军事医学2017，Vol.41Issue(10)：800-804,5.DOI:10.7644/j.issn.1674-9960.2017.10.004
鼠疫耶尔森菌中长度在100nt以内的小RNA缺失株及过表达菌株的构建
Construction of sRNA-deficient and-overexpressing strains of Yersinia pestis
摘要
Abstract
Objective To construct small RNA deletion and overexpression strains with a length of less than 100 nt in Yersinia pestis.Methods Deletion mutants of the target sRNAs were constructed by increasing the length of homologous regions.Meanwhile, the high copy plasmid pBAD/HisA was modified into an inducible transcriptional vector as an sRNA-overexpression plasmid by using QuikChange lightning site-directed mutagenesis kit .The presence , size, and transcription-al initiation sites of the indicated sRNA were predicted by transcriptome sequencing , primer extension , and previous stud-ies.The full-length DNA fragments of target sRNAs were transformed into the transcriptional vector .The overexpressing strains of sRNAs were identified by Northern Blot .Results and Conclusion Four sRNAs deletion mutants of sR01, sR02, sR03 and HmsA and three sRNAs overexpression mutants MicF , HmsA and CpxQ were successfully constructed .A method of construction of sRNA deficient and overexpressing strains of Y.pestis has been quickly and efficiently established by λ-Red homologous recombination technology and QuikChange ? lightning site-directed mutagenesis kit.关键词
鼠疫菌/小RNA/λ-Red同源重组/过表达Key words
Yersinia pestis/small RNA/λ-Red homologous recombination/overexpression分类
医药卫生引用本文复制引用
高肖芳,刘子中,李文亮,王红朵,杨瑞馥,韩延平..鼠疫耶尔森菌中长度在100nt以内的小RNA缺失株及过表达菌株的构建[J].军事医学,2017,41(10):800-804,5.基金项目
国家自然科学基金资助项目(31430006) （31430006）
国家重点基础研究发展计划资助项目(2014CB744405) （2014CB744405）
