical content. The leaves, flowers and seed coat of quinoa contain triterpenoid saponins, which impart bitterness to the grain and make them unpalatable without postharvest removal of the saponins. In this study, we quantified saponin content in quinoa leaves from Ecuadorian sweet and bitter genotypes and assessed the expression of saponin biosynthetic genes in leaf samples elicited with methyl jasmonate. We found saponin accumulation in leaves after MeJA treatment in both ecotypes tested. As no reference genes were available to perform qPCR in quinoa, we mined publicly available RNA-Seq data for orthologs of 22 genes known to be stably expressed in Arabidopsis thaliana using geNorm, NormFinder and BestKeeper algorithms. The quinoa ortholog of At2g28390 (Monensin Sensitivity 1, MON1) was stably expressed and chosen as a suitable reference gene for qPCR analysis. Candidate saponin biosynthesis genes were screened in the quinoa RNA-Seq data and subsequent functional characterization in yeast led to the identification of CqbAS1, CqCYP716A78 and CqCYP716A79. These genes were found to be induced by MeJA, suggesting this phytohormone might also modulate saponin biosynthesis in quinoa leaves. Knowledge of the saponin biosynthesis and its regulation in quinoa may aid the further development of sweet cultivars that do not require postharvest processing.

  2. Saponin determination, expression analysis and functional characterization of saponin biosynthetic genes in Chenopodium quinoa leaves.

    PubMed

    Fiallos-Jurado, Jennifer; Pollier, Jacob; Moses, Tessa; Arendt, Philipp; Barriga-Medina, Noelia; Morillo, Eduardo; Arahana, Venancio; de Lourdes Torres, Maria; Goossens, Alain; Leon-Reyes, Antonio

    2016-09-01

    Quinoa (Chenopodium quinoa Willd.) is a highly nutritious pseudocereal with an outstanding protein, vitamin, mineral and nutraceutical content. The leaves, flowers and seed coat of quinoa contain triterpenoid saponins, which impart bitterness to the grain and make them unpalatable without postharvest removal of the saponins. In this study, we quantified saponin content in quinoa leaves from Ecuadorian sweet and bitter genotypes and assessed the expression of saponin biosynthetic genes in leaf samples elicited with methyl jasmonate. We found saponin accumulation in leaves after MeJA treatment in both ecotypes tested. As no reference genes were available to perform qPCR in quinoa, we mined publicly available RNA-Seq data for orthologs of 22 genes known to be stably expressed in Arabidopsis thaliana using geNorm, NormFinder and BestKeeper algorithms. The quinoa ortholog of At2g28390 (Monensin Sensitivity 1, MON1) was stably expressed and chosen as a suitable reference gene for qPCR analysis. Candidate saponin biosynthesis genes were screened in the quinoa RNA-Seq data and subsequent functional characterization in yeast led to the identification of CqbAS1, CqCYP716A78 and CqCYP716A79. These genes were found to be induced by MeJA, suggesting this phytohormone might also modulate saponin biosynthesis in quinoa leaves. Knowledge of the saponin biosynthesis and its regulation in quinoa may aid the further development of sweet cultivars that do not require postharvest processing. PMID:27457995

  3. Expression of cellulase genes in Rhodobacter capsulatus by use of plasmid expression vectors.

    PubMed Central

    Johnson, J A; Wong, W K; Beatty, J T

    1986-01-01

    Broad-host-range plasmid vectors were constructed for expression of heterologous genes in the photosynthetic bacterium Rhodobacter capsulatus. These plasmids utilize an RK2-derived replicon for maintenance and conjugative transfer and the R. capsulatus rxcA promoter to obtain transcription of genes within appropriately positioned DNA fragments. The expression vectors were used to obtain synthesis of endoglucanase and exoglucanase in R. capsulatus from cellulase genes present on exogenously derived DNA fragments. The cellulase genes were expressed either by use of their native translation initiation signals or by in-frame fusion with the rxcA B870 beta gene translation initiation signals to form a hybrid protein. The level of cellulase gene expression was found to be modulated in response to the extent of aeration of plasmid host cultures. Images PMID:3090019

