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目的 观察我国登革 2型病毒 43株PrM E基因的DNA免疫原性及非甲基化细菌性CpG对DNA免疫效果的影响。方法 采用RT PCR和分子克隆技术构建PrM E基因片段 ,并将其插入真核表达载体pBK CMV中。在证明其可在哺乳动物细胞中高效表达的基础上 ,进一步用重组质粒DNA免疫小鼠。通过间接免疫荧光法对采集的鼠血清中的病毒特异抗体进行检测。结果 用含PrM E基因的重组质粒DNA免疫小鼠可诱导产生登革 2型病毒特异的抗体 ,且产生的抗体在小鼠体内可持续 3周以上。用含CpG的pUC19质粒和重组质粒的共免疫与单独免疫重组质粒所诱导的抗体水平没有显著差别。结论 我国登革 2型病毒PrM E基因重组质粒DNA具有一定的免疫原性。在本实验条件下 ,含非甲基化细菌性CpG的pUC19质粒DNA ,对PrM E基因的DNA免疫效果没有促进作用。
Objective To observe the DNA immunogenicity of PrM E gene of dengue 2 virus 43 and the effect of unmethylated bacterial CpG on DNA immunization in China. Methods The gene fragment of PrM E was constructed by RT PCR and molecular cloning techniques and inserted into eukaryotic expression vector pBK CMV. The mice were further immunized with recombinant plasmid DNA on the basis of their high expression in mammalian cells. The virus-specific antibodies in the collected mouse serum were detected by indirect immunofluorescence. Results Immunization of mice with the recombinant plasmid DNA containing the PrM E gene resulted in the production of Dengue 2 virus-specific antibodies and the resulting antibodies were sustained in mice for more than three weeks. There was no significant difference in the levels of antibodies induced by CpG-containing pUC19 and recombinant plasmids compared to the immunized recombinant plasmids alone. Conclusion The dengue 2 virus PrM E gene recombinant plasmid DNA has some immunogenicity. Under the experimental conditions, pUC19 plasmid DNA containing unmethylated bacterial CpG did not promote the DNA immunization effect of PrM E gene.