Construction,identification and expression of recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis

XIE Ting; et al

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Construction,identification and expression of recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis

更新时间：2024-01-17

- Construction,identification and expression of recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis
- Journal of Chongqing Medical University Issue 6, Pages: 657-660(2008)
- 作者机构：
  
  1. 重庆医科大学附属第一医院传染病寄生虫病研究所
  2. 重庆医科大学附属第一医院传染病寄生虫病研究所,重庆,400016
- 作者简介：
- 基金信息：
- DOI：
  CLC： R392
- Published：2008，
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[1]谢婷,李文桂.结核分枝杆菌重组Bb-MPT64疫苗的构建、鉴定及表达[J].重庆医科大学学报,2008(06):657-660+729.

XIE Ting, et al. Construction,identification and expression of recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis. [J]. Journal of Chongqing Medical University (6):657-660(2008)
[1]谢婷,李文桂.结核分枝杆菌重组Bb-MPT64疫苗的构建、鉴定及表达[J].重庆医科大学学报,2008(06):657-660+729. DOI：

XIE Ting, et al. Construction,identification and expression of recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis. [J]. Journal of Chongqing Medical University (6):657-660(2008) DOI：

摘要

目的:构建和鉴定结核分枝杆菌重组双歧杆菌（Bifidobacteria bifidum

Bb）-MPT64疫苗。方法:以结核分枝杆菌H37Rv标准株基因组DNA为模板

通过PCR扩增得到MPT64抗原编码基因序列

定向克隆入大肠埃希菌-双歧杆菌穿梭表达载体pGEX-1λT中

构建重组质粒pGEX-MPT64;用电穿孔法将该质粒导入Bb及BL21（DE3）

构建结核分枝杆菌重组双歧杆菌-MPT64疫苗。用IPTG诱导重组BL21（DE3）菌表达MPT64/GST融合蛋白;SDS-PAGE及Westernblot对表达产物进行鉴定。结果:PCR成功扩增出688bp的MPT64目的基因;双酶切证实MPT64目的基因成功插入pGEX-1λT中;PCR证实rBb-MPT64疫苗构建成功。免疫印迹分析发现重组质粒pGEX-MPT64的表达产物在相对分子量（50ku）处有明显的蛋白表达条带

且能被活动性结核病人血清特异识别。结论:成功构建了结核分枝杆菌rBb-MPT64疫苗

为发展新型结核病疫苗奠定了基础。

Abstract

Objective:To construct and identify recombinant Bb-MPT64 vaccine of mycobacterium tuberculosis. Methods: MPT64 antigen gene were amplified by PCR from mycobacterium tuberculosis H37Rv

then the gene was cloned into Escherichia coli-Bifidobacteria shuttle plasmid pGEX-1λT to construct pGEX-MPT64.The recombinant plasmid was electroporated into Bifidobacteria bifidum(Bb) to construct rBb-MPT64 vaccine.The expression of pGEX-MPT64 was induced with isopropyl-β-D- thiogalactopyranosid(IPTG) and protein MPT64/GST was examined by SDS-PAGE and Western blotting techniques. Results:688bp MPT64 gene was successfully amplified by PCR and cloned into pGEX-1λT by restriction endonuclease analysis.rBb-MPT64 vaccine was successfully constructed by PCR.SDS-PAGE and Western blotting demonstrated that the protein MPT64/GST were expressed in E.coli

BL21. Conclusion:rBb-MPT64 vaccine of mycobacterium tuberculosis is successfully constructed

which may provide basis for the development of new anti-tuberculosis vaccine.

关键词

结核分枝杆菌rBb-MPT64疫苗构建鉴定表达

Keywords

mycobacterium tuberculosisRecombinant Bb-MPT64 vaccineconstructionidentificationexpression

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