H5N1 亚型禽流感病毒NS1的克隆和原核表达

利用RT-PCR技术扩增了一株H5N1亚型禽流感病毒(AIV)的NS1基因,克隆到pGEM T-easy载体上,经序列测定和分析正确后,将该基因插入到pET-32a载体中构建原核表达载体pET-NS1,阳性重组质粒转化BL21感受态细胞,用1mmol/L的IPTG诱导和SDS-PAGE电泳后获得了分子质量约为51ku的NS1融合蛋白。通过Western-blotting发现NS1蛋白可与H5N1亚型AIV单因子血清反应。为建立H5N1亚型AIV野毒株和疫苗株的特异性鉴别方法奠定了基础。

Cloning and Procaryotic Expression of NS1 Protein of Avian Influenza Virus H5N1 Subtype

The NS1 gene was amplified by RT-PCR from H5N1 subtype of influenza A virus(AIV).After cloning and sequencing,the gene was integrated into pET-32a vector. Recombinant plasmid pET-NS1 was transformed into E.coli BL21(DE3)(pLysS) competent cells and induced with 1mmol/L IPTG.The fusion protein of 51Ku as detected by SDS-PAGE was produced and tested by Western-blotting which could react with the NS1 serum of the H5N1 influenza A virus. The study provides a sound base for constructing new method of differentiating field AIV strain and vaccine strain.