甲型流感病毒H1N1亚型HA基因的克隆及序列分析

Cloning and bioinformatics analysis of influenza A virus subtype H1N1 hemagglutinin gene

目的克隆甲型流感病毒H1N1亚型血凝素HA基因,并对其进行序列分析。方法利用TRIzol Rea-gent法提取甲型流感病毒H1N1亚型的总RNA,RT-PCR后以其cDNA为模板PCR扩增HA基因,经酶切、连接后,构建pET 28a(+)-HA重组质粒。利用生物信息学工具对HA的核酸序列和蛋白功能结构进行分析。结果成功构建了pET 28a(+)-HA重组质粒,经序列比对,与GenBank上公布的HA基因序列同源性达到99%。结论对HA基因进行克隆及生物信息学的分析,为深入研究HA在病毒侵入细胞及抗原变异中的作用机制及后续HA抑制剂的体外筛选奠定基础。

Objective To clone influenza A virus subtype H1 N1 hemagglutinin （HA） gene, and do sequence analy- sis. Methods The total RNA was isolated by TRIzol reagent from influenza A virus subtype H1N1, the cDNA was gained by RT-PCR and then used as a template for PCR, amplified HA gene. Recombinant plasmid pET 28a（ ＋ ）-HA was con- structed. To analyze the nucleic acid sequence and properties of HA by bioinformatics analysis software. Results The re- combinant plasmid pET 28a（ ＋ ）-HA was constructed, sequence alignment result showed that the sequence shares 99% of sequence homology with the HA reported in GenBank. Conclusion The cloning and bioinformatics analysis of HA lay a foundation for the deeply study of its role in virus replication cycle and the in vitro screening of HA inhibitors.