The construction and application of diploid sake yeast with a homozygous mutation in the FAS2 gene.
In Japanese sake brewing, cerulenin-resistant sake yeasts produce elevated levels of ethyl caproate, an important flavor component. The FAS2 mutation FAS2-1250S of Saccharomyces cerevisiae generates a cerulenin-resistant phenotype. This mutation is dominant, and, in general, cerulenin-resistant diploid sake yeast strains carry this mutation heterozygously. Here we constructed diploid sake yeast with a homozygous mutation of FAS2 using the high-efficiency loss of heterozygosity method. The homozygous mutants grew more slowly in YPD medium than did the wild-type and heterozygous mutants, and they produced more ethyl caproate during sake brewing. In addition, although both the wild-type and heterozygous mutant were sensitive to 4 mg/l cerulenin, the homozygous mutant was resistant to more than 4 mg/l cerulenin. Next, we obtained a homozygous mutant of FAS2 without inducing genetic modification. After cultivating the heterozygous FAS2 mutant K-1801 in YPD, homozygous mutants were selected on medium containing high concentrations of cerulenin. Non-genetically modified yeast with a homozygous mutation of FAS2 produced 2.2-fold more ethyl caproate than did heterozygous yeast. Moreover, high-quality Japanese sake with a very rich flavor could be brewed using yeast containing a homozygous mutation in the FAS2 gene.
Kotaka
Atsushi
A
Research Institute, Gekkeikan Sake Co. Ltd., 101 Shimotoba-koyanagi-cho, Fushimi-ku, Kyoto 612-8385, Japan. a_kotaka@gekkeikan.co.jp
Sahara
Hiroshi
H
Hata
Yoji
Y
D000434:Alcoholic Beverages; D002208:Caproates / Q000378:metabolism*; D002569:Cerulenin / Q000494:pharmacology; D004171:Diploidy; D005800:Genes, Fungal; D006720:Homozygote; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism
D002208:Caproates; D002569:Cerulenin; C079237:ethyl hexanoate