分枝杆菌噬菌体Chy2分离鉴定及生物学特性研究 .docVIP

分支杆菌噬菌体Chy2的分离及生物学特性(400016 重庆重庆医科大学附属第一医院呼吸科) 分离鉴定新分支杆菌噬菌体并其生物学特性。 以平板法从泥土中分离纯化噬菌体，紫外线诱导法鉴定溶原性，从噬菌斑大小，颗粒形态和限制性内切酶分析3方面比较Chy2、D29、TM4异同；以不同感染复数(MOI)扩增Chy2，测定最佳MOI；通过一步生长实验测出Chy2潜伏期、裂解期和裂解量；采用单斑法测定MOI的裂解谱；检测MOI在不同pH值培养基中裂解耻垢分支杆菌mc2155的能力。 成功分离纯化1株分支杆菌噬菌体，命名为Chy2，其噬菌斑圆形透明，紫外线诱导法处理宿主菌后不能形成噬菌斑，头部呈椭圆形，长尾，基因组核酸能被双链DNA内切酶EcoRⅠHind Ⅲ、BamH I及XbaI切开，大小约54 kb，与D29、TM4差异大；Chy2最佳MOI为9.58×10-5；Chy2一步生长周期为270 min，潜伏期210 min，裂解期60 min，裂解量为23；Chy2能裂解耻垢分支杆菌mc2155、结核菌标准株H37Rv、多数临床耐药株；在7H9固体培养基pH值为5.0和7.4时，Chy2均能裂解耻垢分支杆菌mc2155。 Chy2属长尾噬菌体科，基因组为双链DNA，宽噬谱广，是一种潜在的抗结核药物资源。 分支杆菌噬菌体；耻垢分支杆菌；结核分支杆菌；生物学特性；耐药结核病 Isolation and biological characteristics of Mycobacteriophage Chy2 Liu Ping, Wu Tingting, Peng Li, Guo Shuliang, Luo Yongai(Department of Respiratory Medicine, the First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, China) [Abstract] Objective To isolate and identify a new mycobacteriophage and research on the biological characteristics to explore its potential of anti-TB treatment. Methods The new mycobacteriophage was isolated and purified by single-plate method from the soil. The phage was induced by ultraviolet ray (UV) to identify its lytic or lysogenic capacity. Chy2, D29 and TM4 were compared by the plaque size, particle morphology and the restriction enzyme analysis. The phage was amplified by double-layer agar plate method in different multiplicity of infection to find the optimal MOI. One step growth experiment was done to find the latent period, burst period and burst size of the phage. The host range of the phage was examined by single-spot examination. The ability of the phage to crack Mycobacterium smegmatis mc2155 at different pH values was examined. Results A new mycobacteriophage was successfully isolated and purified, which was named Chy2. The plaques of Chy2 are transparent and their sizes are about 3mm in the diameter, The host bacteria induced by UV can not form plaques. Chy2 has a oval head，a long tail, base plate and tail fiber. Chy2