  4. Transcriptional expression study in the central nervous system of rats: what gene should be used as internal control?

    PubMed Central

    de Moura, Ana Carolina; Lazzari, Virgínia Meneghini; Agnes, Grasiela; Almeida, Silvana; Giovenardi, Márcia; da Veiga, Ana Beatriz Gorini

    2014-01-01

    Objective A growing number of published articles report the expression of specific genes with different behavior patterns in rats. The levels of messenger ribonucleic acid transcripts are usually analyzed by reverse transcription followed by polymerase chain reaction and quantified after normalization with an internal control or reference gene (housekeeping gene). Nevertheless, housekeeping genes exhibit different expression in the central nervous system, depending on the physiological conditions and the area of the brain to be studied. The choice of a good internal control gene is essential for obtaining reliable results. This study evaluated the expression of three housekeeping genes (beta-actin, cyclophilin A, and ubiquitin C) in different areas of the central nervous system in rats (olfactory bulb, hippocampus, striatum, and prefrontal cortex). Methods Wistar rats (virgin females, n=6) during the diestrum period were used. Total ribonucleic acid was extracted from each region of the brain; the complementary deoxyribonucleic acid was synthesized by reverse transcription and amplified by real-time quantitative polymerase chain reaction using SYBR™ Green and primers specific for each one of the reference genes. The stability of the expression was determined using NormFinder. Results Beta-actin was the most stable gene in the hippocampus and striatum, while cyclophilin A and ubiquitin C showed greater stability in the prefrontal cortex and the olfactory bulb, respectively. Conclusion Based on our study, further studies of gene expression using rats as animal models should take into consideration these results when choosing a reliable internal control gene. PMID:25295456

  5. Use of reporter-gene based bacteria to quantify phenanthrene biodegradation and toxicity in soil.

    PubMed

    Shin, Doyun; Moon, Hee Sun; Lin, Chu-Ching; Barkay, Tamar; Nam, Kyoungphile

    2011-02-01

    A phenanthrene-degrading bacterium, Sphingomonas paucimobilis EPA505 was used to construct two fluorescence-based reporter strains. Strain D harboring gfp gene was constructed to generate green fluorescence when the strain started to biodegrade phenanthrene. Strain S possessing gef gene was designed to die once phenanthrene biodegradation was initiated and thus to lose green fluorescence when visualized by a live/dead cell staining. Confocal laser scanning microscopic observation followed by image analysis demonstrates that the fluorescence intensity generated by strain D increased and the intensity by strain S decreased linearly at the phenanthrene concentration of up to 200 mg/L. Such quantitative increase and decrease of fluorescence intensity in strain D (i.e., from 1 to 11.90 ± 0.72) and strain S (from 1 to 0.40 ± 0.07) were also evident in the presence of Ottawa sand spiked with the phenanthrene up to 1000 mg/kg. The potential use of the reporter strains in quantitatively determining biodegradable or toxic phenanthrene was discussed.

  6. Multiscale Embedded Gene Co-expression Network Analysis

    PubMed Central

    Song, Won-Min; Zhang, Bin

    2015-01-01

    Gene co-expression network analysis has been shown effective in identifying functional co-expressed gene modules associated with complex human diseases. However, existing techniques to construct co-expression networks require some critical prior information such as predefined number of clusters, numerical thresholds for defining co-expression/interaction, or do not naturally reproduce the hallmarks of complex systems such as the scale-free degree distribution of small-worldness. Previously, a graph filtering technique called Planar Maximally Filtered Graph (PMFG) has been applied to many real-world data sets such as financial stock prices and gene expression to extract meaningful and relevant interactions. However, PMFG is not suitable for large-scale genomic data due to several drawbacks, such as the high computation complexity O(|V|3), the presence of false-positives due to the maximal planarity constraint, and the inadequacy of the clustering framework. Here, we developed a new co-expression network analysis framework called Multiscale Embedded Gene Co-expression Network Analysis (MEGENA) by: i) introducing quality control of co-expression similarities, ii) parallelizing embedded network construction, and iii) developing a novel clustering technique to identify multi-scale clustering structures in Planar Filtered Networks (PFNs). We applied MEGENA to a series of simulated data and the gene expression data in breast carcinoma and lung adenocarcinoma from The Cancer Genome Atlas (TCGA). MEGENA showed improved performance over well-established clustering methods and co-expression network construction approaches. MEGENA revealed not only meaningful multi-scale organizations of co-expressed gene clusters but also novel targets in breast carcinoma and lung adenocarcinoma. PMID:26618778

