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目的构建HSV 1型SM44株糖蛋白D(gD)基因的真核表达载体 ,并用此重组质粒直接免疫小鼠 ,探讨HSV 1gD基因作为基因疫苗的可能性。方法从HSV 1基因组中扩增 gD的全编码基因 ,克隆入载体 pUC19中 ,测序鉴定后转入真核表达载体 pcDNA3.1( +)。所得重组质粒pcDNA gD以电穿孔法转染CHO细胞 ,并以荧光染色法鉴定表达效果。用 pcDNA gD免疫小鼠 ,ELISA法检测基因免疫的应答效果。结果成功地构建了可在真核细胞中有效表达的HSV 1gD真核表达载体 ,用其直接免疫小鼠可引起较高水平的特异性抗体应答。结论 gD的真核表达载体有可能作为HSV 1的基因疫苗 ,为进一步研究基于 gD的表位生物学和表位疫苗奠定了基础。
Objective To construct eukaryotic expression vector of glycoprotein D (gD) gene of SMV strain HSV-1 and to directly immunize mice with this recombinant plasmid to explore the possibility of using HSV 1gD gene as a gene vaccine. Methods The full-length gD gene was amplified from HSV 1 genome and cloned into vector pUC19. The recombinant plasmid was transformed into eukaryotic expression vector pcDNA3.1 (+). The recombinant plasmid pcDNA gD was transfected into CHO cells by electroporation, and the expression was confirmed by fluorescence staining. Mice were immunized with pcDNA gD and the effect of gene immunization was tested by ELISA. Results The HSV 1gD eukaryotic expression vector, which can be efficiently expressed in eukaryotic cells, was successfully constructed. Direct immunization of mice with the recombinant eukaryotic vector resulted in a higher level of specific antibody responses. Conclusion The eukaryotic expression vector of gD may be used as a gene vaccine for HSV 1, which lays the foundation for the further study of gD-based epitope biology and epitope vaccines.