Variation in cDNA microarray analysis of gene expression using unamplified poly(A)+ RNA
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Hongjuan Zhao
PROVIDER: E-GEOD-3559 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Publications
Optimization and evaluation of T7 based RNA linear amplification protocols for cDNA microarray analysis.
Zhao Hongjuan H Hastie Trevor T Whitfield Michael L ML Børresen-Dale Anne-Lise AL Jeffrey Stefanie S SS
BMC genomics 20021030 1
<h4>Background</h4>T7 based linear amplification of RNA is used to obtain sufficient antisense RNA for microarray expression profiling. We optimized and systematically evaluated the fidelity and reproducibility of different amplification protocols using total RNA obtained from primary human breast carcinomas and high-density cDNA microarrays.<h4>Results</h4>Using an optimized protocol, the average correlation coefficient of gene expression of 11,123 cDNA clones between amplified and unamplified ...[more]
Similar Datasets
Project description:Summary: Breast cancer cell lines have been used widely to investigate breast cancer pathobiology and new therapies. Breast cancer is a molecularly heterogeneous disease, and it is important to understand how well and which cell lines best model that diversity. In particular, microarray studies have identified molecular subtypes (luminal A, luminal B, ERBB2-associated, basal-like and normal-like) with characteristic gene-expression patterns and underlying DNA copy number alterations (CNAs). Here, we studied a collection of breast cancer cell lines to catalog molecular profiles and to assess their relation to breast cancer subtypes. Whole-genome DNA microarrays were used to profile gene expression and CNAs in a collection of 52 widely-used breast cancer cell lines, and comparisons were made to existing profiles of primary breast tumors. Hierarchical clustering was used to identify gene-expression subtypes, and Gene Set Enrichment Analysis (GSEA) to discover biological features of those subtypes. Genomic and transcriptional profiles were integrated to discover within high-amplitude CNAs candidate cancer genes with coordinately altered gene copy number and expression. Transcriptional profiling of breast cancer cell lines identified one luminal and two basal-like (A and B) subtypes. Luminal lines displayed an estrogen receptor (ER) signature and resembled luminal-A/B tumors, basal-A lines were associated with ETS-pathway and BRCA1 signatures and resembled basal-like tumors, and basal-B lines displayed mesenchymal and stem-cell characteristics. Compared to tumors, cell lines exhibited similar patterns of CNA, but an overall higher complexity of CNA (genetically simple luminal-A tumors were not represented), and only partial conservation of subtype-specific CNAs. We identified 80 high-level DNA amplifications and 13 presumptive homozygous deletions, and the resident genes with concomitantly altered gene-expression, highlighting known and novel candidate breast cancer genes. Overall, breast cancer cell lines were genetically more complex than tumors, but retained expression patterns with relevance to the luminal-basal subtype distinction. The compendium of molecular profiles defines cell lines suitable for investigations of subtype-specific pathobiology, biomarkers and therapies, and provides a resource for discovery of new breast cancer genes. HEEBO oligonucleotide microarrays from the Stanford Functional Genomics Facility were used to perform gene expression profiling of 50 human breast epithelial cell lines, in comparison to a universal RNA reference. Expression data were analyzed by hierarchical clustering to identify subgroups, and gene set enrichment analysis to identify subgroup-specific gene pathways.