  7. DAWN: a framework to identify autism genes and subnetworks using gene expression and genetics

    PubMed Central

    2014-01-01

    Background De novo loss-of-function (dnLoF) mutations are found twofold more often in autism spectrum disorder (ASD) probands than their unaffected siblings. Multiple independent dnLoF mutations in the same gene implicate the gene in risk and hence provide a systematic, albeit arduous, path forward for ASD genetics. It is likely that using additional non-genetic data will enhance the ability to identify ASD genes. Methods To accelerate the search for ASD genes, we developed a novel algorithm, DAWN, to model two kinds of data: rare variations from exome sequencing and gene co-expression in the mid-fetal prefrontal and motor-somatosensory neocortex, a critical nexus for risk. The algorithm casts the ensemble data as a hidden Markov random field in which the graph structure is determined by gene co-expression and it combines these interrelationships with node-specific observations, namely gene identity, expression, genetic data and the estimated effect on risk. Results Using currently available genetic data and a specific developmental time period for gene co-expression, DAWN identified 127 genes that plausibly affect risk, and a set of likely ASD subnetworks. Validation experiments making use of published targeted resequencing results demonstrate its efficacy in reliably predicting ASD genes. DAWN also successfully predicts known ASD genes, not included in the genetic data used to create the model. Conclusions Validation studies demonstrate that DAWN is effective in predicting ASD genes and subnetworks by leveraging genetic and gene expression data. The findings reported here implicate neurite extension and neuronal arborization as risks for ASD. Using DAWN on emerging ASD sequence data and gene expression data from other brain regions and tissues would likely identify novel ASD genes. DAWN can also be used for other complex disorders to identify genes and subnetworks in those disorders. PMID:24602502

  8. Peripheral blood collection: the first step towards gene expression profiling.

    PubMed

    Franken, Carmen; Remy, Sylvie; Lambrechts, Nathalie; Hollanders, Karen; Den Hond, Elly; Schoeters, Greet

    2016-07-01

    A crucial challenge for gene expression analysis in human biomonitoring studies on whole blood samples is rapid sample handling and mRNA stabilization. This study was designed to evaluate the impact of short bench times (less than 30 min) on yield, quality and gene expression of mRNA in the presence of different stabilization buffers (Tempus(TM) Blood RNA tube and RNAlater(®) Stabilization Reagent). Microarray analyzes showed significant changes over short periods of time in expression of a considerate part of the transcriptome (2356 genes) with a prominent role for NFкB-, cancer- and glucocorticoid-mediated networks, and specifically interleukin-8 (IL-8). These findings suggest that even short bench times affect gene expression, requiring to carry out blood collection in a strictly standardized way. PMID:26984061

  9. GENE EXPRESSION PROFILING TO IDENTIFY BIOMARKERS OF REPRODUCTIVE TOXICITY

    EPA Science Inventory

    SOT 2005 SESSION ABSTRACT

    GENE EXPRESSION PROFILING TO IDENTIFY BIOMARKERS OF REPRODUCTIVE TOXICITY

    David J. Dix. National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Research Triangle...

  10. Visually Relating Gene Expression and in vivo DNA Binding Data

    SciTech Connect

    Huang, Min-Yu; Mackey, Lester; Ker?,; nen, Soile V. E.; Weber, Gunther H.; Jordan, Michael I.; Knowles, David W.; Biggin, Mark D.; Hamann, Bernd

    2011-09-20

    Gene expression and in vivo DNA binding data provide important information for understanding gene regulatory networks: in vivo DNA binding data indicate genomic regions where transcription factors are bound, and expression data show the output resulting from this binding. Thus, there must be functional relationships between these two types of data. While visualization and data analysis tools exist for each data type alone, there is a lack of tools that can easily explore the relationship between them. We propose an approach that uses the average expression driven by multiple of ciscontrol regions to visually relate gene expression and in vivo DNA binding data. We demonstrate the utility of this tool with examples from the network controlling early Drosophila development. The results obtained support the idea that the level of occupancy of a transcription factor on DNA strongly determines the degree to which the factor regulates a target gene, and in some cases also controls whether the regulation is positive or negative.

