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Large-scale analysis of the yeast genome by transposon tagging and gene disruption

Abstract

Economical methods by which gene function may be analysed on a genomic scale are relatively scarce. To fill this need, we have developed a transposon-tagging strategy for the genome-wide analysis of disruption phenotypes, gene expression and protein localization, and have applied this method to the large-scale analysis of gene function in the budding yeast Saccharomyces cerevisiae. Here we present the largest collection of defined yeast mutants ever generated within a single genetic background—a collection of over 11,000 strains, each carrying a transposon inserted within a region of the genome expressed during vegetative growth and/or sporulation. These insertions affect nearly 2,000 annotated genes, representing about one-third of the 6,200 predicted genes in the yeast genome1,2. We have used this collection to determine disruption phenotypes for nearly 8,000 strains using 20 different growth conditions; the resulting data sets were clustered to identify groups of functionally related genes. We have also identified over 300 previously non-annotated open reading frames and analysed by indirect immunofluorescence over 1,300 transposon-tagged proteins. In total, our study encompasses over 260,000 data points, constituting the largest functional analysis of the yeast genome ever undertaken.

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Figure 1: The mTn insertion project.
Figure 2: Distribution and phenotypic analysis of 1,917 mTn-mutagenized ORFs within the S. cerevisiae genome.
Figure 3: Phenotypic macroarray analysis.
Figure 4: Graphical representation of clustered phenotypic data.
Figure 5: Immunolocalization of epitope-tagged proteins.

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Acknowledgements

P.S.R.C. is supported by the Fundacao de Amparo a Pesquisa do Estado de Sao Paulo, Brazil. This work was supported by an NIH grant (to G.S.R. and M.S.).

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Correspondence to Michael Snyder.

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Ross-Macdonald, P., Coelho, P., Roemer, T. et al. Large-scale analysis of the yeast genome by transposon tagging and gene disruption. Nature 402, 413–418 (1999). https://doi.org/10.1038/46558

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