2010-06-24 | E-GEOD-15361 | biostudies-arrayexpress
Project description:This experiment set contains the raw data for 8 arrays that were used in the genomic typing of the pre- and post-mouse H. pylori strains SS1 and SS2000. 10700 is the pre-mouse clinical isolate of SS1 and PMSS2000 is the pre-mouse clinical isolate of SS2000. gDNA from these strains were labeled and hybridized to H. pylori microarrays as described in Salama et al. {Salama et al. 2000 PNAS 97:14668-73}. In each case the test gDNA sample was labeled with Cy5 (red) and this was hybridized with Cy3 (green) labeled reference DNA of equal amount. The reference DNA consisted of equal amounts of gDNA from the two H. pylori strains used to make the H. pylori microarray, 26695 and J99. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Computed
2006-01-06 | E-GEOD-3988 | biostudies-arrayexpress
Project description:Using a C. elegans whole genome DNA microarray in this study, the effects of five different xenobiotics on the gene expression of the nematode were investigated. The exposure time for the following five applied compounds beta-NF (5 mg/l), Fla (0.5 mg/l), atrazine (25 mg/l), clofibrate (10 mg/l) and DES (0.5 mg/l) was 48+/-5 h. The analysis of the data showed a clear induction of 203 genes belonging to different families like the cytochromes P450, UDP-glucoronosyltransferases (UDPGT), glutathione S-transferases (GST), carboxylesterases, collagenes, C-type lectins and others. Under the applied conditions, fluoranthene was able to induce most of the induceable genes, followed by clofibrate, atrazine, beta-naphthoflavone and diethylstilbestrol. A decreased expression could be shown for 153 genes with atrazine having the strongest effect followed by fluoranthene, diethylstilbestrol, beta-naphthoflavone and clofibrate. For upregulated genes a change ranging from approximately 2.1- till 42.3-fold and for downregulated genes from approximately 2.1 till 6.6-fold of gene expression could be affected through the applied xenobiotics. Sample Treatments (by exptids) Clofibrate: 14317, 16443, 16505 Fluoranthene: 33664, 33667, 33669, 23484 beta-Naphthoflavone: 6844, 14320, 14316 Atrazin: 33672, 33674, 23487, 23486 DES: 33671, 23485 A compound treatment design type is where the response to administration of a compound or chemical (including biological compounds such as hormones) is assayed. Computed
2006-01-27 | E-GEOD-4111 | biostudies-arrayexpress
Project description:ABSTRACT Background. Acute Kawasaki disease (KD) is difficult to distinguish from other acute rash/fever illnesses, in part because the etiologic agent(s) and pathophysiology remain poorly characterized. As a result, diagnosis and critical therapies may be delayed. Methods. We used DNA microarrays to identify possible diagnostic features of KD. We compared gene expression patterns in the blood of 23 children with acute KD and 18 age-matched febrile children with three illnesses that resemble KD. Results. Genes associated with platelet and neutrophil activation were expressed at higher levels in KD patients than in patients with acute adenovirus infections or systemic adverse drug reactions but not in patients with scarlet fever; genes associated with B cell activation were also expressed at higher levels in KD patients than in controls. A striking absence of interferon-stimulated gene expression in the KD patients was confirmed in an independent cohort of KD subjects. We successfully predicted the diagnosis in 21 of 23 KD patients and 7 of 8 adenovirus patients using a set of 38 gene transcripts. Conclusions. These findings provide insight into the molecular features that distinguish KD from other febrile illnesses, and support the feasibility of developing novel diagnostic reagents for KD based on the host response. A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied. Disease State: One of Kawasaki Disease (KD) or control (C) of Scarlet fever (C-sf), adenovirus infection (C-ai) or drug reaction (C-dr) disease_state_design
2010-06-24 | E-GEOD-15297 | biostudies-arrayexpress
Project description:We generated RNA's and measured expression of 40000 genes using spotted cDNA microarrays from the fifty nine publicly available cell lines of the NCI Developmental Therapeutics Program's NCI60 studies and an additional set of seven cell lines for which GI50 compound sensitivity data were available. All cell lines were grown to 80% confluence in RPMI 1640 supplemented with phenol red, glutamine (2 mM) and 5% fetal calf serum. This expression data, in conjunction with the compound sensitivity data sets available from the DTP, were used to empirically determine whether gene-compound correlates of a sufficiently high correlation coefficient would have a suitable low false discovery rate to support the use of a correlative approach and these datasets for early discovery approaches for new targeted therapies. A cell type comparison design experiment design type compares cells of different type for example different cell lines. Sixty six cell lines were analysed, with no repetitions. Twelve cell lines were pooled for a common reference.