  11. Gene expression defines natural changes in mammalian lifespan

    PubMed Central

    Fushan, Alexey A; Turanov, Anton A; Lee, Sang-Goo; Kim, Eun Bae; Lobanov, Alexei V; Yim, Sun Hee; Buffenstein, Rochelle; Lee, Sang-Rae; Chang, Kyu-Tae; Rhee, Hwanseok; Kim, Jong-So; Yang, Kap-Seok; Gladyshev, Vadim N

    2015-01-01

    Mammals differ more than 100-fold in maximum lifespan, which can be altered in either direction during evolution, but the molecular basis for natural changes in longevity is not understood. Divergent evolution of mammals also led to extensive changes in gene expression within and between lineages. To understand the relationship between lifespan and variation in gene expression, we carried out RNA-seq-based gene expression analyses of liver, kidney, and brain of 33 diverse species of mammals. Our analysis uncovered parallel evolution of gene expression and lifespan, as well as the associated life-history traits, and identified the processes and pathways involved. These findings provide direct insights into how nature reversibly adjusts lifespan and other traits during adaptive radiation of lineages. PMID:25677554

  12. On TADs and LADs: Spatial Control Over Gene Expression.

    PubMed

    Gonzalez-Sandoval, Adriana; Gasser, Susan M

    2016-08-01

    The combinatorial action of transcription factors drives cell-type-specific gene expression patterns. However, transcription factor binding and gene regulation occur in the context of chromatin, which modulates DNA accessibility. High-resolution chromatin interaction maps have defined units of chromatin that are in spatial proximity, called topologically associated domains (TADs). TADs can be further classified based on expression activity, replication timing, or the histone marks or non-histone proteins associated with them. Independently, other chromatin domains have been defined by their likelihood to interact with non-DNA structures, such as the nuclear lamina. Lamina-associated domains (LADs) correlate with low gene expression and late replication timing. TADs and LADs have recently been evaluated with respect to cell-type-specific gene expression. The results shed light on the relevance of these forms of chromatin organization for transcriptional regulation, and address specifically how chromatin sequestration influences cell fate decisions during organismal development. PMID:27312344

  13. Super-paramagnetic clustering of yeast gene expression profiles

    NASA Astrophysics Data System (ADS)

    Getz, G.; Levine, E.; Domany, E.; Zhang, M. Q.

    2000-04-01

    High-density DNA arrays, used to monitor gene expression at a genomic scale, have produced vast amounts of information which require the development of efficient computational methods to analyze them. The important first step is to extract the fundamental patterns of gene expression inherent in the data. This paper describes the application of a novel clustering algorithm, super-paramagnetic clustering (SPC) to analysis of gene expression profiles that were generated recently during a study of the yeast cell cycle. SPC was used to organize genes into biologically relevant clusters that are suggestive for their co-regulation. Some of the advantages of SPC are its robustness against noise and initialization, a clear signature of cluster formation and splitting, and an unsupervised self-organized determination of the number of clusters at each resolution. Our analysis revealed interesting correlated behavior of several groups of genes which has not been previously identified.

  14. Expression of streptavidin gene in bacteria and plants

    SciTech Connect

    Guan, Xueni; Wurtele, E.S.; Nikolau, B.J. )

    1990-05-01

    Six biotin-containing proteins are present in plants, representing at least four different biotin enzymes. The physiological function of these biotin enzymes is not understood. Streptavidin, a protein from Streptomyces avidinii, binds tightly and specifically to biotin causing inactivation of biotin enzymes. One approach to elucidating the physiological function of biotin enzymes in plant metabolism is to create transgenic plants expressing the streptavidin gene. A plasmid containing a fused streptavidin-beta-galactosidase gene has been expressed in E. coli. We also have constructed various fusion genes that include an altered CaMV 35S promoter, signal peptides to target the streptavidin protein to specific organelles, and the streptavidin coding gene. We are examining the expression of these genes in cells of carrot.

