摘要
目的筛选能感染结核分枝杆菌的溶源性噬菌体,了解其生物学及遗传学信息。方法从土壤中分离纯化噬菌体;电镜观察噬菌体形态;单斑法测定宿主谱;提取噬菌体DNA,鸟枪法随机测序。采用DNAStar软件包分析基因组成分,Glimmer3.0、GeneMark软件预测基因功能,同时做共线性分析。分离溶源菌,通过紫外线诱导试验、超免反应验证噬菌体溶源性。结果分离到一株新的噬菌体Chy5,其头部直径(61.5±1.4)nm,尾部长度(114.1±2.1)nm,为长尾噬菌体,可感染敏感及耐药结核菌株。Chy5基因组全长51214 bp,G+C含量63.60%,与噬菌体D29基因组最相似,有88个推定基因,含有编码整合酶和阻遏蛋白的基因以及attP位点。经紫外线诱导Chy5溶源菌可形成噬菌斑,Chy5与其溶源菌共培养会发生超免反应。结论分离到一株新的能感染敏感及耐药结核菌的溶源性噬菌体Chy5,其基因组与噬菌体D29相似,有望应用于构建荧光报告噬菌体。
To select the phage infecting Mycobacterium tuberculosis,and understand its biological characteristics and genetic information,mycobacteriophage was isolated and purified from soil.The morphological characteristics were observed by electron-microscopy,the host range was then examined by single-spot examination.Mycobacteriophage DNA was extracted.Shutgun sequences were assembled,then the gaps were filled with PCR fragments.DNAStar software Packages was used to analyze the general characteristic of genome,Glimmer 3.0 and GeneMark were applied to predict the function of the coding gene.In addition,the collinearity analysis were conducted.Lysogens was isolated to conduct an Ultraviolet induction test and super infection immunity.In results,a novel mycobacteriophage Chy5 was isolated.The head diameter was(61.5±1.4 nm)and the tail length was(114.1±2.1 nm).It could infect different clinical isolates of Mycobacterium tuberculosis,which was classified into Siphoviridae.The full length of Chy5 genome was 51214 bp with a G+C content of 63.60%.Chy5 genome was most similar with D29,which were 88 predicted genes encoding integrase and repressor as well as attP site.The Chy5 lysogen could release free phage particles to form plaques after UV induction.Chy5 could not clear lawn of Chy5 lysogens.So Chy5 is a novel lysogenic mycobacteriophage,which could lyse mycobacterium tuberculosis and be applied to construct Luciferase reporter phages(LRP).
作者
江莉莎
刘平
邬亭亭
郭述良
JIANG Li-sha;LIU Ping;WU Ting-ting;GUO Shu-liang(Department of Respiratory Medicine,The First Branch,The First Affiliated Hospital of Chongqing Medical University,Chongqing 400010,China;Department of Respiratory Medicine,The First Affiliated Hospital of Chongqing Medical University,Chongqing 400016,China;Department of Respiratory Medicine,People’s Hospital of Changshou District,Chongqing 401220,China;Department of Respiratory Medicine,The Third People’s Hospital of Chengdu,Chengdu 610031,China)
出处
《中国人兽共患病学报》
CAS
CSCD
北大核心
2020年第6期462-468,共7页
Chinese Journal of Zoonoses
基金
国家“十二五”科技重大专项课题(No.2012ZX10003009)资助。
