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首页> 外文期刊>Genes & Genetic Systems >Isolation of thermotolerant mutants by using proofreading-deficient DNA polymerase delta as an effective mutator in Saccharomyces cerevisiae
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Isolation of thermotolerant mutants by using proofreading-deficient DNA polymerase delta as an effective mutator in Saccharomyces cerevisiae

机译：使用校对缺陷的DNA聚合酶δ作为酿酒酵母中有效的突变体，分离耐热突变体

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摘要

Eukaryotic DNA polymerases delta and epsilon, both of which are required for chromosomal DNA replication, contain proofreading 3'-->5'exonuclease activity. DNA polymerases lacking proofreading activity act as strong mutators. Here we report isolation of thermotolerant mutants by using a proofreading-deficient DNA polymerase delta variant encoded by pol3-01 in the yeast Saccharomyces cerevisiae. The parental pol3-01 strain grew only poorly at temperatures higher than 38 degrees C. By stepwise elevation of the incubation temperature, thermotolerant mutants that could proliferate at 40 degrees C were successfully obtained; however, no such mutants were isolated with the isogenic POL3 strain. The recessive hot1-1 mutation was defined by genetic analysis of a weak thermotolerant mutant. Strong thermotolerance to 40 degrees C was attained by multiple mutations, at least one of which was recessive. These results indicate that a proofreading-deficient DNA delta polymerase variant is an effective mutator for obtaining yeast mutants that have gained useful characteristics, such as the ability to proliferate in harsh environments.
机译：染色体DNA复制所需的真核DNA聚合酶δ和epsilon均具有校对的3'-> 5'核酸外切酶活性。缺乏校对活性的DNA聚合酶可作为强突变体。在这里我们报告通过使用酿酒酵母中的pol3-01编码的校对缺陷型DNA聚合酶δ变体来分离耐热突变体。亲代pol3-01菌株在高于38摄氏度的温度下仅生长缓慢。通过逐步升高孵育温度，成功获得了可以在40摄氏度下增殖的耐热性突变体。但是，没有用等基因POL3菌株分离出这样的突变体。隐性hot1-1突变是通过对耐热性较弱的突变体进行遗传分析确定的。通过多次突变获得了高达40摄氏度的强耐热性，其中至少一种是隐性的。这些结果表明，校对缺陷的DNA三角洲聚合酶变异体是一种有效的突变体，可用于获得具有有用特性（例如在恶劣环境中增殖的能力）的酵母突变体。

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