Broach J R, Guarascio V R, Jayaram M
Cell. 1982 May;29(1):227-34. doi: 10.1016/0092-8674(82)90107-6.
The multicopy yeast plasmid, 2mu circle, encodes a specialized recombination system. It contains two regions, each 599 bp in length, that are precise inverted repeats of each other and between which recombination occurs readily. In addition, this recombination requires the product of a 2mu circle gene, designated FLP. By examining the products of FLP-mediated recombination of plasmids containing single insertions within one of the repeated regions, we show that this recombination occurs only at a specific site within the repeat. This result was confirmed from analysis of the ability of plasmids containing various deletions within one of the repeated regions to serve as substrates for FLP-mediated recombination. These experiments limit the recombination site to a sequence of less than 65 bp. In addition, by mutational analysis of the recombination potential of a hybrid plasmid containing the entire 2mu circle genome, we have shown that FLP is only the 2mu circle gene necessary for this site-specific recombination. Finally, we describe a sensitive assay for recombination between the repeated sequences of 2mu circle; using it, we demonstrate that even in the absence of FLP gene product, recombination between the repeats occurs at a low but detectable level during meiosis.
多拷贝酵母质粒2μm环编码一种特殊的重组系统。它包含两个区域,每个区域长度为599 bp,它们是彼此精确的反向重复序列,并且在它们之间很容易发生重组。此外,这种重组需要一个2μm环基因的产物,命名为FLP。通过检查在其中一个重复区域内含有单个插入片段的质粒的FLP介导的重组产物,我们发现这种重组仅发生在重复序列内的一个特定位点。通过分析在其中一个重复区域内含有各种缺失的质粒作为FLP介导的重组底物的能力,证实了这一结果。这些实验将重组位点限制在小于65 bp的序列内。此外,通过对含有整个2μm环基因组的杂交质粒的重组潜力进行突变分析,我们表明FLP只是这种位点特异性重组所必需的2μm环基因。最后,我们描述了一种用于检测2μm环重复序列之间重组的灵敏检测方法;使用该方法,我们证明即使在没有FLP基因产物的情况下,重复序列之间的重组在减数分裂期间也会以低但可检测的水平发生。