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目的获得HIV-1 CRF07_BC株gp41重组抗原表达效率最佳的体系。方法采用基因工程技术将gp41重组蛋白基因分别构建到pThioHis A,pThioHis A-1(不含标签),PBV220,PGEX-6p-1原核表达载体。转化大肠杆菌BL21和Rosetta两种宿主菌,通过IPTG诱导表达并经WB鉴定。结果 4种原核表达质粒构建完成,蛋白成功表达。PGEX-6p-1表达量明显高于pThioHis A载体;而无标签载体组中,PBV220表达量优于pThioHis A-1。BL21和Rosetta宿主菌对表达体系无影响。结论 HIV-1 CRF07_BC株gp41重组抗原在大肠杆菌中以包涵体形式存在,构建在PGEX-6p-1和PBV220载体上gp41重组蛋白表达效率较高,为gp41疫苗的研究和血清学检测提供充足的抗原。
Objective To obtain the best expression system of gp41 recombinant antigen of HIV-1 CRF07_BC strain. Methods Recombinant gp41 protein was constructed into prokaryotic expression vector pThioHis A, pThioHis A-1 (without tag), PBV220 and PGEX-6p-1 respectively by genetic engineering. Two host strains, E. coli BL21 and Rosetta, were transformed and induced by IPTG and identified by WB. Results Four prokaryotic expression plasmids were constructed and the protein was successfully expressed. The expression level of PGEX-6p-1 was significantly higher than that of pThioHis A vector, but PBV220 was higher than pThioHis A-1 in the vector without tag. The host strains BL21 and Rosetta had no effect on the expression system. Conclusion The gp41 recombinant antigen of HIV-1 CRF07_BC strain exists as inclusion bodies in E. coli. The expression of recombinant gp41 protein in PGEX-6p-1 and PBV220 vectors is high, and it provides abundant gp41 vaccine and serological tests antigen.