汉坦病毒包膜糖蛋白G2的基因克隆与序列分析

目的 建立汉坦病毒(HV)包膜糖蛋白G2的克隆载体;进行系统发生树分析,研究G2基因的变异情况.方法 应用逆转录聚合酶链反应(RT-PCR)扩增山东省HV G2基因片段,克隆于PMD-18T载体,经氨卞西林筛选,酶切鉴定后,进行序列测定,应用DNASTAR软件将其与世界范围内的病毒株基因序列进行分析.结果 扩增得到山东省高密、淄川、莒南、荣成四地G2基因.序列同源性分析表明,四地G2基因都属于SEO型HV,与Z37株核苷酸同源性最高,与其他SEO型各株的同源性为82.3%～99.8%;绘出了G2基因及氨基酸的系统发生树.结论 成功地建立了山东省四地HV G2基因克隆载体;四地HV G2基因同源性高,为山东省SEO型HV的遗传与变异、分子流行病学研究,以及制备有效的亚单位疫苗提供了依据.

Cloning and sequence analysis of envelope glycoprotein G2 gene of hantavirus in Shandong province

OBJECTIVE: To construct the cloning vector of glycoprotein G2 gene of hantavirus (HV), to analyze the sequence of G2 gene by the phylogenetic tree, and to study the differences among glycoprotein G2 genes from the world around. METHODS: Envelope glycoprotein G2 gene was amplified from four specimens of Shandong province by RT-PCR, and the product recombined into the PMD-18T vector. The clones that carry the G2 gene were identified. After sequencing, the gene sequence was handled with the software DNASTAR, compared with 24 strains worldwide and the phylogenetic tree was drawn. RESULTS: HV G2 gene was amplified by RT-PCR from 4 specimens, named GM04-38.G2, ZB8.G2, JUN5-14.G2, RCH5.G2, respectively. The map of the phylogenetic tree showed that all the 4 strains belonged to SEO-type hantavirus. The analysis of the sequence showed that all the four HV strains had the highest rates of homology with Z37 strain. The sequence homology of SEO-type HV strains was from 82.3% to 99.8%. CONCLUSION: The four cloning vectors containing the glycoprotein G2 genes were successfully constructed. Envelope glycoprotein G2 gene of four specimens from Shandong province had high homology rates.