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pubmed:abstractText |
The post-genome sequencing era of Saccharomyces cerevisiae is defined by the analysis of newly discovered open reading frames of unknown function. In this report, we describe a genetic method for the rapid identification and characterisation of genes involved in a given phenotype. This approach is based on the ability of overexpressed genomic DNA fragments to cure an induced phenotype in yeast. To validate this concept, yeast cells carrying a yeast DNA library present on multicopy plasmid vectors were screened for resistance to the antifungal drug ketoconazole. Among 1.2 million colonies 13 clones tested positive, including those expressing the lanosterol C-14 demethylase, known to be a cellular target for azole drugs, and the cytochrome-c oxidase of mitochondria, regulating the respiratory chain electron transport. Several other resistant clones were identified, which code for yeast proteins of so far unknown function. These genes may represent potential candidates for antifungal drug effects. Together with the availability of the entire yeast genome sequence, the described genetic screening method is a powerful tool for the effective functional analysis of yeast genes.
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