Waldman A S, Liskay R M
Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, Connecticut 06510.
Mol Cell Biol. 1988 Dec;8(12):5350-7. doi: 10.1128/mcb.8.12.5350-5357.1988.
Recombination between a 360-base-pair (bp) segment of a wild-type thymidine kinase gene (tk) from each of three different strains (F, MP, and 101) of herpes simplex virus type one and a complete herpes simplex virus type 1 (strain F) tk gene containing an 8-bp insertion mutation was studied. The pairs of tk sequences resided as closely linked repeats within the genome of mouse LTK- cells. The frequency of recombination between sequences exhibiting 232 bp of uninterrupted homology and containing no mismatches other than the insertion mutation was comparable to the frequency of recombination between two sequences exhibiting four additional nucleotide mismatches distributed in such a way to preserve the 232-bp stretch of contiguous homology. In contrast, the placement of only two single-nucleotide mismatches (in addition to the insertion mutation) in such a manner to reduce the longest uninterrupted homology to 134 bp resulted in a 20-fold reduction in recombination. We conclude that the rate of intrachromosomal recombination in mammalian cells is determined by the amount of uninterrupted homology available and not by the total number of mismatches within a given interval of DNA. Furthermore, efficient recombination appears to require between 134 and 232 bp of uninterrupted homology; single-nucleotide heterologies are most likely sufficient to disrupt the minimal efficient recombination target. We also observed that if recombination was allowed to initiate within sequences exhibiting perfect homology, the event could propagate through and terminate within adjacent sequences exhibiting 19% base pair mismatch. We interpret this to mean that heterology exerts most of its impact on early rather than late steps of intrachromosomal recombination in mammalian cells.
研究了来自单纯疱疹病毒1型三种不同毒株（F、MP和101）的野生型胸苷激酶基因（tk）的360个碱基对（bp）片段与含有8 bp插入突变的完整单纯疱疹病毒1型（F毒株）tk基因之间的重组。tk序列对作为紧密连锁的重复序列存在于小鼠LTK-细胞的基因组中。在具有232 bp不间断同源性且除插入突变外无其他错配的序列之间的重组频率，与在另外两个序列之间的重组频率相当，这另外两个序列存在四个额外的核苷酸错配，其分布方式可保持232 bp的连续同源性延伸。相比之下，仅以将最长不间断同源性减少到134 bp的方式放置两个单核苷酸错配（除插入突变外），导致重组减少了20倍。我们得出结论，哺乳动物细胞中染色体内部重组的速率由可用的不间断同源性的量决定，而不是由给定DNA区间内错配的总数决定。此外，有效的重组似乎需要134至232 bp的不间断同源性；单核苷酸异源序列很可能足以破坏最小有效重组靶点。我们还观察到，如果允许重组在表现出完美同源性的序列内启动，该事件可以传播并在相邻的表现出19%碱基对错配的序列内终止。我们将此解释为意味着异源性对哺乳动物细胞染色体内部重组的早期而非晚期步骤产生了大部分影响。
