A Nested Parallel Experiment Demonstrates Differences in Intensity-Dependence Between RNA-Seq and Microarrays
ABSTRACT: We carried out a parallel nested experiment performed simultaneously on RNA-Seq and microarrays that systematically split variation into four stages (treatment, biological variation, library preparation, and chip/lane noise), allowing a separation and comparison of the sources of variation in a well-controlled cellular system, Saccharomyces cerevisiae. Nested design. Two conditions (E for ethanol and G for glucose as sole carbon source), each grown on two biological replicates (1 and 2), prepared in two technical preparations (A and B), then hybridized to two chips (1 and 2) This submission represents microarray component of study.
Project description:Understanding the structure and interplay of cellular signalling pathways is one of the great challenges in molecular biology. Boolean Networks can infer signalling networks from observations of protein activation. In situations where it is difficult to assess protein activation directly, Nested Effect Models are an alternative. They derive the network structure indirectly from downstream effects of pathway perturbations. To date, Nested Effect Models cannot resolve signalling details like the formation of signalling complexes or the activation of proteins by multiple alternative input signals. Here we introduce Boolean Nested Effect Models (B-NEM). B-NEMs combine the use of downstream effects with the higher resolution of signalling pathway structures in Boolean Networks. We show that B-NEMs accurately reconstruct signal flows in simulated data. Using B-NEM we then resolve BCR signalling via PI3K and TAK1 kinases in BL2 lymphoma cell lines. 84 BL2 cell-line samples were hybridized to HGU133+2 Affymetrix GeneChips.
Project description:The purpose of this study was to improve prediction of patients at high-risk for metastatic disease utilizing a nested case-control design that uniquely enables enrichment for relevant phenotypes. We identified all women diagnosed with primary breast cancer from January 1, 1997, to December 31, 2005, in the Stockholm health care region. Patients developing distant metastatic disease (cases) were selected and controls (free from distant disease) were randomly matched by adjuvant therapy, age and calendar period at diagnosis. The nested case-control study included 768 study subjects with clinical information and gene expression arrays (Human Cancer G110). Study subjects were randomly and equally divided into training set (discovery) or testing (validation) set. Metastatic onset prediction was then compared including either clinical variables only or combining clinical and genetic information. Differentially expressed genes and pathways between cases and controls included a wide-spectrum of well known as well as candidate regulators of the metastatic cascade. The nested case-control study included 768 study subjects corresponding to 623 primary tumor samples. Details concerning case-control status are given in the samples section. Each case and its' matching controls form risk sets, indicated by the setnr variable.
Project description:Aneuploidy, or an aberrant karyotype, results in developmental disabilities and has been implicated in tumorigenesis. However, the causes of aneuploidy-induced phenotypes and the consequences of aneuploidy on cell physiology remain poorly understood. We have performed a meta-analysis on gene expression data from aneuploid cells in diverse organisms, including yeast, plants, mice, and humans. We found highly-related gene expression patterns that are conserved between species: genes that were involved in the response to stress were consistently upregulated, while genes associated with the cell cycle and cell proliferation were downregulated in aneuploid cells. Within species, different aneuploidies induced similar changes in gene expression, independent of the specific chromosomal aberrations. Taken together, our results demonstrate that aneuploidies of different chromosomes and in different organisms impact similar cellular pathways and cause a stereotypical anti-proliferative response that must be overcome prior to transformation. These experiments are two-color hybridizations of RNA isolated from aneuploid samples vs matched wt cells, all grown to midlog phase.
Project description:Telomeres and the protein/RNA complexes involved in maintaining them are rapidly evolving systems across eukaryotes. Using two Saccharomyces species, S. cerevisiae and S. bayanus, we provide evidence that the telomere systems of these two closely related yeasts have evolved significantly apart and that the gene in one species can not maintain the set-point of telomere length of the other species in the hybrid. Each array has co-hybridized RNA vs DNA for the indicated species (BB9=S. bayanus, CC5=S. cerevisiae) or hybrid (BC11). The cultures for RNA preparation were grown to midlog phase at the indicated temperature.
