[1]黄丹丹,鲍朗#,杨晓玲,等.结核分枝杆菌MycP1蛋白的原核表达及其对巨噬细胞的毒性作用*[J].郑州大学学报(医学版),2012,(04):438.
 HUANG Dandan,BAO Lang,YANG Xiaoling,et al.Prokaryotic expression of MycP1 of Mycobacterium tuberculosis and its cytotoxicity to macrophages[J].JOURNAL OF ZHENGZHOU UNIVERSITY(MEDICAL SCIENCES),2012,(04):438.
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结核分枝杆菌MycP1蛋白的原核表达及其对巨噬细胞 的毒性作用*
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《郑州大学学报(医学版)》[ISSN:1671-6825/CN:41-1340/R]

卷:
期数:
2012年04期
页码:
438
栏目:
论著
出版日期:
2012-07-20

文章信息/Info

Title:
Prokaryotic expression of MycP1 of Mycobacterium tuberculosis and its cytotoxicity to macrophages
作者:
黄丹丹鲍朗#杨晓玲邓仪昊廖丹
四川大学华西基础医学与法医学院感染免疫研究室 成都 610041
Author(s):
HUANG DandanBAO LangYANG XiaolingDENG YihaoLIAO Dan
Research Unit of Infection and Immunity, West China School of Preclinical and Forensic Medicine,Sichuan University,Chengdu 610041
关键词:
结核分枝杆菌MycP1巨噬细胞细胞毒性
Keywords:
Mycobacterium tuberculosisMycP1macrophagecytotoxicity
分类号:
R378.91
摘要:
目的:构建结核分枝杆菌MycP1蛋白原核表达系统。方法:构建pGEX4T1Rv3883c重组质粒,转化大肠杆菌,经IPTG诱导表达,采用SDSPAGE和Western blot法检测目标蛋白的表达情况。目标蛋白经亲和层析纯化后,以不同质量浓度MycP1作用于小鼠巨噬细胞ANA1, 48 h后XTT法检测活细胞数,同时测定细胞上清液中乳酸脱氢酶(LDH)活力,以评价其细胞毒性。结果:成功构建pGEX4T1Rv3883c质粒,该质粒能在大肠杆菌中表达,经鉴定,表达的融合蛋白为目标蛋白MycP1。该融合蛋白作用后可使ANA1活细胞数下降(F=34.327,P<0.001),培养上清液中LDH活力升高(F=336.720,P<0.001)。结论:成功构建了MycP1原核表达系统,目标蛋白对巨噬细胞有一定的毒性效应。
Abstract:
Aim:To establish the prokaryotic expression system of MycP1 of Mycobacterium tuberculosis.Methods:The recombinant plasmid of pGEX4T1Rv3883c was constructed and expressed in E.coli BL21(DE3),and then induced by IPTG. The product was identified by SDSPAGE and Western blot and purified with GSTaffinity chromatography. The purified protein was applied to mice macrophages ANA1 cells,and the cytotoxicity was evaluated by detecting LDH activity in culture medium and XTT absorption.Results:The recombinant plasmid pGEX4T1Rv3883c was constructed successfully,and the purified product could decrease XTT absorption(F=34.327,P<0.001),increase the level of LDH activity in culture medium of ANA1 cells (F=336.720,P<0.001).Conclusion:The prokaryotic expression system of MycP1 has been successfully constructed,and the targeted protein has a toxic effect on macrophages.

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备注/Memo

备注/Memo:
*国家传染病重大专项基金资助项目2008ZX10003013;国家自然科学基金资助项目30872257 #通讯作者,男,1954年6月生,博士,教授,研究方向:分子感染免疫,Email:baolang@scu.edu.cn
更新日期/Last Update: 2012-08-14