【摘要】 目的:克隆、表达结核分枝杆菌PPE家族蛋白PPE68的编码基因Rv3873,并初步探索PPE68诱导Balb/c小鼠脾淋巴细胞的增殖活性,为进一步研究PPE68的免疫功能提供实验依据。方法:以结核分枝杆菌H37Rv基因组为模板,PCR扩增Rv3873基因编码序列,定向克隆入融合蛋白原核表达载体pET32a(+),获得重组表达质粒,转化大肠杆菌后用IPTG进行诱导表达,纯化表达产物,通过XTT比色法检测重组蛋白PPE68诱导脾淋巴细胞的增殖活性。结果:重组质粒pETRv3873构建成功,57kDa的His-PPE68融合蛋白在大肠杆菌中稳定表达,并具有刺激Balb/c小鼠脾淋巴细胞增殖的功能。结论:PPE68蛋白具有较强的免疫原性,其免疫学功能具有进一步研究的价值。更多还原

【Abstract】 Objective: To express and purify recombinant PPE68, and study the proliferation activity of splenic lymphocyte induced by recombinant PPE68. Methods: Rv3873 was amplified from Mycobacterium tuberculosis H37Rv genomic DNA by PCR. PCR product was cloned into prokaryotic expression vector pET32a(+). The recombinant plasmid with PPE68 gene was transformed into E.coli BL21(DE3) to express the fusion protein His-PPE68 induced by IPTG .Stimulating lymphocyte of spleen with purified recombinant PPE68 and detecting lymphocyte proliferation activity by XTT colorimetry method. Results: Recombinant expression vector pETRv3873 was successfully constructed and the recombinant plasmid could express the 57 kDa fusion protein His-PPE68 stably. Conclusion: It has the potential of inducing splenic lymphocyte proliferation, providing the basis for the further study of immunogenicity and immunoprotection of PPE68 from TB infection.更多还原