核酸疫苗双表达载体的构建及表达

Construction and Expression of Co-Expression Vector for DNA Vaccine

将微小病毒内部核糖体进入位点(IRES)基因克隆到质粒pVAXI载体多克隆位点,构建出核酸疫苗双表达载体pVI。将绿色荧光蛋白(EGFP)基因和新霉素磷酸转移酶(neor)基因作为报告基因,连接到pVI载体IRES基因的前后两处多克隆位点,构建出表达载体pEIN。通过脂质体介导的方法将该载体转染COS-7细胞,筛选到同时表达绿色荧光蛋白和新霉素磷酸转移酶的表达株,表明成功地构建了核酸疫苗双表达载体,为构建多价核酸疫苗及带有分子佐剂的核酸疫苗打下了基础。

Internal ribosome entry site （IRES） gene was cloned into plasmid pVAXI to construct a eukaryotic co-expression plasmid pVI for DNA vaccine. Then enhanced green fluorescent protein （EGFP） gene and neomycin phosphotransferase （neo^r） gene fragment were ligated with pVI as reporter genes into its two multiple cloned sites by IRES respectively to construct co-expression plasmid pEIN containing two expression units of EGFP and neor. Recombinant vector was transfected into COS-7 cell by means of lipid mixtures. The cell expressing the two proteins was obtained by screening. The successful construction of the eukaryotic co-expression vector became the foundation for multivalency DNA vaccine and DNA vaccine containing molecular adjuvant.