结核分枝杆菌Rv1512基因的特异性和克隆表达

摘要

Objective To identification of specific antigens RvlS12 of Mycobacterium tuberculosis. To construct expression vector carrying Mycobacterium tuberculosis (M. tb) Rvl5l2 gene and express it in E. coli. Methods Design RvlS12 gene-specific primers, and identify its specificity. The positive clones of expression recombinant plasmids pET30a( + ) ;Rvl512 were constructed and then transformed into BL21. The fusion protein was purified by Ni * -NTA column and detected by SDS-PAGE. Results Mycobacterium tuberculosis Rvl512 gene specificity was confirmed. Sequence analysis confirmed that the identification of prokaryotic expression plasmid was correct. The SDS-PACE results showed that the molecular weight of Rvl512 fusion protein was about 40 kD. Conclusion Mycobacterium tuberculosis RvlS12 gene has better specificity. The expression recombinant plasmids pET30a( + ) :Rvl512 was successfully constructed, recombinant RvlS12 protein is produced, which laid the foundation for application in the diagnosis of Mycobacterium tuberculosis .%目的 鉴定结核分枝杆菌Rv1512基因的特异性,克隆表达该基因并获得其重组蛋白.方法 设计Rv1512基因的特异性引物,并鉴定其特异性.构建pET30a(+):Rv1512重组质粒,阳性克隆转化入大肠杆菌BL21.经Ni+ -NTA层析柱纯化融合蛋白,通过SDS-PAGE鉴定该蛋白及其纯度.结果 结核分枝杆菌Rv1512基因的特异性被证实；经测序分析证实Rv1512原核表达质粒构建正确,SDS-PAGE结果显示在40 kD处呈现单一蛋白条带.结论 结核分枝杆菌Rv1512基因具有较好的特异性；成功构建表达载体pET30a(+ ):Rv1512并获得重组蛋白,为辅助诊断结核分枝杆菌的感染奠定基础.