【摘要】 将鹅源新城疫病毒的NP、P和L基因通过RT PCR方法从尿囊液中扩增后分别克隆进pGEM Teasy载体 ,再分别亚克隆到真核表达载体pCI neo上 ,通过酶切、PCR和测序验证克隆正确。利用P基因开放性阅读框 (ORF)上靠近终止密码上游的AgeI位点 ,将报告基因绿色荧光蛋白 (GFP)基因克隆进P基因真核表达重组质粒 ,分别转染COS 1细胞和CEF细胞 ,在倒置荧光显微镜下可见到绿色荧光 ,表明GFP基因已得到表达 ,由此证明P基因也已得到表达。鹅源新城疫病毒NP、P和L基因的克隆成功 ,为即将进行的鹅源新城疫病毒的反向遗传操作以及功能基因组研究打下基础更多还原

【Abstract】 NP, P and L gene of Newcastle disease virus of goose origin were amplified and cloned into pGEM-T easy vector and then subcloned into pCI-neo expression vector respectively, the positive clones were identified by enzyme cutting, PCR and sequencing. GFP reporter gene was inserted into the downstream of recombinant expression plasmid of P gene, which of stop codon was deleted. The experiment of transfection of P and GFP recombinant plasmid on COS-1 cells and CEF showed that GFP gene expressed, and this demonstrated that P gene was also expressed. This research may be helpful for further study of reverse genetics and functional genome of NDV of goose origin.更多还原