湖南省猬迭宫绦虫ITS及5.8S rDNA的克隆及序列分析

Cloning and sequence analysis of ITS and 5.8S rDNA of Spirometra erinaceieuropaei isolates from Hunan province

利用聚合酶链反应（PCR）扩增猬迭宫绦虫rDNA的ITS-1、5.8S及ITS-2片段,将PCR扩增出的片段纯化后克隆至pGEM-T Easy载体,重组质粒通过菌落PCR鉴定后,对阳性菌落进行序列测定并进行序列分析。结果显示,所获得的ITS及5.8S rDNA序列总长存在一定差异,为1 369~1 393 bp,包含部分的18、28S及全部的ITS-1（662~687 bp）5、.8S（138 bp）及ITS-2（457~475 bp）序列。由于猬迭宫绦虫ITS序列种内相对保守,种间差异较大,故可作为种间遗传变异研究的标记。

To examine sequence variation in the internal transcribed spacer（ITS） and 5.8S rDNA of Spirometra erinaceieuropaei isolates from Hunan province,and to reconstruct their phylogenetic relationship using ITS sequences,the ITS sequences were amplified from each S.erinaceieuropaei sample and the amplicons were cloned into pGEM-T Easy vector,respectively.The inserts were successfully sequenced,and the length of ITS in 17 S.erinaceieuropaei isolates were different（1 369-1 393 bp）.Sequence analysis revealed that the ITS-1,5.8S and ITS-2 rDNA of these samples were 662-687 bp,153 bp and 457-475 bp in length,respectively.There is no significant variation in ITS sequences within S.erinaceieuropaei,while inter-species difference is obvious.The results indicated that the ITS sequences provide useful genetic markers for molecular identification of S.erinaceieuropaei.