论文部分内容阅读
目的 :构建登革 3型病毒NS1基因的真核重组表达质粒 ,观察其免疫原性 ,为登革新型疫苗的研究提供依据。方法 :从感染鼠脑中提取登革 3型病毒RNA ,经RT_PCR获得NS1基因片段 ,然后将其克隆到真核表达载体中。用电穿孔法将重组质粒DNA转入COS7细胞 ,通过免疫荧光法检测外源基因在真核细胞中的表达。将重组质粒DNA免疫BALB c小鼠 ,观察其免疫原性。结果 :通过酶切鉴定和序列测定证实了NS1基因片段已经插入真核表达载体。用免疫荧光法证明转染了重组质粒的COS7细胞的胞浆中有特异荧光。重组质粒DNA免疫小鼠后可观察到诱发产生的细胞免疫应答。结论 :含有登革 3型病毒NS1基因的真核重组表达质粒可以在COS7细胞中进行表达 ,而且能够诱导小鼠产生细胞免疫应答
Objective: To construct eukaryotic recombination plasmid of dengue 3 virus NS1 gene and observe its immunogenicity, so as to provide basis for the study of new dengue vaccine. Methods: The dengue virus type 3 RNA was extracted from infected mouse brain and the NS1 gene fragment was obtained by RT_PCR. The NS1 gene fragment was cloned into eukaryotic expression vector. The recombinant plasmid DNA was transfected into COS7 cells by electroporation, and the expression of foreign gene in eukaryotic cells was detected by immunofluorescence. The recombinant plasmid DNA was immunized BALB c mice to observe its immunogenicity. Results: The NS1 gene fragment was inserted into the eukaryotic expression vector by restriction enzyme digestion and sequencing. The results of immunofluorescence showed that there was specific fluorescence in the cytoplasm of COS7 cells transfected with recombinant plasmids. The induced cellular immune response was observed after immunization of mice with recombinant plasmid DNA. Conclusion: The eukaryotic recombinant plasmid containing NS1 gene of dengue virus type 3 can be expressed in COS7 cells, and can induce mice to produce cellular immune response