Gracheva L M, Evstiukhina T A, Koval'tsova S V, Korolev V G
Genetika. 1992 May;28(5):56-65.
A group of Saccharomyces cerevisiae mutants deficient in repair of induced premutation lesions (him mutants) were previously isolated in our laboratory. Recessive him1 mutant had enhanced level of spontaneous and induced mutagenesis as well as specific altered mitotic conversion. This HIM1 gene was supposed to be involved in the process of mismatch correction of heteroduplexes. In this paper the correction efficiency of in vitro constructed heteroduplex DNA in wild-type cells and him1 mutant was studied. In the former cells heteroduplex DNA was repaired highly efficiently (about 90%), this repair efficiency being reduced in him cells approx. two times as compared with the wild-type cells. Molecular cloning of yeast chromosomal DNA fragments containing HIM1 gene was carried out. The clones were selected from the bank of yeast DNA fragments by complementing him1-1 mutation which enhances conversion frequency in ADE2 gene. One of the DNA fragments was analysed by restriction endonuclease digestion and shown to contain an insert of 6 Kb. Chromosomal integrants were obtained by homologous recombination between the plasmid and chromosomal gene him1.
我们实验室先前分离出了一组酿酒酵母突变体,这些突变体在诱导前突变损伤修复方面存在缺陷(him突变体)。隐性him1突变体具有增强的自发和诱导诱变水平以及特定的有丝分裂转换改变。推测该HIM1基因参与异源双链体错配校正过程。本文研究了体外构建的异源双链体DNA在野生型细胞和him1突变体中的校正效率。在野生型细胞中,异源双链体DNA得到高效修复(约90%),而在him细胞中,这种修复效率与野生型细胞相比降低了约两倍。进行了包含HIM1基因的酵母染色体DNA片段的分子克隆。通过补充him1 - 1突变(该突变提高了ADE2基因中的转换频率)从酵母DNA片段文库中筛选出这些克隆。对其中一个DNA片段进行了限制性内切酶消化分析,结果显示其含有一个6 kb的插入片段。通过质粒与染色体基因him1之间的同源重组获得了染色体整合体。