  15. Gene Expression Profiling of Soft and Firm Atlantic Salmon Fillet

    PubMed Central

    Larsson, Thomas; Mørkøre, Turid; Kolstad, Kari; Østbye, Tone-Kari; Afanasyev, Sergey; Krasnov, Aleksei

    2012-01-01

    Texture of salmon fillets is an important quality trait for consumer acceptance as well as for the suitability for processing. In the present work we measured fillet firmness in a population of farmed Atlantic salmon with known pedigree and investigated the relationship between this trait and gene expression. Transcriptomic analyses performed with a 21 K oligonucleotide microarray revealed strong correlations between firmness and a large number of genes. Highly similar expression profiles were observed in several functional groups. Positive regression was found between firmness and genes encoding proteasome components (41 genes) and mitochondrial proteins (129 genes), proteins involved in stress responses (12 genes), and lipid metabolism (30 genes). Coefficients of determination (R2) were in the range of 0.64–0.74. A weaker though highly significant negative regression was seen in sugar metabolism (26 genes, R2 = 0.66) and myofiber proteins (42 genes, R2 = 0.54). Among individual genes that showed a strong association with firmness, there were extracellular matrix proteins (negative correlation), immune genes, and intracellular proteases (positive correlation). Several genes can be regarded as candidate markers of flesh quality (coiled-coil transcriptional coactivator b, AMP deaminase 3, and oligopeptide transporter 15) though their functional roles are unclear. To conclude, fillet firmness of Atlantic salmon depends largely on metabolic properties of the skeletal muscle; where aerobic metabolism using lipids as fuel, and the rapid removal of damaged proteins, appear to play a major role. PMID:22745718

  16. Rotation gene set testing for longitudinal expression data.

    PubMed

    Dørum, Guro; Snipen, Lars; Solheim, Margrete; Saebø, Solve

    2014-11-01

    Gene set analysis methods are popular tools for identifying differentially expressed gene sets in microarray data. Most existing methods use a permutation test to assess significance for each gene set. The permutation test's assumption of exchangeable samples is often not satisfied for time-series data and complex experimental designs, and in addition it requires a certain number of samples to compute p-values accurately. The method presented here uses a rotation test rather than a permutation test to assess significance. The rotation test can compute accurate p-values also for very small sample sizes. The method can handle complex designs and is particularly suited for longitudinal microarray data where the samples may have complex correlation structures. Dependencies between genes, modeled with the use of gene networks, are incorporated in the estimation of correlations between samples. In addition, the method can test for both gene sets that are differentially expressed and gene sets that show strong time trends. We show on simulated longitudinal data that the ability to identify important gene sets may be improved by taking the correlation structure between samples into account. Applied to real data, the method identifies both gene sets with constant expression and gene sets with strong time trends.

  17. Expression of a human placental alkaline phosphatase gene in transfected cells: Use as a reporter for studies of gene expression

    SciTech Connect

    Henthorn, P.; Zervos, P.; Raducha, M.; Harris, H.; Kadesch, T.

    1988-09-01

    The human placental alkaline phosphatase gene has been cloned and reintroduced into mammalian cells. When a plasmid carrying the gene under control of the simian virus 40 early promoter (pSV2Apap) is transfected into a variety of different cell types, placental alkaline phosphatase activity can readily be detected by using whole cell suspensions or cell lysates. Alkaline phosphatase activity can also be visualized directly in individual transfected cells by histochemical staining. The gene is appropriate for use as a reporter in studies of gene regulation since its expression is dependent on the presence of exogenous transcription control elements. The overall assay to detect the expression of the gene is quantitative, very rapid, and inexpensive. Cotransfections of cells with pSV2Apap and a related plasmid carrying the bacterial chloramphenicol acetyltransferase gene (pSV2Acat) indicate that transcription of these two genes is detected with roughly the same sensitivity.

  18. Equivalent Gene Expression Profiles between Glatopa™ and Copaxone®

    PubMed Central

    D’Alessandro, Josephine S.; Duffner, Jay; Pradines, Joel; Capila, Ishan; Garofalo, Kevin; Kaundinya, Ganesh; Greenberg, Benjamin M.; Kantor, Daniel; Ganguly, Tanmoy C.

