4株不同亚型流感病毒NP基因的原核表达

为了进一步研究A型流感病毒核蛋白功能及在诊断中的应用,采用RT-PCR技术分别扩增H1N1、H3N2、H5N1、H9N2等4株不同亚型流感病毒的NP基因,分别将其克隆到原核表达载体pET30(a)上,在大肠埃希菌中进行诱导表达.SDS-PAGE和Western blot检测结果表明,H1N1亚型流感病毒NP蛋白得到表达,并且能分别与H1N1和H5N1亚型流感病毒的鼠高免血清发生特异性反应,具有良好的反应活性;而H3N2、H5N1、H9N2亚型流感病毒的重组NP蛋白未获得表述.本研究结果表明,不同毒株的NP蛋白在大肠埃希菌中的表达具有一定差异性,为把NP蛋白作为诊断抗原的开发、ELISA诊断方法的建立以及蛋白功能的研究奠定了基础.

Prokaryotic Expression of NP Genes of Four Influenza A virus

In order to research the function and application in diagnosis of nucleoprotein of influenza A virus,the NP genes of four influenza A viruses(H1N1,H3N2,H5N1,H9N2)were amplified by RT-PCR,then the NP genes were cloned into the prokaryotic expression vector pET30(a) and expressed in E.coli induced by IPTG respectively.The results of SDS-PAGE indicated that the recombinant NP of H1N1 subtype of swine influenza virus was produced while that of H3N2,H5N1,H9N2 were not.The immune reactions were confirmed by Western blotting after being reacted with the antisera of H1N1 and H5N1 subtype influenza viruses,respectively.On the one hand the results had great impact on the exploitation of diagnosis reagent depending on NP,establishment of ELISA and research on the function of NP,on the other hand it indicated that there were something different between the NP genes of different strains which expressed in E.coli.