变形链球菌耐氟菌株gtfB基因失活菌株的构建  

Construction of Streptococcus Mutans Fluoride-resistant Strain with GtfB Gene Inactive

作　　者：李文月[1] 高爽[1] 张志鹰[1] 张红[1] 超博 武玉凤[1] 李贺[1] 王春萌 张志民[1] LI Wen- yue;GAO Shuang;ZHANG Zhi ying;ZHANG Hong;CHAO Bo;WU Yu -- feng;LI He;WANG Chun - meng;ZHANG Zhi -- min(Department of Endoclontics School of Stomatology , Jilin University, Changchun 130021, China.)

机构地区：[1]吉林大学口腔医院牙体牙髓科,吉林长春130021

出　　处：《口腔医学研究》2018年第5期490-494,共5页Journal of Oral Science Research

摘　　要：目的:构建变形链球菌(S.mutans)耐氟菌株UA159-FR中gtfB基因失活株,为研究耐氟菌株gtfB基因的功能奠定基础。方法:培养UA159-FR并以其为模板扩增gtfB基因上游、下游同源臂片段,以质粒pEGFP-N1为模板扩增kan基因;通过重叠延伸聚合酶链反应法(overlap extension polymerase chain reaction,OE-PCR)获取上述3个片段的同源重组片段;与pEASY-Blunt Cloning Vector连接形成重组质粒后,进行PCR鉴定和测序鉴定;将重组片段电转化入UA159-FR感受态细胞中得到gtfB基因失活菌株,并进行PCR鉴定。结果:经PCR鉴定和测序鉴定,含有gtfB基因重组片段的重组质粒构建成功;经PCR鉴定,UA159-FR的gtfB基因失活株构建成功。结论:成功构建了含有gtfB基因重组片段的重组质粒和S.mutans耐氟菌株UA159-FR的gtfB基因失活株,可用于gtfB基因功能的研究。Objective：To construct Streptococcus mutans（S.mutans）fluoride-resistant strain UA159-FR with gtfB gene inactive to investigate the function of gtfB gene in fluoride-resistant strain.Methods：The S.mutans fluoride-resistant strain UA159-FR was cultured and used as a template to amplify the upstream and downstream homologous arm fragments of the gtfB gene.The kan gene was amplified by using the plasmid pEGFP-N1 as a template.Homologous recombination fragments of three fragments were obtained by overlap extension polymerase chain reaction（OE-PCR）and ligated with pEASY-Blunt Cloning Vector to form recombinant plasmids.The recombinant plasmids were identified by PCR and sequencing.The recombinant fragment was electrotransformed into UA159-FR competent cells to obtain the inactivated strain and identified by PCR.Results：The recombinant plasmids were successfully constructed via being identified by PCR and sequencing.It was identified by PCR that UA159-FR with gtfB gene inactive was obtained.Conclusion：The recombinant fragments of gtfB gene of S.mutans with its recombinant plasmids and the S.mutans fluoride-resistant strain UA159-FR with gtfB gene inactive were successfully constructed and could be used to study the function of gtfB gene.

关 键 词：变形链球菌耐氟菌株 gtfB基因 重叠延伸聚合酶链反应 同源重组 基因失活 

分 类 号：R780.2[医药卫生—口腔医学]