Department of Biophysics and Medical Physics and Donner Laboratory, University of California, 94720, Berkeley, CA, USA.
Curr Genet. 1983 Apr;7(2):85-92. doi: 10.1007/BF00365631.
The RAD52 gene of Saccharomyces cerevisiae has previously been shown to be involved in both recombination and DNA repair. Here we report on the cloning of this gene. A plasmid containing a 5.9 kb yeast DNA fragment inserted into the BamH1 site of the YEp13 vector has been isolated and shown to complement the X-ray sensitive phenotype of the rad52-1 mutation. The rad52-1 cells containing the plasmid form larger colonies than similar cells having lost the plasmid. This plasmid has been shown not to complement either the U.V. sensitivity or the recombination defect of the E. coli recA mutation. From the insert various fragments have been subcloned into the YRp7 and YIp5 vectors. Integration events of two of the subclones have been genetically mapped to the chromosomal location of RAD52, indicating that the structural gene has been cloned. A 1.97 kb BamH1 fragment subcloned into YRp7 in one orientation complements the rad52-1 mutation, while the same fragment in the opposite orientation fails to complement. Various other subclones indicate that a BglII site, within the BamH1 fragment, is in the RAD52 gene. This BglII site has been deleted by Sl-nuclease digestion and the resulting deletion inactivates the RAD52 gene. BAL31 deletions from one end of a 1.9 kb Sal1-BamH1 fragment have been isolated; up to 0.9 kb can be deleted without loss of RAD52 activity, indicating that the RAD52 gene is approximately 1 kb or less in length.
酿酒酵母的 RAD52 基因先前已被证明参与重组和 DNA 修复。在这里，我们报告了该基因的克隆。已经分离出并证明含有插入 YEp13 载体 BamH1 位点的 5.9 kb 酵母 DNA 片段的质粒可弥补 rad52-1 突变的 X 射线敏感表型。含有质粒的 rad52-1 细胞比失去质粒的类似细胞形成更大的菌落。该质粒未证明可弥补大肠杆菌 recA 突变的 U.V. 敏感性或重组缺陷。从插入物中，已经将各种片段亚克隆到 YRp7 和 YIp5 载体中。两个亚克隆的整合事件已在遗传上被映射到 RAD52 的染色体位置，表明结构基因已被克隆。一个定向亚克隆到 YRp7 中的 1.97 kb BamH1 片段可弥补 rad52-1 突变，而相同的片段反向则不能弥补。其他各种亚克隆表明，BamH1 片段内的 BglII 位点位于 RAD52 基因内。该 BglII 位点已被 Sl-核酸酶消化删除，导致 RAD52 基因失活。从 1.9 kb Sal1-BamH1 片段的一个末端分离出 BAL31 缺失；在不失去 RAD52 活性的情况下可以删除高达 0.9 kb，表明 RAD52 基因的长度约为 1 kb 或更短。
