目的:构建可表达融合蛋白IL-12p70-Ag85A的重组卡介苗,为结核病新疫苗的研制奠定基础.方法:以穿梭表达质粒pMV361为载体,构建重组质粒rpMV361-IL-12p70-Ag85A.将成功构建的重组质粒以电转化方式导入BCG基因组构建重组卡介苗rBCG-IL-12p70-Ag85A.通过抗生素筛选、重组卡介苗基因组PCR扩增及测序,以及western-blotting对重组卡介苗进行鉴定.结果:经鉴定,重组卡介苗成功构建,并可表达融合蛋白IL-12p70-Ag85A.结论:重组卡介苗rBCG-12p70-Ag85A构建成功,并可高效表达目的蛋白.%Objective To construct a recombinant BCG expressing fusion protein of human IL-12p70-Ag85A, to lay the foundation for production of new tuberculosis vaccine. Methods Construct recombinant plasmid of rpMV361-IL-12p70-Ag85A using plasmid pMV361 as the carrier. Then, the recombinant plasmid was imported BCG genome by electric transformation mode to construct recombinant BCG vaccine of rBCG-IL-12p70-Ag85A.The recombinant BCG was certified by antibiotic selection, PCR amplification and sequencing of recombinant BCG genome and westem-blotting. Results The recombinant BCG was certified to be constructed successfully expressing fusion protein of IL-12p70-Ag85A. Conclusion Recombinant rBCG-IL-12p70-Ag85A is constructed successfully, and can highly express the target protein.
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