湖南省华支睾吸虫线粒体cox1和nad1基因的克隆及进化分析

为了研究湖南省华支睾吸虫分离株线粒体细胞色素c氧化酶第Ⅰ亚基(cox1)基因部分序列(pcox1)和烟酰胺腺嘌呤二核苷酸脱氢酶亚单位Ⅰ基因(nad1)部分序列(pnad1)的遗传变异情况,并用pcox1和pnad1序列构建华支睾吸虫与其他吸虫的种群遗传进化树。应用聚合酶链反应扩增华支睾吸虫虫株的pcox1和pnad1,应用ClustalX 1.81程序对序列进行比对,然后用Phylip3.67程序中的MP法和MEGA4.0程序NJ法绘制种系发育树,并用Puzzle5.2程序构建最大似然树。结果显示,所获得的pcox1和pnad1序列长度分别为1 100bp和790bp,种系发育进化表分析表明,25个华支睾吸虫分离株位于同一分枝。证实华支睾吸虫pcox1和pnad1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,从而为华支睾吸虫的分类鉴定以及进一步的分子流行病学调查和群体遗传研究奠定基础。

Cloning and phylogenetic analysis of mitochondrial cox1 and nad1 genes of Clonorchis sinensis isolates from Hunan Province

The objectives of the present study were to examine the sequence variation in the partial mitochondrial cytochrome c oxidase subunit 1(pcox1) and NADH dehydrogenase subunit 1(pnad1) genes among Clonorchis sinensis isolates from Hunan Province,and to study the phylogenetic relationship among trematodes of the other family using pcox1 and pnad1 sequences.The pcox1 and pnad1 were amplified from each C.sinensis sample by PCR,and pcox1 and pnad1 sequences were aligned using the ClustalX 1.81.MP and NJ trees were constructed using the software Phylip version 3.67 and MEGA version 4.0 respectively,and ML tree was also constructed using Puzzle version 5.2.The lengths of the pcox1 and pnad1 sequences were 1 100 bp and 790 bp,respectively.The phylogenetic tree showed that 25 Hunan isolates were clustered in the same clade.There was no significant variation in pcox1 and pnad1 sequences within C.sinensis,while inter-species difference was obvious.It was concluded that pcox1 and pnad1 sequences could be used as genetic markers for population genetic studies of C.sinensis.The results of this study lay foundations for further studies on classifications identification molecular epidemiology and population genetic of C.sinensis.