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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 |
