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目的:构建草原兔尾鼠卵透明带3 DNA疫苗pVAX1-sig-LTB-lZP3-C3d3进行小鼠的黏膜免疫,增强该疫苗的免疫不育效果。方法:将两种佐剂分子大肠杆菌不耐热肠毒素B亚基和C3d基因通过酶切鉴定,构建重组质粒pVAX1-sig-LTB-lZP3-C3d3,采用RT-PCR和W estern b lot技术,检测其在mRNA和蛋白水平的表达,并通过肌肉注射、滴鼻和灌服3种途径免疫雌性C57BL/6小鼠,通过ELISA检测抗体水平及分型。结果:酶切鉴定,RT-PCR和W estern b lot结果表明重组质粒构建正确并可在mRNA和蛋白水平的表达,ELISA结果表明以重组质粒pVAX1-sig-LTB-lZP3-C3d3免疫诱导的特异性IgG、IgA的水平明显高于对照组(P<0.01)。抗生育实验表明该疫苗免疫的小鼠平均生仔数与对照组相比差异极显著(P<0.01)。结论:构建的草原兔尾鼠卵透明带3DNA疫苗重组质粒pVAX1-sig-LTB-lZP3-C3d3可高效地激发小鼠特异性免疫应答,提高抗生育的效果。
OBJECTIVE: To construct mucosal immunity of mice and to enhance the immunostimulatory effect of the vaccination vaccine against pVAX1-sig-LTB-1ZP3-C3d3. Methods: The recombinant plasmids pVAX1-sig-LTB-1ZP3-C3d3 were constructed by restriction endonuclease digestion of the two adjuvant E. coli heat-labile enterotoxin B subunits and C3d gene. RT-PCR and Western blotting The mRNA and protein levels were detected. Female C57BL / 6 mice were immunized by intramuscular injection, intranasal instillation and oral administration. Antibody levels and typing were detected by ELISA. Results: The results of restriction enzyme digestion, RT-PCR and Western blot showed that the recombinant plasmid was correctly constructed and expressed at the mRNA and protein levels. The ELISA results showed that the specific immunization induced by the recombinant plasmid pVAX1-sig-LTB-1ZP3-C3d3 IgG, IgA levels were significantly higher than the control group (P <0.01). Anti-fertility experiments showed that the vaccine-immunized mice average litter size compared with the control group, the difference was significant (P <0.01). CONCLUSION: The recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 of the zona pellucida with 3DNA vaccine of the zoo of the Prairie Dog can efficiently stimulate the mouse-specific immune response and improve the anti-fertility effect.