猪流行性腹泻病毒Nsp7基因真核表达载体的构建及表达

Construction of Eukaryotic Expression Vector of Porcine Epidemic Diarrhea Virus Nsp7 Gene and Expression of Fusion Protein

PEDV Nsp7蛋白是猪流行性腹泻病毒的一种非结构蛋白,它在病毒复制过程中表达,并与感染细胞的内质网和细胞膜相关,是病毒基因组翻译的两个大型复制酶的关键辅助因子。为了对PEDV Nsp7蛋白进行更加深入的研究,试验从病变猪小肠中提取总RNA,并通过RT-PCR扩增获取PEDVNsp7基因,最终将其连接到pCMV-Myc载体上。试验结果显示,成功构建了pCMV-Myc-Nsp7的重组质粒,并通过Western Blot技术和免疫荧光技术在真核细胞中验证了其蛋白表达情况,为后续研究PEDV Nsp7的生物学功能奠定了良好的基础。

PEDV Nsp7 protein was a non-structural protein of porcine epidemic diarrhea virus.It was expressed during viral replication and associated with the endoplasmic reticulum and cell membrane of infected cells.It was a key cofactor of two large replicases in viral genome translation.In order to further study PEDV Nsp7 protein,total RNA was extracted from the small intestine of diseased pigs,and PEDV Nsp7 gene was obtained by RT-PCR amplification,and finally connected to pCMV-Myc vector.The experimental results showed that the recombinant plasmid pCMV-Myc-Nsp7 was successfully constructed,and its protein expression was verified in eukaryotic cells by Western Blot and immunofluorescence techniques,which laid a good foundation for subsequent research on the biological function of PEDV Nsp7.