Yang C L, Liu Y H, Wang T C
Department of Molecular and Cellular Biology, Chang Gung College of Medicine and Technology, Kwei-San, Tao-Yuan, Taiwan.
Mutat Res. 1996 Jan 2;362(1):21-8.
The recO gene is required for the RecF pathway of recombination and the repair of DNA daughter-strand gaps in Escherichia coli. In this work, the structural portion of recO was synthesized by the polymerase chain reaction (PCR) and cloned onto expression vectors at their Nco1 fusion cloning site, to eliminate the presence of mRNA leader sequence. While the plasmid carrying a Ptac promoter failed to overproduce RecO, the plasmid carrying a T7O10 promoter overproduced RecO in large quantity, indicating that the natural recO may be overexpressed. An increase of intracellular RecO, which may be due to the increased recO gene copies or to the induction of RecO synthesis, increased the UV resistance of recA, recF, and ssb cells, but did not increase the UV resistance of uvrB, uvrB recF, uvrB recA and uvrB ssb cells. We suggest that an increase of intracellular RecO may allow some recombination-deficient cells to perform more excision repair, thus increasing the survival. The possible causes for RecO overproduction on excision repair, and for the differential expression of recO by the Ptac and T7 promoter plasmids are discussed.
recO基因是大肠杆菌中重组的RecF途径和DNA子链缺口修复所必需的。在这项工作中，recO的结构部分通过聚合酶链反应（PCR）合成，并克隆到表达载体的Nco1融合克隆位点上，以消除mRNA前导序列的存在。携带Ptac启动子的质粒未能过量产生RecO，而携带T7O10启动子的质粒大量过量产生RecO，这表明天然recO可能会被过度表达。细胞内RecO的增加，这可能是由于recO基因拷贝数增加或RecO合成的诱导，增加了recA、recF和ssb细胞的紫外线抗性，但没有增加uvrB、uvrB recF、uvrB recA和uvrB ssb细胞的紫外线抗性。我们认为细胞内RecO的增加可能使一些重组缺陷细胞能够进行更多的切除修复，从而提高存活率。讨论了RecO在切除修复上过量产生的可能原因，以及Ptac和T7启动子质粒对recO的差异表达。
