摘要
目的:观察密码子优化能否增强乙型肝炎病毒表面抗原中蛋白核酸疫苗的免疫原性。方法:根据乙型肝炎病毒(adr亚型)表面抗原中蛋白(MHBs)的氨基酸序列,在不改变其氨基酸序列的基础上,设计并人工合成了密码子优化的基因,并将该基因克隆到核酸疫苗载体pSW3891中,构建了密码子优化的表达MHBs核酸疫苗(命名为:pSW3891/MHBs/adr/opt,简称opt)。用上述核酸疫苗与表达野生型MHBs核酸疫苗(命名为:pSW3891/MHBs/adr,简称adr)及空载体质粒pSW3891一起瞬时转染293T细胞,应用蛋白质印迹法检测转染细胞中MHBs的表达。采用肌肉注射法,以opt、adr及pSW3891,分别对BALB/c小鼠进行免疫。用ELISA方法检测免疫后小鼠血清中HBs抗体的滴度,ELISPOT法检测免疫小鼠表面抗原特异性分泌IFN-γ的脾细胞。结果:opt和adr均可在体外293T细胞中高效表达,且opt的蛋白表达量要高于野生型。opt免疫组小鼠特异性抗体滴度和免疫小鼠表面抗原特异性分泌IFN-γ脾细胞数量都要显著的高于adr免疫组小鼠。结论:密码子优化可以增强乙型肝炎病毒DNA疫苗在体外的蛋白表达量及免疫小鼠的体液免疫和细胞免疫原性。
Objective：To investegate whether codon optimization of middle hepatitis B surface antigen could enhance the immunogenicity of HBV DNA vaccine. Methods：According to the sequence of amino acids of MHBs （adr subtype）, the codon optimized genes were designed and synthesized without changing the sequence of amino acids and then cloned into vector pSW3891, which was named pSW3891/MHBs/adr/opt. 293T cells were transiently transfected with opt and wild-type MHBs DNA vaccine （pSW3891/ MHBs/adr） and empty vector pSW3891. The protein expression level was assessed by western blot. BALB/c mice were intramuscularly injected with opt, adr or pSW3891. Anti-HBs in sera was tested by ELISA. IFN-γ,secretion splenocytes of mice immunized with opt, adr and pSW3891 were tested by ELISPOT. Results：The level of protein expression in both supernatant and ]ysate of 293T cells transfected with opt is higher than that in 293T cells transfected with adr. The BALB/c mice immunized with opt showed stronger anti-HBs response and more IFN-γsecretion splenocytes than those immunized with adr. Conclusion： Codon optimization of MHBs DNA vaccine could improve protein expression and induce significant humoral and cellular immune responses in BALB/c mice.
出处
《南京医科大学学报（自然科学版）》
CAS
CSCD
北大核心
2009年第3期286-290,共5页
Journal of Nanjing Medical University(Natural Sciences)
基金
国家自然科学基金资助(30371276)
