Lee T C, Lin N T, Tseng Y H
Institute of Molecular Biology, Department of Botany, National Chung Hsing University, Taichung, Taiwan.
Biochem Biophys Res Commun. 1996 Apr 16;221(2):459-65. doi: 10.1006/bbrc.1996.0617.
A 1.8-kb NsiI-StuI fragment containing the recA gene of Xanthomonas campestris pv. campestris was cloned by a PCR-based approach and complementation of Escherichia coli HB 101. Sequence analysis of this fragment revealed an ORF (orf343) of 1,032 bp able to encode a protein of 343 amino acids with a calculated MW of 37,021 Da, a size similar to the values detected by in vitro system and Western blotting. It showed 69.6% identity to the E. coli RecA in amino acid sequence. Amino acid residues of the E coli RecA associated with functional activities are conserved in this Xc17 RecA. The recA mutant, L1, constructed by gene replacement, was sensitive to ultraviolet irradiation and methyl methanesulfonate, and deficient in homologous recombination.
通过基于PCR的方法并利用大肠杆菌HB 101进行互补,克隆了一个包含野油菜黄单胞菌野油菜致病变种recA基因的1.8 kb NsiI - StuI片段。对该片段的序列分析揭示了一个1032 bp的开放阅读框(orf343),其能够编码一个由343个氨基酸组成的蛋白质,计算分子量为37021 Da,该大小与体外系统和蛋白质免疫印迹检测到的值相似。其氨基酸序列与大肠杆菌RecA的一致性为69.6%。在这个Xc17 RecA中,与功能活性相关的大肠杆菌RecA氨基酸残基是保守的。通过基因置换构建的recA突变体L1对紫外线照射和甲磺酸甲酯敏感,并且在同源重组方面存在缺陷。