Gómez-González Belén, Ruiz José F, Aguilera Andrés
Centro Andaluz de Biología Molecular y Medicina Regenerativa, Universidad de Sevilla-CSIC, Sevilla, Spain.
Methods Mol Biol. 2011;745:151-72. doi: 10.1007/978-1-61779-129-1_10.
Many systems have been developed for the study of mitotic homologous recombination (HR) in the yeast Saccharomyces cerevisiae at both genetic and molecular levels. Such systems are of great use for the analysis of different features of HR as well as of the effect of mutations, transcription, etc., on HR. Here we describe a selection of plasmid- and chromosome-borne DNA repeat assays, as well as plasmid-chromosome recombination systems, which are useful for the analysis of spontaneous and DSB-induced recombination. They can easily be used in diploid and, most importantly, in haploid yeast cells, which is a great advantage to analyze the effect of recessive mutations on HR. Such systems were designed for the analysis of a number of different HR features, which include the frequency and length of the gene conversion events, the frequency of reciprocal exchanges, the proportion of gene conversion versus reciprocal exchange, or the molecular analysis of sister chromatid exchange.
已经开发了许多系统,用于在遗传和分子水平上研究酵母酿酒酵母中的有丝分裂同源重组(HR)。此类系统对于分析HR的不同特征以及突变、转录等对HR的影响非常有用。在这里,我们描述了一系列基于质粒和染色体的DNA重复检测方法,以及质粒-染色体重组系统,这些方法对于分析自发和双链断裂(DSB)诱导的重组很有用。它们可以很容易地用于二倍体酵母细胞,最重要的是,也可用于单倍体酵母细胞,这对于分析隐性突变对HR的影响是一个很大的优势。此类系统旨在分析许多不同的HR特征,包括基因转换事件的频率和长度、相互交换的频率、基因转换与相互交换的比例,或姐妹染色单体交换的分子分析。