Fishel R A, Siegel E C, Kolodner R
J Mol Biol. 1986 Mar 20;188(2):147-57. doi: 10.1016/0022-2836(86)90300-1.
We have constructed heteroduplex plasmid DNA that is similar in structure to the heteroduplex DNA expected to be produced during genetic recombination of plasmids, and studied its repair after transformation into different Escherichia coli strains. The heteroduplex DNA was constructed using two different parental plasmids, each of which contained a different ten-nucleotide insertion mutation. The effect of different defined states of dam-methylation on repair was also examined. We found that heteroduplex DNA repair occurred prior to the replication of the substrate DNA 60 to 80% of the time, regardless of the state of DNA methylation. Most excision/synthesis tracts covered two markers separated by 1243 base-pairs, and this process has been termed co-repair. The most efficient co-repair pathway was the Dam-instructed repair pathway that required the mutH, mutL, mutS and uvrD gene products and preferentially used the methylated strand as the template for DNA synthesis. If there was no methylation asymmetry, mismatch nucleotide repair occurred with a similar frequency; however, no strand bias was observed. Co-repair of symmetrically methylated heteroduplex DNA required the mutS and uvrD gene products, while repair of unmethylated heteroduplex DNA also required the mutL and mutH gene products.
我们构建了结构与质粒基因重组过程中预期产生的异源双链 DNA 相似的异源双链质粒 DNA，并研究了其转化到不同大肠杆菌菌株后的修复情况。异源双链 DNA 是使用两种不同的亲本质粒构建的，每种质粒都含有不同的十个核苷酸插入突变。还研究了不同定义状态的 dam 甲基化对修复的影响。我们发现，无论 DNA 甲基化状态如何，60% 至 80% 的情况下异源双链 DNA 修复发生在底物 DNA 复制之前。大多数切除/合成片段覆盖了由 1243 个碱基对隔开的两个标记，这个过程被称为协同修复。最有效的协同修复途径是 Dam 指导的修复途径，该途径需要 mutH、mutL、mutS 和 uvrD 基因产物，并优先使用甲基化链作为 DNA 合成的模板。如果没有甲基化不对称性，错配核苷酸修复以相似的频率发生；然而，未观察到链偏向。对称甲基化异源双链 DNA 的协同修复需要 mutS 和 uvrD 基因产物，而未甲基化异源双链 DNA 的修复也需要 mutL 和 mutH 基因产物。
