CONSTRUCTION OF URACIL AND TRYPTOPHAN AUXOTROPHIC MUTANTS FROM SAKE YEASTS BY DISRUPTION OF URA3 AND TRP1 GENES
CONSTRUCTION OF URACIL AND TRYPTOPHAN AUXOTROPHIC MUTANTS FROM SAKE YEASTS BY DISRUPTION OF URA3 AND TRP1 GENES
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DOI：
10.1080/00021369.1990.10870447
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发表时间：
1990-11-01
期刊：
影响因子：
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通讯作者：
TAKAHASHI, K
中科院分区：
文献类型：
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作者：
KITAMOTO, K;ODA, K;TAKAHASHI, K
以清酒酵母K-701和K-901为出发菌株，通过URA 3基因的连续缺失，构建了尿嘧啶营养缺陷型突变株。首先，由于清酒酵母是二倍体，用pURA 38（Δ URA 3 SMR 1）破坏一个URA 3基因，并在SM（甲嘧磺隆）平板上分离杂合破坏物。用pURA 36（Δ URA 3）破坏另一个URA 3基因，并在FOA（5-氟-乳清酸）平板上分离纯合的URA 3破坏物，在该平板上只有ura 3突变体可以生长。还用pURA 36（.EPSILON.DURA3）进行直接URA 3基因破坏，并分离尿嘧啶营养缺陷型突变体。分离出四种类型的URA 3破坏物，其中两种不含细菌DNA。使用pTRP 14（Δ TRP 1 URA 3）通过基因破坏从一种URA 3破坏剂构建色氨酸营养缺陷型突变体。这种TRP 1破坏物也缺乏细菌DNA。实验室规模的清酒酿造使用的营养缺陷型突变体表明，这些菌株是非常有用的受体菌株的清酒酵母的分子育种。
Uracil auxotrophic mutants were constructed from the sake yeasts K-701 and K-901 by successive URA3 gene disruption. First, as sake yeast is diploid, one URA3 gene was disrupted with pURA38 (.DELTA.URA3 SMR1) and the heterozygous disruptant was isolated on an SM (sulfometuron methyl) plate. The other URA3 gene was disrupted with pURA36 (.DELTA.URA3) and homozygous URA3 disruptants were isolated on FOA (5-fluoro-orotic acid) plate on which only ura3 mutants can grow. Direct URA3 gene disruption with pURA36 (.EPSILON.DURA3) was also done and the uracil auxotrophic mutant was isolated. Four types of URA3 disruptants were isolated, two of which had no bacterial DNA. A tryptophan auxotrophic mutant was constructed from one of the URA3 disruptant using pTRP14(.DELTA.TRP1 URA3) by gene disruption. This TRP1 disruptant was also lacking bacterial DNA. Laboratory scale sake brewing using the auxotrophic mutants showed that these strains are very useful as recipient strains for molecular breeding of sake yeasts.