A combination of flow cytometry and traditional screening using chemicals to isolate high glutathione-producing yeast mutants.
Traditional screening using chemicals or flow cytometry (FCM) alone is not sufficient to isolate the high glutathione (GSH)-producing yeast strains used in food production. Therefore, to improve screening efficiency, we investigated a combination of both methods. A mutated Saccharomyces cerevisiae strain was labeled with 5-chloromethylfluorescein diacetate and sorted by FCM according to emitted fluorescence intensity. Moderate GSH (1%-2%)-producing mutants were isolated, whereas high GSH (>2%)-producing mutants were not. Traditional screening using cerulenin resulted in similar findings, but a combination of both methods resulted in a 40% increase in the screening yield of high GSH-producing mutants. An analysis of model strains indicated that the ratio of high GSH-producing cells in a sample affected the FCM results. By combining FCM with traditional screening using chemicals, we succeeded in isolating high GSH-producing mutants from several parental strains.
Ajinomoto Co., Inc., Kanagawa, Japan. hiroaki_nishiuchi@ajinomoto.com
Hiroaki
H
Nishiuchi
Yukiko
Y
Tabira
Kazuo
K
Yamagishi
D002569:Cerulenin / Q000494:pharmacology; D054872:Fatty Acid Synthesis Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005524:Food Technology; D005978:Glutathione / Q000096:biosynthesis* / Q000235:genetics; D057166:High-Throughput Screening Assays; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
D054872:Fatty Acid Synthesis Inhibitors; D005452:Fluoresceins; D005456:Fluorescent Dyes; C069306:5-chloromethylfluorescein; D002569:Cerulenin; D005978:Glutathione