Effect of high fat diet feeding on gene expression
Subtle metabolic and liver gene transcriptional changes underlie diet-induced fatty liver susceptibility in insulin-resistant mice.
Sex, Age, Specimen part, Subject
View SamplesBackground: RNA-seq is revolutionizing the way we study transcriptomes. mRNA can be surveyed without prior knowledge of gene transcripts. Alternative splicing of transcript isoforms and the identification of previously unknown exons are being reported. Initial reports of differences in exon usage, and splicing between samples as well as quantitative differences among samples are beginning to surface. Biological variation has been reported to be larger than technical variation. In addition, technical variation has been reported to be in line with expectations due to random sampling. However, strategies for dealing with technical variation will differ depending on the magnitude. The size of technical variance, and the role of sampling are examined in this manuscript. Results: Independent Solexa/Illumina experiments containing technical replicates are analyzed. When coverage is low, large disagreements between technical replicates are apparent. Exon detection between technical replicates is highly variable when the coverage is less than 5 reads per nucleotide and estimates of gene expression are more likely to disagree when coverage is low. Although large disagreements in the estimates of expression are observed at all levels of coverage. Conclusions: Technical variability is too high to ignore. Technical variability results in inconsistent detection of exons at low levels of coverage. Further, the estimate of the relative abundance of a transcript can substantially disagree, even when coverage levels are high. This may be due to the low sampling fraction and if so, it will persist as an issue needing to be addressed in experimental design even as the next wave of technology produces larger numbers of reads. We provide practical recommendations for dealing with the technical variability, without dramatic cost increases. Overall design: Three independent samples of D. melanogaster female heads were collected with each sample representing a unique pool of biological material. Each sample was prepared according to manufacturer's instructions and then the same library was run on two lanes of a Solexa/Illumina flow cell, resulting in two technical replicates for each biological replicate, runs were 36 base-pair paired end.
A flexible Bayesian method for detecting allelic imbalance in RNA-seq data.
Sex, Specimen part, Cell line, Subject
View Samples99 individual ovarian tumors (37 endometrioid, 41 serous, 13 mucinous, and 8 clear cell carcinomas) and 4 individual normal ovary samples, each assayed on an Affymetrix HG_U133A array
Fibroblast growth factor 9 has oncogenic activity and is a downstream target of Wnt signaling in ovarian endometrioid adenocarcinomas.
Disease stage
View SamplesFifty million plaque-forming units of AdCre was injected into the right ovarian bursal cavity of 56- 70 day old female mice. Mice were euthanized 63 days later to obtain ovary tumors and normal ovary tissue. Seven individual ovarian tumors and 4 individual normal ovary samples were each assayed on an Affymetrix Mouse Genome 430 2.0 array.
Fibroblast growth factor 9 has oncogenic activity and is a downstream target of Wnt signaling in ovarian endometrioid adenocarcinomas.
Sex
View SamplesDrosophila melanogaster adult males perform an elaborate courtship ritual to entice females to mate. fruitless (fru), a gene that is one of the key regulators of male courtship behavior, encodes multiple male-specific isoforms (FruM). These isoforms vary in their carboxy-terminal zinc finger domains, which are predicted to facilitate DNA binding. By over-expressing individual FruM isoforms in fru-expressing neurons in either males or females and assaying the global transcriptional response by RNA-sequencing, we show that three FruM isoforms have different regulatory activities that depend on the sex of the fly. We identified several sets of genes regulated downstream of FruM isoforms. Overall design: RNA seqeuncing was performed on mRNA derived from adult male or female heads, for a total of 39 samples. These samples included two wild type genotypes (Berlin and Canton-S), two transheterozygous mutants for fru P1 (Df(3R)P14/Df(3R)fru4-40 and fruw12/ Df(3R)ChaM5), and 3 overexpressing genotypes (fru P1-Gal4: UAS-FruMA, UAS-FruMB, UAS-FruMC). There were at least 3 replicates from biological samples for all sex by genotype combinations.
Sex Differences in Drosophila Somatic Gene Expression: Variation and Regulation by doublesex.
Sex, Specimen part, Cell line, Subject
View SamplesThe Drosophila sex determination hierarchy consists of a splicing cascade with sex-specific transcription directing somatic sexual dimorphism. Our understanding of this pathway, and many others, is incomplete. Here we pioneer an approach to expand our knowledge of gene regulatory networks (GRNs) by leveraging natural genetic variation. This approach is generalizable to any natural population, including humans. Two studies from Drosophila female head tissue were used – the DSPR collection (alleles from 15 natural variants) and F1-hybrid collection (alleles from heterozygotes of 75 isogenic lines crossed to w1118) – in a structural equation model (SEM) analysis. We expanded the sex hierarchy GRN by adding novel links among genes in the pathway and by adding novel genes to the pathway. A link from fruitless (fru) to Sex-lethal (Sxl) was found in both populations, which is supported by the presence of fru binding sites in the Sxl locus. The splicing factors male-specific lethal 2 and Rm62 were correctly identified as downstream targets of Sxl. There were 754 additional candidate genes for an expanded sex hierarchy GRN. These candidates were enriched for genes with sex-biased splicing and many components of the spliceosome were placed in the GRN. As with other population-genetic analyses, the number of alleles limits the number of observable interactions. Network expansion was only clear in the F1-hybrid population, which has an average of twice as many alleles as the DSPR population. Independent studies of doublesex and transformer mutants support many novel connections, including evidence for a link between the sex hierarchy and metabolism, with the inclusion of Insulin-like receptor in the sex hierarchy GRN. Overall design: RNA sequencing was performed on mRNA derived from adult male or female heads, for a total of 9 samples. These samples included females that produce the male isoform of dsx [w/+;DsxD/dsxm+r15 (XX)], and two dsx mutants: females [w/+; dsxm+r15/dsxd+r3 (XX)] and males [w;dsxm+r15/dsxd+r3 (XY)]. Two wild type genotypes (Berlin and Canton-S) were sequenced at the same time, but have previously been published as part of GSE50515. There were at least 3 replicates from biological samples.
