校对
生物
DNA聚合酶
核酸外切酶
酿酒酵母
突变体
遗传学
DNA聚合酶Ⅱ
DNA复制
聚合酶
DNA聚合酶Ⅰ
DNA
DNA聚合酶δ
DNA钳
突变
分子生物学
酵母
聚合酶链反应
基因
逆转录酶
作者
Chikashi Shimoda,Akiko Itadani,Akio Sugino,Mitsuru Furusawa
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.
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