小反刍兽疫病毒H基因DNA疫苗的构建及对小鼠的免疫原性

Immunogenicity of DNA Vaccine Expressing H Gene of Peste des Petits Ruminants Virus in Mice

旨在构建表达小反刍兽疫病毒(PPRV)H蛋白的核酸疫苗,并评价其对小鼠的免疫原性。利用RT-PCR扩增了PPRV的H基因并将其克隆到pcDNA3.1(+)载体中,构建真核表达质粒pcDNA3.1-H,然后转染鸡胚成纤维(CEF)细胞,采用间接免疫荧光和Western blot检测H蛋白的瞬时表达情况。将重组质粒通过后腿肌肉注射的方式免疫BALB/c小鼠,采用间接ELISA、淋巴细胞增殖试验和细胞因子检测评价该DNA疫苗的免疫效果,结果表明,成功构建了重组真核表达质粒pcDNA3.1-H,并且能够在CEF细胞中表达。免疫小鼠后,pcDNA3.1-H能够诱导小鼠产生较高的特异性抗体水平,DNA疫苗免疫组小鼠的脾淋巴细胞增殖试验刺激指数(SI)均高于空载体和PBS免疫组,并且能够分泌较高水平的IFN-γ和IL-4。因此,制备的DNA疫苗免疫小鼠后可诱导体液免疫应答和细胞免疫应答,具有较好的应用前景。

The purpose of the study was to construct DNA vaccines expressing Peste des petits ruminants virus(PPRV)H protein,and to evaluate the immunogenicity in mice.The H gene of PPRV was amplified by RT-PCR,and was cloned into the eukaryotic expression vector pcDNA3.1(+)to construct recombinant plasmids pcDNA3.1-H.The H protein transiently expressed in chicken embryo fibroblast cells(CEF)was detected by indirect immunofluorescence and Western blot.Mice were inoculated with recombinant plasmids via intramuscular injection in back legs,the immunogenicity of the vaccine was evaluated by indirect ELISA,lymphocyte proliferation assay and cytokines detection.The results showed that recombinant eukaryotic expression plasmids pcDNA3.1-H was successfully constructed and could express H protein in CEF cell.High specific antibodies were induced by the vaccine.The antigen specific lymphoproliferative test stimulation index of mice immunized with the DNA vaccine was significantly enhanced compared with the control immunized group,and high level of IFN-γand IL-4were secreted by the immunized mice.Hence the DNA vaccine could induce humoral and cellular immune in mice and could be promising for PPR control.