基于核糖体RNA ITS1-5.8S-ITS2基因序列的十二指肠钩虫分子系统发育分析

Molecular phylogenetic analysis of Ancylostoma duodenales based on ITS1-5.8SrRNA-ITS2 gene sequence

目的通过克隆十二指肠钩虫的ITS1-5.8S-ITS2,初步分析钩口属的系统发育,构建基于ITS1、ITS2的圆线目线虫的系统进化树,为进一步研究其遗传进化关系奠定基础。方法从厦门海沧东孚镇收集标本,分离,形态鉴定为十二指肠钩虫,分别克隆了供试钩虫的ITS1-5.8S-ITS2序列,经NCBI网站的BLAST比对以及基于ITS1和ITS2序列的钩口属的系统发育分析。结果供试钩虫的ITS1、5.8S和ITS2序列,它们的长度分别为366bp、153bp和221bp,登陆http://www.ncbi.nlm.nih.gov进行BLAST,在数据库中没有与之相匹配的序列,将获得的序列作为新的序列上传至GenBank中进行注册,各序列的GenBank登录号分别为:5.8SrRNA基因:EU344796、ITS1-5.8S-ITS2:EU344797。结论从系统进化树中,本实验的供试十二指肠钩虫均与GenBank中已公布的十二指肠钩口线虫自然聚类在一起。

ITS1-5.8S-ITS2 genes of Ancylostoma duodenales was cloned and sequenced in this study. We collected speci- mens from Dongfu Town, Haieang District, Xiamen City after morphological identification for Ancylostoma duodenale. Hook- worm ITS1-5.8S-ITS2 sequence was tested, and the length of ITS1, 5.8S and ITS2 sequences were 366 bp, 153 bp and 221 bp, respectively. The genes of 5.8S rRNA and ITS1-5.8S-ITS2 as new sequences were registered in GenBank, and their num- ber was EU344796 and EU344797, respectively. Based on ITS1 and ITS2 sequence and phylogenetic analysis of Ancylostoma, results have further confirmed that above materials was Ancylostoma duodenales, and the molecular level were accordance well with that of morphological identification.