替代失活法构建变形链球菌LuxS基因缺陷株  

Construction of Streptococcus mutans LuxS gene deletion mutants by long flanking homology polymerase chain reaction

作　　者：李月恒[1] 陈惠珍[1] 周智[1] 陈娇[1] 李锐[1] 尹一兵[2] 张雪梅[2] 

机构地区：[1]重庆医科大学口腔医学院,重庆400015 [2]重庆医科大学检验系,重庆400015

出　　处：《中国微生态学杂志》2011年第3期201-203,共3页Chinese Journal of Microecology

基　　金：重庆市自然科学基金(CSTC2007BB5272);重庆市卫生局科研课题(2008-2-249);重庆市卫生局中医药科研计划项目(2010-2-64);重庆市教委科研课题(KJ060317)

摘　　要：目的 LuxS基因是变形链球菌生物膜早期形成过程中的关键基因,构建该基因的缺陷菌。方法采用长臂同源多聚酶链反应(LFH-PCR)方法构建含红霉素耐药基因片段的LuxS基因上、下游同源序列的连接片段,转化到变形链球菌中,在红霉素的平板上筛选缺陷菌株,并采用PCR鉴定。结果对变形链球菌LuxS基因缺陷菌株进行PCR和DNA序列测定分析证实构建成功。结论成功构建出变形链球菌LuxS基因的缺陷菌株,为后期针对变形链球菌LuxS基因的相关研究奠定基础。Objective To construct LuxS deletion mutant of Streptococcus mutans and get the capsule deficient strain. Method Long flanking homology polymerase chain reaction（LFH-PCR） was introduced to generate a gene disruption construct consisting of Emr cassette with long flanking homology regions to the target gene.The electroporation competence of Streptococcus mutans was then transformed with this PCR product.Then the positive transformants were counted on selective agar containing erythromycin and identified by PCR. Result Identification by PCR and sequencing confirmed the validity of the LuxS deletion mutant of Streptococcus mutans. Conclusion The successful construction of the LuxS deletion mutant can be used in further functional genome research.

关 键 词：变形链球菌 基因缺陷 PCR鉴定 失活 多聚酶链反应 同源序列 DNA序列 S基因 

分 类 号：Q78[生物学—分子生物学]