【摘要】 目的构建单纯疱疹病毒Ⅰ型(herpes simplex virus I,HSV-I)SM44株截短糖蛋白B基因序列,为研制联合基因疫苗奠定基础,用于角膜炎的防治。方法利用聚合酶链反应(polymerase chain response,PCR)技术从感染HSV-I SM44株的vero细胞中扩增出HSV-gBt编码基因,经双酶切鉴定后将目的基因定向插入真核表达质粒pcDNA3载体中,构建出重组真核表达质粒pcDNA3-gBt,并对其进行酶切分析和测序鉴定。结果对pcDNA3-gBt双酶切后,电泳可见目的基因(1.5kb)和线性质粒pcDNA3(5.4kb)两条带;测序结果表明,克隆基因插入方向正确,与基因库中登录的HSV-1F株gB基因序列比较,同源性达99.5%。结论经证实成功地构建了重组真核表达质粒pcDNA3-gBt,为进一步研究其免疫学效应及构建病毒糖蛋白联合疫苗并最终用于单疱病毒角膜炎的预防和治疗奠定理论基础更多还原

【Abstract】 Objective To construct an eukaryotic expression plasmid of herpes simplex virus I(HSV-1)truncated glycoprotein B and to make the foundation for the construction of multi-valent DNA vaccine.Methods The encoding sequence of the truncated glycoprotein B was amplified from HSV-I SM44 DNA genome by means of polymerase chain response,incised by two different endonucleases,and then was directionally integrated into eukaryotic expression vector pcDNA3,and the recombinant vector pcDNA3-gBt was constructed,which was confirmed by the restriction endonuclease analysis and DNA sequencing.Results Electrophoresis of the products of double-enzyme digestion and analysis of the recombinant vector pcDNA3-gBt showed two bands,one was HSV-1 gBt gene(1.5 kb)and the other was linear pcDNA3(5.4 kb);DNA sequencing showed that the insertion orientation was right and the rate of homology was 99.5% compared with HSV-F of GeneBank.Conclusions The recombinant vector was constructed successfully,which would provide a theoretical basis to evaluate the possibility of the construction of multi-valent DNA vaccine for HSV-Ⅰ更多还原