Novel method for systematic analysis of essential yeast genes

Masashi Yukawa(1), Daisuke Fukuoka(1), Kintake Sonoike(1), Hiroshi Sawai(2), Shinichi Morishita(3), Yoshikazu Ohya(1). (1) Integrated Biosciences, University of Tokyo, Kashiwanoha 5-1-5, Kashiwa, 277-8562, JAPAN; (2) Institute for Bioinformatics Research and Development, Japan Science and Technology Agency; (3) Department of Computational Biology, University of Tokyo.

More than 1,000 of the ~6,000 known or predicted protein-coding yeast genes are essential for cell viability. Utilizing the collection of heterozygous diploid deletion strains (produced by EUROFAN project), we established a novel method for phenotypic analysis of essential gene deletion mutants. This method makes use of the yeast differentiation pathway that diploid cells form four haploid cells in meiosis, two of which are wild-type cells and the other two are gene-disrupted cells. First, we have replaced the KanMX module with the NLS-EGFP cassette. Additionally, we have devised the way to prevent the morphological change by response to mating pheromone. By these two genetic manipulations that can be performed systematically, we can select the haploid cells deleted with essential gene as the cells with nuclear fluorescence after release them into growth medium. To demonstrate the availability of the constructed strains, we confirmed phenotypes of several essential gene deletion mutants including cdc mutants. Next, we defined the critical time points to observe the phenotype of each strain, and described their phenotypes by the image-processing program. With this program, we can obtain various quantitative data about cell morphology from microscopic images. Combining the way of the strain construction and its phenotypic description, we will present that this method is valuable to systematically analyze the phenotypes of essential gene disruptants.

Program Nr. 508A from 2004 Yeast meeting

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