摘要
目的 克隆表达结核分枝杆菌Rv1884基因 ,序列测定正确后进行融合表达、纯化。方法 采用聚合酶链反应(PCR)从结核分枝杆菌H37Rv基因组中扩增出Rv1884编码基因 ,用限制性内切酶消化后插入到pGEM Teasy中 ,序列测定正确后 ,再亚克隆到融合表达载体 pPro EXHT中 ,转化大肠杆菌DH5α ,目的基因经IPTG诱导 ,由T7启动子调控表达了氨基端带 6个连续组氨酸残基的Rv1884蛋白 ,在变性条件下对目的蛋白进行纯化。结果 获得了结核分枝杆菌H37Rv株Rv1884蛋白基因 ,得到融合 6个组氨酸残基的Rv1884蛋白纯度大于 85 %。结论 构建了结核分枝杆菌Rv1884基因的重组表达载体 ,并获得了高纯度的融合表达蛋白 ,为以后的深入研究奠定了基础。
To clone the RV1884 gene and to purify the fusion protein of Mycobacterium tuberculosis, the gene encoding RV1884 protein was amplified from genome from M.tuberculosis H37Rv strain by PCR, and was inserted into cloning vector pGEM Teasy. After sequencing, it was then subcloned into expression vector PPro EX HT and transfected to host strain E.coli DH5α. The strain controlled by T7 promotor expressed the fused RV1884 protein with a hexa histidine tail at its N terminal end after induction with IPTG and the target protein was purified under denaturing condition. It was found that the recombinant expression vector for gene encoding M.tuberculosis RV1884 was successfully constructed and the fusion protein with high purity was obtained. These results would provide the basis for the further studies of the RV1884 protein.
出处
《中国人兽共患病杂志》
CSCD
北大核心
2005年第1期18-21,共4页
Chinese Journal of Zoonoses
