Successive truncation of yeast genome toward the construction of 'minimal genome' yeast.
Kiriko Murakami (1), Yuki Ito (1), Eriko Tao (1), Minetaka Sugiyama (2), Yoshinobu Kaneko (2), Satoshi Harashima (2), Takahiro Sumiya (3), Atsushi Nakamura (3), Masafumi Nishizawa (1)
(1) Dept. Microbiol. Immunol., Keio Univ. School of Medicine, Shinjuku, Tokyo, 160-8582, Japan; (2) Department of Biotechnology, Graduate School of Engineering, Osaka University, Suita, Japan; (3) RIISE, Hiroshima University, Higashi-Hiroshima, Japan
In the yeast Saccharomyces cerevisiae, the PCR-mediated Chromosome Splitting (PCS) method has been developed by Harashima's group to retrieve any chromosome regions of interest into a mini-chromosome. The PCS method uses two chromosome splitting modules, one is composed of a selectable marker, the target sequence, and (C 4 A 2 ) 6 sequence which can serve as telomere in yeast, and the other contains the CEN4 sequence in place of the marker. A yeast strain having more than 20 chromosomes has been generated by this method. When using a splitting module that lacks the CEN4 sequence, we can delete the resulting split fragment since a fragment without CEN is lost after chromosome segregation. This deletion technique is useful for improvement of yeast by deleting genes unnecessary for or inhibitory to production of useful materials, and also enables us to know how many genes are required for yeast growth. We developed a program by which a region where essential, functionally important, or synthetic lethal genes are not present can be selected. We carried out successive deletion of these regions, and so far obtained a strain whose genome had been deleted by a total of 320 kb. We also constructed strains with different combination of deletions. Properties of these yeast strains having truncated genome will be presented. This study was carried out as a part of The Project for Development of a Technological Infrastructure for Industrial Bioprocesses on R&D of NISTF by METI and NEDO.