摘要
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。
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.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2006年第6期429-433,共5页
Chinese Veterinary Science
基金
国家自然科学基金资助项目(30371076)