Project description:These RNA-seq samples represent ten different tissue types within a diverse Nested Association Mapping (NAM) maize population that has been sequenced by the NAM Consortium Group. These samples correspond to project IDs PRJEB31061.
Project description:Aneuploidy and aging are correlated; however, a causal link between these two phenomena has remained elusive. Here we show that yeast disomic for a single native yeast chromosome generally have a decreased replicative lifespan. In addition, the extent of this lifespan deficit correlates with the size of the extra chromosome. We identified a mutation in BUL1 that rescues both the lifespan deficit and a protein trafficking defect in yeast disomic for chromosome 5. Bul1 is an E4 ubiquitin ligase adaptor involved in a protein quality-control pathway that targets membrane proteins for endocytosis and destruction in the lysosomal vacuole thereby maintaining protein homeostasis. Concurrent suppression of the aging and trafficking phenotypes suggests that disrupted membrane protein homeostasis in aneuploid yeast may contribute to their accelerated aging. The data reported here demonstrate that aneuploidy can impair protein homeostasis, shorten lifespan, and may contribute to age-associated phenotypes. These are all CGH arrays comparing DNA content between the indicated strain of interest and a wt control.
Project description:This is a quality control (QC) substudy of GSE48091. The QC substudy comprises gene-expression profiling of re-extracted tumor RNA for a subset of the tumours in the full study. As background, a population-based cohort study of metastatic breast cancer patients was first designed. Thereafter, a case-control study nested in the corresponding population-based cohort of primary breast cancer patients was designed by selecting distant metastasis-free controls to each case. Tumor RNA was extracted in the same order. All RNA was profiled on microarrays in randomized order. For quality control, RNA was also re-extracted (new tumor piece) in a randomized order for randomly selected cases-controls sets and profiled with the rest. Keywords: Expression profiling by array The nested case-control study included 768 study subjects corresponding to 623 primary tumor samples. This QC substudy comprises 97 of the study subjects (all different primary tumor samples). Details concerning case-control status are given in the samples section. This Series includes a re-analysis of 97 samples from GSE48091. The file "full_data_matrix.txt" includes the re-normalized values for the 97 samples from GSE48091 and the normalized values for the 97 new samples from the same patients that were analyzed together.
Project description:In oligotrophic ocean waters where bacteria are often subjected to chronic nutrient limitation, community transcriptome sequencing has pointed to the presence of highly abundant small RNAs (sRNAs). The role of sRNAs in regulating response to nutrient stress was investigated in a model heterotrophic marine bacterium Ruegeria pomeroyi grown in continuous culture under carbon and nitrogen limitation. RNAseq analysis identified 98 sRNAs, of which 69 were cis-encoded and located antisense to their target genes, and 30 were trans-encoded and linked to predicted target genes through complementarity analysis. The most prevalent functional roles of target genes were transport, cell-cell interactions, signal transduction, and transcriptional regulation. Thirty-two percent of the sRNAs had been identified in a previous study of R. pomeroyi growth on organic sulfur compounds, and may be constitutively expressed, while 69% were not identified in previous studies. Eighty-six percent and were transcribed equally under both carbon and nutrient limitation, and may be involved in a general stress response; 14% were differentially regulated under carbon versus nitrogen stress, and may respond to specific nutrient limitations. A network analysis of the predicted target genes of the R. pomeroyi sRNAs indicated that they average fewer connections than typical protein-encoding genes, and appear to be more important in peripheral or niche-defining functions encoded in the pan genome rather than central metabolism encoded in the core genome.
Project description:The Illumina Infinium 27k Human DNA methylation Beadchip v1.2 was used to obtain DNA methylation profiles across approximately 27,000 CpGs in normal cells from the uterine cervix (liquid based cytology samples), obtained from 152 women in a nested prospective case-control study within the ARTISTIC trial. Out of the 152 women, 75 developed a cervical intraepitelial neoplasia of grade 2 or higher (CIN2+) after 3 years of sample collection. The rest of women (77) remained disease free (CIN2-). Overall design: Bisulphite converted DNA from the 152 samples were hybridised to the Illumina Infinium 27k Human Methylation Beadchip v1.2