多药耐药草绿色链球菌LuxS基因同源重组质粒的构建  

Construction of Streptococcus Viridans LuxS Gene Homologous Recombinant Plasmid

作　　者：陆笑[1] 刘少娟[1] 章锦才[2] 刘芹[1] 林珊[1] 彭湘明[1] 

机构地区：[1]广州市红十字会医院口腔科,广东广州510220 [2]南方医科大学附属口腔医院.广东省口腔医院,广东广州510280

出　　处：《口腔医学研究》2015年第10期984-987,共4页Journal of Oral Science Research

基　　金：广州市卫生局基金资助项目(编号:2060402)

摘　　要：目的:构建一个含有红霉素抗性基因和多药耐药草绿色链球菌LuxS基因上下游区同源序列的重组质粒,为后续进行LuxS基因敲除实验做准备。方法:设计合成引物,分别以质粒PMG36E、草绿色链球菌DNA为模板,应用聚合酶链反应(PCR)扩增得到红霉素抗性基因DNA片断和LuxS基因上下游序列,经双酶切反应插入pUC19质粒的多克隆酶切位点中,转化大肠杆菌的感受态中,氨苄青霉和红霉素培养基筛选。结果:红霉素抗性基因和LuxS基因两侧同源序列成功连入到Puc19质粒相应酶切位点,PCR凝胶电泳、测序结果正确。结论:成功构建多药耐药草绿色链球菌LuxS基因敲除同源重组质粒,为进一步构建LuxS突变株打下基础。Objective：To construct a recombination plasmid containing erythromycin resistance gene and the upstream and downstream homologous sequence of LuxS gene of streptococcus viridans（S.viridans）with multidrug resistance.Methods：Erythromycin resistance gene and the upstream and downstream homologous sequence of LuxS were cloned respectively by using plasmid PMG36 Eand DNA of S.viridans template.Then the genes were ligated into multiple cloning site（MCS）of vector pUC19 and transformed into competent E.coli.Finally,the transformants which were resistant to erythromycin and ampicillin were selected.Results：Erythromycin resistance gene and the upstream and downstream homologous sequence of LuxS were successfully ligated into enzyme digestion site of vector pUC19 accurately.The PCR agarose gel electrophoretic analysis and the sequencing results were correct.Conclusion：The S.viridans recombinant plasmid with LuxS geneknock-out was constructed and could be used for constructing S.viridans LuxS mutans in future.

关 键 词：草绿色链球菌 LUXS基因 pUC19质粒 PMG36E质粒 

分 类 号：R780.2[医药卫生—口腔医学]