幽门螺杆菌ArsRS双组分系统基因突变株的构建

Construction of Mutants of Helicobacter pylori with Two-component System ArsRS

目的:构建幽门螺杆菌ArsRS双组分系统基因突变株。方法:以幽门螺杆菌标准菌株11637为模板,PCR扩增ArsS、ArsR基因;扩增产物分别克隆于载体pID700A1中,化学法转化感受态大肠杆菌E.coli DH5α,使其在细菌内同源重组,双酶切筛选得到重组载体,然后将重组载体经电转化法转化幽门螺杆菌标准菌株,获得幽门螺杆菌ArsS、ArsR突变株。结果:PCR获得了360bp、350bpArsS、ArsR基因,构建了插入ArsS、ArsR基因的重组载体pID700A1-ArsS、pID700A1-ArsR。经双酶切分析显示,所得条带与设计结果完全一致。PCR方法扩增突变株结果显示ArsS、ArsR基因已缺失。结论:成功构建了幽门螺杆菌ArsS、ArsR突变株,为ArsS、ArsR基因的检测和新型药物靶点以及疫苗的研究奠定基础。

Objective： To construct the gene mutants of Helicobacter pylori with two-component system ArsRS. Methods： ArsS and ArsR genes were amplified by PCR from total DNA of standard Helicobacter pylori strains and inserted into vector plD700A1. The recombinant vectors were transformed into E. coli DH5α and identified by restriction enzyme digestion and DNA sequencing. The correct recombinant vectors plD700A1-ArsS and pID700A1-ArsR were electroporated into standard Helicobacter pylori strains to get the mutants. Results： Recombinant gene vectors plD700A1-ArsS and plD700A1-ArsR which were inserted with ArsS and ArsR have been successfully constructed. The recombinant vectors were generated by homologous recombination in E. coli DH5α and the result was confirmed by enzyme digestion. The gene mutants of ArsS and ArsR were successfully constructed. Conclusions： This study has successfully constructed the Helicobacter pylori two-component system ArsRS gene mutants, and has made groundwork for ArsS and ArsR gene de- tections, new drug targets and the vaccine research.