结核分枝杆菌rv0199基因在耻垢分枝杆菌中的表达及检测

中国畜牧兽医

结核分枝杆菌rv0199基因在耻垢分枝杆菌中的表达及检测

崔保亮，陈利苹，张娈娈，李华芳，刘思国

（中国农业科学院哈尔滨兽医研究所，兽医生物技术国家重点实验室动物细菌病研究室，黑龙江哈尔滨 150001）

收稿日期:2014-03-14 出版日期:2014-07-20 发布日期:2014-08-21
通讯作者: 刘思国。E-mail：siguo_liu@hvri.ac.cn
作者简介:崔保亮（1986—），男，河北人，硕士生，研究方向：动物结核病。
基金资助:
国家自然科学基金（31201920、31272538）；兽医生物技术国家重点实验室自主研究课题（SKLVBP201307）。

Expression and Detection of Mycobacterium tuberculosis rv0199 Gene in Mycobacterium smegmatis

CUI Bao-liang, CHEN Li-ping, ZHANG Luan-luan, LI Hua-fang, LIU Si-guo

(Division of Animal Bacterial Diseases, State Key Laboratory of Veterinary Biotechnology，Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China)

Received:2014-03-14 Online:2014-07-20 Published:2014-08-21

摘要/Abstract

摘要： 为研究结核分枝杆菌（M.tb）rv0199基因在该病原致病性中可能发挥的作用，本研究克隆了rv0199基因，在耻垢分枝杆菌（Ms）中异源表达，并对其表达进行检测。将大肠杆菌—分枝杆菌穿梭表达载体pMV261进行改造，向载体中引入常用的6His和Strep Ⅱ重组蛋白标签，命名为pMV262。使用Ms/pMV262表达系统对rv0199基因进行超表达，用抗6His标签抗体和抗Strep Ⅱ标签抗体均能特异的检测到重组蛋白。本试验改造的pMV262穿梭表达载体可很方便的检测M.tb的基因在Ms中的异源表达，不用制备针对蛋白的多克隆抗体或单克隆抗体。构建的重组菌Ms/262-99为进一步研究rv0199基因的功能提供了材料，奠定了基础。

关键词: font-size: 10.5pt; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-font-kerning: 1.0pt; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA">结核分枝杆菌; rv0199基因; pMV261; pMV262; 耻垢分枝杆菌

Abstract: To study the role of Mycobacterium tuberculosis(M.tb) rv0199 gene may play in the pathogenicity of the pathogen, rv0199 gene was cloned, the protein was heterologous expressed in M.smegmatis(Ms) and its expression was detected. The E.coli-Mycobacteria shuttle vector pMV261 was improved by introducing the commonly used 6His and Strep Ⅱ recombinant protein tags, the improved vector was named as pMV262. The rv0199 gene was over-expressed in Ms/pMV262 expression system, the recombinant protein was specifically detected by the anti-6His tag antibody and anti-Strep Ⅱ tag antibody. The heterologous expression of M.tb gene in Ms could be easily detected by the shuttle vector pMV262 constructed in this study, without preparing monoclonal or polyclonal antibody against the protein. The recombinant bacteria Ms/262-99 provided material and laid foundation for further study of the potential function of rv0199 gene.

Key words: font-size: 10.5pt; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-font-kerning: 1.0pt; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA" lang="EN-US">Mycobacterium tuberculosisfont-size: 10.5pt; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-font-kerning: 1.0pt; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA" lang="EN-US">; rv0199 gene; pMV261; pMV262; M.smegmatis

崔保亮，陈利苹，张娈娈，李华芳，刘思国. 结核分枝杆菌rv0199基因在耻垢分枝杆菌中的表达及检测[J]. 中国畜牧兽医.

CUI Bao-liang, CHEN Li-ping, ZHANG Luan-luan, LI Hua-fang, LIU Si-guo. Expression and Detection of Mycobacterium tuberculosis rv0199 Gene in Mycobacterium smegmatis[J]. China Animal Husbandry & Veterinary Medicine.

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