结核杆菌多价核酸疫苗的构建及其免疫原性的初步研究

目的构建包含结核杆菌3种抗原蛋白Ag85A、ESAT-6和CFP-10真核表达质粒的多价DNA疫苗,并对其在小鼠体内的免疫原性进行初步分析。方法PCR扩增获得结核杆菌ag85 a、esat-6和cfp-10基因片段,分别克隆入真核表达质粒VR1020的BamHⅠ位点构建DNA疫苗,以此多价DNA疫苗免疫BALB/c小鼠,检测小鼠体内的特异性抗体应答和细胞免疫反应,并与BCG免疫组、质粒VR1020免疫组和生理盐水对照组相比较。结果多价DNA疫苗免疫小鼠后能产生特异性的细胞和体液免疫应答,DNA疫苗免疫组小鼠脾细胞IFN-γ的表达量显著高于BCG免疫组,而DNA疫苗免疫组小鼠所产生的特异性抗体水平低于BCG免疫组。结论成功构建了结核杆菌多价DNA疫苗,免疫小鼠后检测到特异性细胞和体液免疫应答。

Preliminary Study on the Construction and Immunogenicity of a Multivalent DNA Vaccine Encoding Ag85A,ESAT-6 and CFP-10 Antigens from Mycobacterium tuberculosis

Objective To construct a multivalent DNA vaccine encoding Ag85A,ESAT-6 and CFP-10 antigens from Mycobacterium tuberculosis,and to preliminarily study its immunogenicity.Methods Mycobacterium tuberculosis ag85a、esat-6 and cfp-10 gene fragments were obtained by using Polymerase Chain Reaction(PCR).The gene fragments were cloned into the BamHⅠsite of plasmid VR1020 to construct three recombinant plasmids(multivalent DNA vaccine).BALB/c mice were immunized with the multivalent DNA vaccine、plasmid vector VR1020 or BCG.The hormonal immune response and cell-mediated immune response of the mice were detected and analyzed.Results Specific hormonal immune response and cell-mediated immune response were detected in multivalent DNA vaccine immunized mice and BCG immunized mice.The IFN-γ production of lymphocytes obtained from the multivalent DNA vaccine immunized mice were significantly higher than that from BCG immunized mice,while the specific antibody level of the multivalent DNA vaccine immunized mice was lower than that of BCG immunized mice.Conclusion A multivalent DNA vaccine against Mycobacterium tuberculosis was constructed successfully.BALB/c mice immunized with this kind of DNA vaccine can induce both the hormonal immune response and the cell-mediated immune response.