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Universal Microarray System (UMAS) to investigate yeast response to galactose

ABSTRACT: We describe a transcriptional analysis platform consisting of a universal micro-array system (UMAS) combined with an enzymatic manipulation step that is capable of generating expression profiles from any organism without requiring a priori species-specific sequence information. The transcriptome is converted to cDNA and processed with restriction endonucleases to generate low-complexity pools (80–120) of equal length DNA fragments. The resulting material is amplified and detected with the UMAS system, comprising all possible 4,096 (4^6) DNA hexamers. Ligation to the arrays yields thousands of 14-mer sequence tags. The compendium of signals from all pools in the array-of-universal arrays comprises a full-transcriptome expression profile. The technology was validated by analysis of the galactose response of Saccharomyces cerevisiae, and the resulting profiles showed excellent agreement with the literature and real-time PCR assays. Keywords: other

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE993 | GEO | 2004/03/12

SECONDARY ACCESSION(S): PRJNA88043

REPOSITORIES: GEO

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Project description:This SuperSeries is composed of the following subset Series: GSE3972: Contribution of unique genes by individual cell lines to Universal Human Reference RNA GSE3973: Microarray Coverage of Universal Human Reference RNA on microarrays with yeast controls GSE3974: Contribution of unique genes by individual cell lines to Universal Mouse Reference RNA Abstract: BACKGROUND: Obtaining reliable and reproducible two-color microarray gene expression data is critically important for understanding the biological significance of perturbations made on a cellular system. Microarray design, RNA preparation and labeling, hybridization conditions and data acquisition and analysis are variables difficult to simultaneously control. A useful tool for monitoring and controlling intra- and inter-experimental variation is Universal Reference RNA (URR), developed with the goal of providing hybridization signal at each microarray probe location (spot). Measuring signal at each spot as the ratio of experimental RNA to reference RNA targets, rather than relying on absolute signal intensity, decreases variability by normalizing signal output in any two-color hybridization experiment. RESULTS: Human, mouse and rat URR (UHRR, UMRR and URRR, respectively) were prepared from pools of RNA derived from individual cell lines representing different tissues. A variety of microarrays were used to determine percentage of spots hybridizing with URR and producing signal above a user defined threshold (microarray coverage). Microarray coverage was consistently greater than 80% for all arrays tested. We confirmed that individual cell lines contribute their own unique set of genes to URR, arguing for a pool of RNA from several cell lines as a better configuration for URR as opposed to a single cell line source for URR. Microarray coverage comparing two separately prepared batches each of UHRR, UMRR and URRR were highly correlated (Pearson's correlation coefficients of 0.97). CONCLUSION: Results of this study demonstrate that large quantities of pooled RNA from individual cell lines are reproducibly prepared and possess diverse gene representation. This type of reference provides a standard for reducing variation in microarray experiments and allows more reliable comparison of gene expression data within and between experiments and laboratories. Refer to individual Series

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Universal Microarray System (UMAS) to investigate yeast response to galactose

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