新城疫病毒F48E9株M NP F和HN基因的真核表达

将新城疫病毒(NDV)F48E9株的M、NP、F和HN基因克隆到真核细胞表达载体 pCAGG上,经酶切、PCR鉴定和序列分析,分别筛选出了含有M、NP、F和HN基因的重组质粒, 命名为pCAGG-M、pCAGG-NP、pCAGG-F和pCAGG-HN。纯化后的重组质粒通过脂质体单独转染HeLa细胞,经间接免疫荧光试验检测到了M、NP、F和HN蛋白的表达。pCAGG-M、 pCAGG-NP、pCAGG-F和pCAGG-HN重组质粒组合共转染HeLa细胞48 h后,可以观察到明显的细胞融合现象。试验结果表明,NDV F48E9株的M、NP、F和HN蛋白均在HeLa细胞内得到了成功表达,同时证明在该表达系统中F蛋白不足以诱导细胞融合。

Eukaryotic expression of M, NP, F and HN genes of NDV strain F48E9

M,NP,F and HN genes amplified from Newcastle disease virus F48E9 strain were cloned into the eukaryotic expression vector pCAGGS, respectively. The positive recombinant plasmids were named pCAGG-M,pCAGG-NP, pCAGG-F and pCAGG-HN after identification by restriction digestion,PCR amplification and sequencing analyses. Then the recombinant plasmids were amplified, purified and transfected into HeLa cells. Expression of M, NP, F and HN genes were detected and confirmed by the indirect immunofluorescence analysis.The syncytium formation was observed in the HeLa cells co-transfected with pCAGG-M, pCAGG-NP, pCAGG-F and pCAGG-HN recombinant plasmids at 48 h post transfection. These results indicated that all four recombinant proteins were successfully expressed in the HeLa cells, but F protein was not efficiently enough to induce cell fusion in the co-expression system.