H9N2禽流感病毒HA全长基因的高效可溶表达及免疫原性研究  

High Soluble Expression and Immunogenicity Assessment of HA Protein of Avian Influenza Virus

作　　者：谢青梅[1] 李少璃[1] 陈丽[1] 周庆丰[1] 马静云[1] 毕英佐[1] 曹永长[1] 

机构地区：[1]华南农业大学动物科学学院,广东广州510642

出　　处：《华南农业大学学报》2006年第3期93-96,共4页Journal of South China Agricultural University

基　　金：广东省科技计划项目(2004A2090102)

摘　　要：采用RT-PCR方法扩增了H9N2亚型禽流感病毒(AIV)广东株的血凝素基因HA.将HA基因克隆到笔者所设计的原核表达载体pMBX上,成功构建重组表达质粒pMBX-HA.将其转化到E.coliBL-21感受态细胞中表达,经SDS-PAGE可检测到相对分子质量约为96 200的融合蛋白,表达产物占菌体总蛋白的29.5%.经蛋白可溶性分析看出,融合蛋白90%以上是以可溶形式表达的.W estern-b lot证实,可溶表达的融合蛋白与H9N2 AIV阳性血清具有良好的免疫反应性.将可溶表达的融合蛋白用体积分数为50%的N i-NTA树脂过柱纯化,用融合蛋白作抗原,通过ELISA方法,能特异性地检测出禽流感阳性血清.The HA gene of H9N2 subtype avian influenza virus （A1T） isolated from Guangdong Province was successfully amplified by RT-PCR. The HA fragment was inserted into plasmid pMBX to obtain recombinant plasmid in which HA gene was fused into. The pMBX-HA was transformed into E. coli BL-21 （DE3） to induce HA gene fusion expression with 1 mmol/L IPTG. The fusion protein band with relative molecular mass of 96 200 was detected on the SDS-PAGE gel, and the soluble expession products accounted for 29.5 % of the total E. coli proteins. The soluble form of the expression products was up to 90% the fusion protein. The expression products specifically reacted with the antisera against H9N2 subtype avian influenza virus. The soluble expression products was purified by 50% （φ） Ni-NTA affinity chromatography and used as an antigen, AIV antibody could be detected by ELISA method.

关 键 词：禽流感病毒 HA基因 高效可溶表达 免疫原性 

分 类 号：S852.65[农业科学—基础兽医学]