2008-02-07 | E-GEOD-7947 | biostudies-arrayexpress
Project description:The first goal of the study was to find the genes that explain the difference in progression of age-related retinal degeneration (ageRD) between the inbred strains C57BL/6J-c2J (B6a) and BALB/cByJ (C). The specific goal was to find the candidate genes that explain the chromosome 6 and chromosome 10 quantitative trait loci (QTL) found in a previous study. Genes differentially expressed in retina between both strains and that map to the QTL are candidate genes. By integrating different approaches: microarray study, knowledge-based candidate gene search and screening of genetic variants, we found 35 candidate genes for chromosome 6 QTL and 14 candidate genes for chromosome 10 QTL. Five genes for each QTL were selected for sequencing and qPCR analysis. One gene, Tnfaip3 was selected for phenotypic testing and was excluded as the quantitative trait gene for chr10 QTL. The second goal of the study was to find biological processes and pathways associated with ageRD by Gene Ontology analysis of the microarray data. We concentrated on GO terms overrepresented in either strain at 8 months; the age when there is a difference in the phenotype (outer nuclear layer thickness). The GO analysis revealed that transcripts associated with inflammation, mitochondrial envelope, DNA repairing and oxidative stress were increased in the ageRD sensitive C strain with age (8 months). In the ageRD resistant B6a strain transcripts associated with antioxidant response, regulation of blood vessel size and growth factor activity were increased with age. Transcripts associated with cell remodeling and lipid metabolism were increased in either strain with age. This study aims to identify gene expression differences in posterior eyecups between the strains C and B6a at 4 months and at 8 months. Twelve animals were used per strain and time point for a total of 48 mice. Total RNA for 3 animals (6 eyes) were pooled giving a total of 4 biological replicates per strain and time point. The 4-month C samples labeled with Cy5 were hybridized with the 4-month B6a samples labeled with Cy3 (4 biological replicates); the 8-month C samples labeled with Cy5 were hybridized with the 8-month B6a samples labeled with Cy3, for 8 arrays (4 biological replicates). A second set of eight arrays were used to analyzed the same RNA samples but with the dye orientation reversed, yielding a total of 16 arrays in the experiment. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc.
2010-05-12 | E-GEOD-18151 | biostudies-arrayexpress
Project description:Spots were selected that had an expression level higher than 1.6 the local background. The use of a common reference sample allows the comparison of the relative expression levels across the tissue samples. All genes were expressed relative to their median expression level across the arrays of the same microarray platform, allowing us to combine the data from the 2 platforms. Data from Lymphochip and Stanford Human cDNA arrays was combined per patient. Only genes with reliable datapoints in at least 80% of the samples were used. Subsequently, genes with the same unigene identifier were averaged in SMD. For all further analysis genes were selected that showed at least a two-fold difference in expression level in at least one array. A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied. Using regression correlation
2007-09-13 | E-GEOD-9027 | biostudies-arrayexpress
Project description:Measurement of the relative changes of gene expression of S. cerevisiae cells lacking either trf4 (trf4delta) or trf5 (trf5delta), and trf4delta/TRF4-DADA mutants compared to wild-type (WT) cells using yeast oligo microarrays that contain features representing all annotated yeast ORFs, ncRNAs, introns, rRNA precursors, as well as some intergenic regions (IGRs) and tiled regions downstream of a few genes. Total RNA was isolated by hot phenol extraction from exponentially growing cells, and reverse transcribed with a mixture of random nonamers and oligo(dT) primers in the presence of amino-allyl dUTP/dNTP mixture. Cy5 fluorescently labeled cDNAs derived from total RNA isolated from either the trf4delta or the trf5delta mutants, and the trf4delta/TRF4-DADA mutants were then competitively hybridized with Cy3 labeled cDNAs from WT cells. cDNAs were hybridized on yeast oligo microarrays over night at 42 degrees in formamide-based hybridization buffer. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. mutated gene: paralogous non-canonical poly(A) polymerases Trf4p and Trf5p forming TRAMP4 and TRAMP5 complexes Computed
2010-06-24 | E-GEOD-16103 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE16103: Wild-type versus trf4, trf5, and trf4-DADA mutant cells GSE16105: Trf4p in vivo crosslinking and ribonucleoprotein-immunopurification-chip analysis (X-RIP-Chip) Refer to individual Series
2010-06-24 | E-GEOD-16107 | biostudies-arrayexpress
Project description:Streptococcus pneumoniae normally resides in the human nasopharynx in a non-disease state. In response to yet unknown triggers it can descend to the lower respiratory tract and/or invade the bloodstream. Regulation and activation of virulence genes play essential roles in this process of disease development. A putative transcriptional regulator in S. pneumoniae, MgrA, with homology to a virulence gene activator, mga, of Group A streptococcus (GAS) was previously identified as being required for development of pneumonia in a murine model. In this work we confirm that mgrA is required for both nasopharyngeal carriage and pneumonia. Transcriptional profiling by microarray technology through the growth course of a strain that bears a deletion of mgrA (AC1500) with that of a strain that over expresses Mgra (AC1481) is used to show that MgrA . This is manifested phenotypically by a decrease in adherence to epithelial cells in tissue culture since rlrA pathogenicity islet contains genes mediating adherence. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Computed
2005-08-11 | E-GEOD-3107 | biostudies-arrayexpress