您的位置:首页  > 论文页面

采用PCR连接诱变技术构建变异链球菌dexA基因缺陷株

发表时间:2016-03-15  浏览量:1485  下载量:594
全部作者: 阳燕,洪潇,胡涛,杨英明
作者单位: 四川大学华西口腔医(学)院,口腔疾病研究国家重点实验室
摘 要: 变异链球菌dexA基因合成的右旋糖酐酶(dextranase,DexA)通过水解水溶性葡聚糖(water soluble glucan,WSG)内部的α-1,6糖苷键,为细菌提供糖源,在胞外多糖的代谢和生物膜的结构中有着重要影响。为进一步研究变异链球菌dexA基因的功能,本研究拟通过PCR连接诱变技术构建变异链球菌dexA基因敲除突变株。通过PCR连接诱变技术,构建变异链球菌dexA基因敲除株SmudexA;PCR技术和凝胶电泳验证目的基因的完整转入和dexA基因的敲除;荧光定量实时PCR(real-time PCR,RT-PCR)技术进一步验证SmudexA株dexA基因的表达缺失;核苷酸序列分析验证dexA基因的正确敲除。结果表明,本研究成功构建并鉴定了变异链球菌UA159的dexA基因缺陷株,为后续实验奠定了基础。
关 键 词: 口腔医学;变异链球菌;dexA基因;PCR连接诱变;突变株构建
Title: Construction of Streptococcus mutans dexA gene deletion strain by PCR ligation mutagenesis
Author: YANG Yan, HONG Xiao, HU Tao, YANG Yingming
Organization: State Key Laboratory of Oral Diseases, West China School/Hospital of Stomatology, Sichuan University
Abstract: Dextranase (DexA) synthesized by dexA gene of Streptococcus mutans has an important influence on the metabolism of extracellular polysaccharides and the structure of biofilm, via hydrolysis of α-1,6glycosides bond inside water soluble glucan (WSG). In the purpose of studying the further function of dexA gene, this paper was to construct the dexA gene knockout mutants of S. mutans by PCR ligation mutagenesis. In this study, the dexA gene deletion strain of S. mutans (SmudexA) was constructed with PCR ligation mutagenesis. The successful construction of the mutation was verified by PCR, gel electrophoresis. Fluorescence quantitative real-time PCR (RT-PCR) showed dexA gene expression deficiency and nucleotide sequence analysis confirmed dexA gene was knocked out. We have successfully constructed and authenticated the mutant strains of S. mutans for dexA gene deletion which lay foundation for the further subsequent exploration.
Key words: stomatology; Streptococcus mutans; dexA gene; PCR ligation mutagenesis; mutant construction
发表期数: 2016年3月第5期
引用格式: 阳燕,洪潇,胡涛,等. 采用PCR连接诱变技术构建变异链球菌dexA基因缺陷株[J]. 中国科技论文在线精品论文,2016,9(5):443-448.
 
2 评论数 0
暂无评论
友情链接