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Methods Mol Biol. 2009;548:115-43. doi: 10.1007/978-1-59745-540-4_7.

Chemogenomic approaches to elucidation of gene function and genetic pathways.

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  • 1Department of Pharmaceutical Sciences, University of Toronto, Toronto, ON, Canada M5S 3E1.

Abstract

The approximately 6,000 strains in the yeast deletion collection can be studied in a single culture by using a microarray to detect the 20 bp DNA "barcodes" or "tags" contained in each strain. Barcode intensities measured by microarray are compared across time-points or across conditions to analyze the relative fitness of each strain. The development of this pooled fitness assay has greatly facilitated the functional annotation of the yeast genome by making genome-wide gene-deletion studies faster and easier, and has led to the development of high throughput methods for studying drug action in yeast. Pooled screens can be used for identifying gene functions, measuring the functional relatedness of gene pairs to group genes into pathways, identifying drug targets, and determining a drug's mechanism of action. This process involves five main steps: preparing aliquots of pooled cells, pooled growth, isolation of genomic DNA and PCR amplification of the barcodes, array hybridization, and data analysis. In addition to yeast fitness applications, the general method of studying pooled samples with barcode arrays can also be adapted for use with other types of samples, such as mutant collections in other organisms, siRNA vectors, and molecular inversion probes.

PMID:
19521822
[PubMed - indexed for MEDLINE]
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