摘要： 参考GenBank上H5N1亚型禽流感病毒(AIV)的核衣壳蛋白(NP)基因序列设计引物,PCR扩增NP基因,对其进行序列分析并构建分子进化树。将克隆的NP基因插入原核表达载体pET32a,在大肠杆菌中进行诱导表达,对诱导表达的NP重组蛋白进行纯化、Western-Blot鉴定及免疫原性分析。结果表明,克隆的NP基因与GenBank上发表的另外2个河南AIV毒株亲缘关系较近,诱导表达的NP重组蛋白主要以可溶性形式存在,分子量约为70 kDa,并且能够与H5亚型AIV阳性血清发生特异性反应,具有良好的抗原性。
关键词:
禽流感病毒,
H5亚型,
核衣壳蛋白(NP),
原核表达
Abstract: The Nucleocapsid Protein ( NP) gene of Avian influenza virus( AIV) subtype H5 was amplified by PCR according to the published sequence, and its molecular evolution was analyzed.The coding sequence of NP was cloned into the prokaryotic vector of pET32a to express and obtain the recombinant protein inE． coli.The expressed NP was purified and identified with Western-Blot strategy.and its immunogenicity was further characterized with enzyme linked immunosorbent assay.The result suggested that the cloned NP was evolutionary more similar to others of Henan.and the fusion protein was successfully expressed and mainly existed as soluble bodies,Also, the protein could be specifically recognized by the serum against H5 subtype of AIV．
Key words:
Avian influenza virus,
Subtype H5,
Nucleocapsid protein,
Prokaryotic expression
中图分类号:
邬成业, 王爱萍, 郝慧芳, 史平玲, 游雷鸣, 李培培, 张改平. H5N1亚型禽流感病毒核衣壳蛋白(NP)基因的进化及原核表达[J]. 华北农学报, 2010, 25(3): 19-22. doi: 10.7668/hbnxb.2010.03.005.
WU Cheng-ye,, WANG Ai-ping, HAO Hui-fang, SHI Ping-ling, YOU Lei-ming, LI Pei-pei, ZHANG Gai-ping. Phylogenetic Analysis and Prokaryotic Expression of NP Gene from Subtype H5N1 of Avian Influenza Virus in Escherichia coli[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2010, 25(3): 19-22. doi: 10.7668/hbnxb.2010.03.005.