Sex Differences in Drosophila Somatic Gene Expression: Variation and Regulation by doublesex.
Sex, Specimen part, Cell line, Subject
View SamplesWe treated 6-8 week old mice that had floxed alleles of both Apc and Pten (for both alleles in each case) that also carry an Ovgp1-iCre-ERT2 transgene, with one of two treatments; a third group received neither treatment. The Ovgp1-iCre-ERT2 expresses Cre recombinase fused to a tamoxifen-inducible fragment of the estrogen receptor, in tissues where the Ovgp1 gene (oviductal glycoprotein 1) is expressed, which is almost exclusively in mouse oviductal epithelium (equivalent to human fallopian tube epithelium = FTE). Treating the mice with tamoxifen permits the Cre recombinase to enter the cell nucleus and inactivate the Apc and Pten genes. Six of the mice were treated with intraperitoneal injection of tamoxifen (0.1g/kg of body weight) dissolved in corn oil on days 1 and 3 and developed oviductal tumors (OdT) yielding 6 of the samples. Four mice (yielding 5 samples) were instead injected with 50 million plaque-forming units of replication-incompetent AdCre into both ovarian bursal cavities on day 1, which inactivated Apc and Pten in the ovarian surface epithelium (OSE), and lead to ovarian tumors (OT). Ovaries were also harvested from four untreated 6-8 week old mice with the same genotype, with two ovaries from each mouse comprising one control sample. RNA was purified from tumor or normal tissue, and targets for Affymetrix arrays synthesized from the mRNAs. We used Affymetrix Mouse Gene 2.1 ST arrays, which hold 41345 probe-sets, but we largely analyzed just those 25216 probe-sets that were mapped to Entrez gene IDs. Raw data was processed with Robust Multi-array Average algorithm (RMA). Data is log2-transformed transcript abundance estimates. We fit a one-way ANOVA model to the three groups of samples. We supply a supplementary excel workbook that holds the same data as the data matrix file, but also holds the probe-set annotation at the time we analyzed the data, and some simple statistical calculations, which select subsets of the probe-sets as differentially expressed. Consumers should consider obtaining more up-to-date probe-set annotation for the array platform. We also provide supplementary excel files that show our simple analysis of GSE6008, which consists of 99 human ovarian tumor samples of 4 types, and 4 normal ovary samples, where we fit an ANOVA model to the 5 groups. In yet another supplementary file we show the correlation between each human tumor and mouse tumor, where we correlate the difference in log2-transformed values of each tumor from the average of the normals for the same species, for just those genes that were 1-to-1 best homologs according to build 68 of NCBI's Homologene, in order to see how much the human tumors resemble the mouse tumors.
Impact of oviductal versus ovarian epithelial cell of origin on ovarian endometrioid carcinoma phenotype in the mouse.
Sex
View Samples10 normal squamous cervical epitheilia samples, 7 high grade squamous intraepithelial lesions, and 21 invasive squamous cell carcinomas of the cervix samples were obtained using laser capture miicrodissection. Two rounds of T7-based linear RNA amplification using the Arcturus RiboAmp kit were performed for each sample, and assayed using Affymetrix HG_U133A arrays.
Gene expression analysis of preinvasive and invasive cervical squamous cell carcinomas identifies HOXC10 as a key mediator of invasion.
No sample metadata fields
View SamplesWe mated mice with floxed alleles of both Apc and Pten, to mice with floxed alleles for Arid1a, to obtain female mice with both copies of all three genes floxed. At 7 to 8 weeks of age the right ovarian bursal cavites of the mice were injected with 50 million plaque-forming units of adenovirus expressing Cre recombinase, which causes the floxed genes to be knocked out. Tumor tissue from 3 mice for each group was obtained at necropsy, RNA purified, and targets for Affymetrix arrays synthesized from the mRNAs. We used Affymetrix Mouse Genome 430 2.0 arrays, which hold 45101 probe-sets. Raw data was processed with Robust Multi-array Average algorithm (RMA). Data is log2-transformed transcript abundance estimates. We performed T-tests to compare the 3 vs 3 arrays. We supply a supplementary excel workbook that holds the same data as the data matrix file, but also holds the probe-set annotation at the time we analyzed the data, and some very simple statistical calculations, which select subsets of the probe-sets as differentially expressed. Consumers should consider obtaining more up-to-date probe-set annotation for the array platform. We have also supplied a second supplementary tar archive holding software and files to 1) perform permutation testing of the probe-set selection in order to estimate false discovery rates for the probe-sets we selected as differentially expressed, 2) perform enrichment testing of GO terms, and 3) to perform enrichment testing of KEGG pathways and 3000 curated gene sets from version 4 of the Molecular Signatures Database (MSigDB). The software is in "C".
Arid1a inactivation in an Apc- and Pten-defective mouse ovarian cancer model enhances epithelial differentiation and prolongs survival.
Sex
View SamplesThe mechanisms underlying muscle wasting in cancer patients remain poorly understood, and consequently there remains an unmet clinical need for new biomarkers and treatment strategies.
Suppression of skeletal muscle turnover in cancer cachexia: evidence from the transcriptome in sequential human muscle biopsies.
Sex, Age, Specimen part
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