新城疫病毒F和HN基因真核表达载体的构建

Construction of eukaryotic expression vector harbouring F and HN gene of Newcastle disease virus

根据GenBank中新城疫病毒(NDV)F、HN基因序列和真核细胞表达载体pEGPF-N1的克隆位点,分别设计一对引物,通过PCR方法获得了F、HN基因,经回收纯化,将产物连接到克隆载体pMD18-T上。酶切鉴定和序列分析后,再将目的基因亚克隆到真核细胞表达载体pEGPF-N1上,经鉴定,成功构建含有NDVF和HN基因的真核细胞表达载体pEGPF-N1-F和pEGPF-N1-HN,为下一步研究NDVF和HN蛋白在细胞凋亡中的生物学功能奠定基础。

According to the encoding sequence of the NDV F and HN protein and the multiple Cloning sites characteristic of the eukaryotic expressional vector pEGPF-N1 in GenBank, appropriate primers specific to NDV F and HN gene were designed. The target DNA fragments were obtained by PCR amplification and then were cloned into the pMD18-T vector respectively. After identification by restriction digestion and sequencing analysis, the specific DNA fragments were cloned into the eukaryotic expression vector pEGPF-NI, respectively. The positive recombinant plasmids were named pEGPF-NI-F and pEGPF-NI-HN after identification by restriction enzyme digestion, PCR amplification. Results showed that recombinant expression plasmids pEGPF-NI-F and pEGPF-N1-HN were successfully constructed and could be the foundation of analyzing biological role of F and HN protein of NDV in cell apoptosis.