    2015-01-01

    Glatopa™ is a generic glatiramer acetate recently approved for the treatment of patients with relapsing forms of multiple sclerosis. Gene expression profiling was performed as a means to evaluate equivalence of Glatopa and Copaxone®. Microarray analysis containing 39,429 unique probes across the entire genome was performed in murine glatiramer acetate—responsive Th2-polarized T cells, a test system highly relevant to the biology of glatiramer acetate. A closely related but nonequivalent glatiramoid molecule was used as a control to establish assay sensitivity. Multiple probe-level (Student’s t-test) and sample-level (principal component analysis, multidimensional scaling, and hierarchical clustering) statistical analyses were utilized to look for differences in gene expression induced by the test articles. The analyses were conducted across all genes measured, as well as across a subset of genes that were shown to be modulated by Copaxone. The following observations were made across multiple statistical analyses: the expression of numerous genes was significantly changed by treatment with Copaxone when compared against media-only control; gene expression profiles induced by Copaxone and Glatopa were not significantly different; and gene expression profiles induced by Copaxone and the nonequivalent glatiramoid were significantly different, underscoring the sensitivity of the test system and the multiple analysis methods. Comparative analysis was also performed on sets of transcripts relevant to T-cell biology and antigen presentation, among others that are known to be modulated by glatiramer acetate. No statistically significant differences were observed between Copaxone and Glatopa in the expression levels (magnitude and direction) of these glatiramer acetate-regulated genes. In conclusion, multiple methods consistently supported equivalent gene expression profiles between Copaxone and Glatopa. PMID:26473741

  19. Equivalent Gene Expression Profiles between Glatopa™ and Copaxone®.

    PubMed

    D'Alessandro, Josephine S; Duffner, Jay; Pradines, Joel; Capila, Ishan; Garofalo, Kevin; Kaundinya, Ganesh; Greenberg, Benjamin M; Kantor, Daniel; Ganguly, Tanmoy C

    2015-01-01

    Glatopa™ is a generic glatiramer acetate recently approved for the treatment of patients with relapsing forms of multiple sclerosis. Gene expression profiling was performed as a means to evaluate equivalence of Glatopa and Copaxone®. Microarray analysis containing 39,429 unique probes across the entire genome was performed in murine glatiramer acetate--responsive Th2-polarized T cells, a test system highly relevant to the biology of glatiramer acetate. A closely related but nonequivalent glatiramoid molecule was used as a control to establish assay sensitivity. Multiple probe-level (Student's t-test) and sample-level (principal component analysis, multidimensional scaling, and hierarchical clustering) statistical analyses were utilized to look for differences in gene expression induced by the test articles. The analyses were conducted across all genes measured, as well as across a subset of genes that were shown to be modulated by Copaxone. The following observations were made across multiple statistical analyses: the expression of numerous genes was significantly changed by treatment with Copaxone when compared against media-only control; gene expression profiles induced by Copaxone and Glatopa were not significantly different; and gene expression profiles induced by Copaxone and the nonequivalent glatiramoid were significantly different, underscoring the sensitivity of the test system and the multiple analysis methods. Comparative analysis was also performed on sets of transcripts relevant to T-cell biology and antigen presentation, among others that are known to be modulated by glatiramer acetate. No statistically significant differences were observed between Copaxone and Glatopa in the expression levels (magnitude and direction) of these glatiramer acetate-regulated genes. In conclusion, multiple methods consistently supported equivalent gene expression profiles between Copaxone and Glatopa. PMID:26473741

  20. Single Cell Quantification of Reporter Gene Expression in Live Adult Caenorhabditis elegans Reveals Reproducible Cell-Specific Expression Patterns and Underlying Biological Variation.

