Use of Protoplast Fusion for the Development of Rapid Starch-fermenting Strains of Saccharomyces diastaticus

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We bred potent starch fermenting strains of Saccharomyces diastaticus by means of protoplast fusion. To improve the starch fermentation ability, hybrids carrying unlinked STA genes, which permit fermentation of starch, were introduced by protoplast fusion of haploid strains of identical mating type. Combination of protoplasts prepared from cells of haploid strains with amino acid requirements, cycloheximide resistance or respiration deficiency were fused in the presence of CaCl2 and polyethyleneglycol 6000. Such fusants thus obtained were selected as regenerated colonies on a minimal medium. The frequencies of fusant formation varied depending upon the strains used and were 7.6 × 10-6 to 4.1 × 10-4 for the regenerated protoplasts. Most of the fusants could mate with strains of the opposite mating type, although spore formation by the resultant non-maters was poor. The DNA content and cell volume of the fusants were greater than those of the parental strains. Some fusants produced more ethanol and amylase than the parental strains. Thus, the method we used seemed to be favorable for the breeding of potent starch fermenting strains.
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