    PubMed

    Mendenhall, Alexander R; Tedesco, Patricia M; Sands, Bryan; Johnson, Thomas E; Brent, Roger

    2015-01-01

    In multicellular organisms such as Caenorhabditis elegans, differences in complex phenotypes such as lifespan correlate with the level of expression of particular engineered reporter genes. In single celled organisms, quantitative understanding of responses to extracellular signals and of cell-to-cell variation in responses has depended on precise measurement of reporter gene expression. Here, we developed microscope-based methods to quantify reporter gene expression in cells of Caenorhabditis elegans with low measurement error. We then quantified expression in strains that carried different configurations of Phsp-16.2-fluorescent-protein reporters, in whole animals, and in all 20 cells of the intestine tissue, which is responsible for most of the fluorescent signal. Some animals bore more recently developed single copy Phsp-16.2 reporters integrated at defined chromosomal sites, others, "classical" multicopy reporter gene arrays integrated at random sites. At the level of whole animals, variation in gene expression was similar: strains with single copy reporters showed the same amount of animal-to-animal variation as strains with multicopy reporters. At the level of cells, in animals with single copy reporters, the pattern of expression in cells within the tissue was highly stereotyped. In animals with multicopy reporters, the cell-specific expression pattern was also stereotyped, but distinct, and somewhat more variable. Our methods are rapid and gentle enough to allow quantification of expression in the same cells of an animal at different times during adult life. They should allow investigators to use changes in reporter expression in single cells in tissues as quantitative phenotypes, and link those to molecular differences. Moreover, by diminishing measurement error, they should make possible dissection of the causes of the remaining, real, variation in expression. Understanding such variation should help reveal its contribution to differences in complex

  1. On the robustness of complex heterogeneous gene expression networks.

    PubMed

    Gómez-Gardeñes, Jesús; Moreno, Yamir; Floría, Luis M

    2005-04-01

    We analyze a continuous gene expression model on the underlying topology of a complex heterogeneous network. Numerical simulations aimed at studying the chaotic and periodic dynamics of the model are performed. The results clearly indicate that there is a region in which the dynamical and structural complexity of the system avoid chaotic attractors. However, contrary to what has been reported for Random Boolean Networks, the chaotic phase cannot be completely suppressed, which has important bearings on network robustness and gene expression modeling.

  2. Freedom of expression: cell-type-specific gene profiling.

    PubMed

    Otsuki, Leo; Cheetham, Seth W; Brand, Andrea H

    2014-01-01

    Cell fate and behavior are results of differential gene regulation, making techniques to profile gene expression in specific cell types highly desirable. Many methods now enable investigation at the DNA, RNA and protein level. This review introduces the most recent and popular techniques, and discusses key issues influencing the choice between these such as ease, cost and applicability of information gained. Interdisciplinary collaborations will no doubt contribute further advances, including not just in single cell type but single-cell expression profiling.

  3. Strong Magnetic Field Induced Changes of Gene Expression in Arabidopsis

    NASA Astrophysics Data System (ADS)

    Paul, A.-L.; Ferl, R. J.; Klingenberg, B.; Brooks, J. S.; Morgan, A. N.; Yowtak, J.; Meisel, M. W.

    2005-07-01

    We review our studies of the biological impact of magnetic field strengths of up to 30 T on transgenic arabidopsis plants engineered with a stress response gene consisting of the alcohol dehydrogenase (Adh) gene promoter driving the β-glucuronidase (GUS) gene reporter. Field strengths in excess of 15 T induce expression of the Adh/GUS transgene in the roots and leaves. Microarray analyses indicate that such field strengths have a far reaching effect on the genome. Wide spread induction of stress-related genes and transcription factors, and a depression of genes associated with cell wall metabolism are prominent examples.

  4. Caffeine exposure alters cardiac gene expression in embryonic cardiomyocytes.

    PubMed

    Fang, Xiefan; Mei, Wenbin; Barbazuk, William B; Rivkees, Scott A; Wendler, Christopher C

    2014-12-15

    Previous studies demonstrated that in utero caffeine treatment at embryonic day (E) 8.5 alters DNA methylation patterns, gene expression, and cardiac function in adult mice. To provide insight into the mechanisms, we examined cardiac gene and microRNA (miRNA) expression in cardiomyocytes shortly after exposure to physiologically relevant doses of caffeine. In HL-1 and primary embryonic cardiomyocytes, caffeine treatment for 48 h significantly altered the expression of cardiac structural genes (Myh6, Myh7, Myh7b, Tnni3), hormonal genes (Anp and BnP), cardiac transcription factors (Gata4, Mef2c, Mef2d, Nfatc1), and microRNAs (miRNAs; miR208a, miR208b, miR499). In addition, expressions of these genes were significantly altered in embryonic hearts exposed to in utero caffeine. For in utero experiments, pregnant CD-1 dams were treated with 20-60 mg/kg of caffeine, which resulted in maternal circulation levels of 37.3-65.3 μM 2 h after treatment. RNA sequencing was performed on embryonic ventricles treated with vehicle or 20 mg/kg of caffeine daily from E6.5-9.5. Differential expression (DE) analysis revealed that 124 genes and 849 transcripts were significantly altered, and differential exon usage (DEU) analysis identified 597 exons that were changed in response to prenatal caffeine exposure. Among the DE genes identified by RNA sequencing were several cardiac structural genes and genes that control DNA methylation and histone modification. Pathway analysis revealed that pathways related to cardiovascular development and diseases were significantly affected by caffeine. In addition, global cardiac DNA methylation was reduced in caffeine-treated cardiomyocytes. Collectively, these data demonstrate that caffeine exposure alters gene expression and DNA methylation in embryonic cardiomyocytes.

  5. Gene Expression Profiling in the Hibernating Primate, Cheirogaleus Medius

    PubMed Central

    Faherty, Sheena L.; Villanueva-Cañas, José Luis; Klopfer, Peter H.; Albà, M. Mar; Yoder, Anne D.

    2016-01-01

    Hibernation is a complex physiological response that some mammalian species employ to evade energetic demands. Previous work in mammalian hibernators suggests that hibernation is activated not by a set of genes unique to hibernators, but by differential expression of genes that are present in all mammals. This question of universal genetic mechanisms requires further investigation and can only be tested through additional investigations of phylogenetically dispersed species. To explore this question, we use RNA-Seq to investigate gene expression dynamics as they relate to the varying physiological states experienced throughout the year in a group of primate hibernators—Madagascar’s dwarf lemurs (genus Cheirogaleus). In a novel experimental approach, we use longitudinal sampling of biological tissues as a method for capturing gene expression profiles from the same individuals throughout their annual hibernation cycle. We identify 90 candidate genes that have variable expression patterns when comparing two active states (Active 1 and Active 2) with a torpor state. These include genes that are involved in metabolic pathways, feeding behavior, and circadian rhythms, as might be expected to correlate with seasonal physiological state changes. The identified genes appear to be critical for maintaining the health of an animal that undergoes prolonged periods of metabolic depression concurrent with the hibernation phenotype. By focusing on these differentially expressed genes in dwarf lemurs, we compare gene expression patterns in previously studied mammalian hibernators. Additionally, by employing evolutionary rate analysis, we find that hibernation-related genes do not evolve under positive selection in hibernating species relative to nonhibernators. PMID:27412611

  6. [Gene expression profile of spinal ventral horn in ALS].

    PubMed

    Yamamoto, Masahiko; Tanaka, Fumiaki; Sobue, Gen

    2007-10-01

    The causative pathomechanism of sporadic amyotrophic lateral sclerosis (ALS) is not clearly understood. Using microarray technology combined with laser-captured microdissection, gene expression profiles of degenerating spinal motor neurons as well as spinal ventral horn from autopsied patients with sporadic ALS were examined. Spinal motor neurons showed a distinct gene expression profile from the whole spinal ventral horn. Three percent of genes examined were significantly downregulated, and 1% were upregulated in motor neurons. In contrast with motor neurons, the total spinal ventral horn homogenates demonstrated 0.7% and 0.2% significant upregulation and downregulation of gene expression, respectively. Downregulated genes in motor neurons included those associated with cytoskeleton/axonal transport, transcription and cell surface antigens/receptors, such as dynactin 1 (DCTN1) and early growth response 3 (EGR3). In particular, DCTN1 was markedly downregulated in most residual motor neurons prior to the accumulation of pNF-H and ubiquitylated protein. Promoters for cell death pathway, death receptor 5 (DR5), cyclins C (CCNC) and A1 (CCNA), and caspases were upregulated, whereas cell death inhibitors,