河豚鱼Cyt b基因部分DNA序列分析与应用

Partial DNA Sequence Analysis of Cyt b Gene in Puffer Fish

建立河豚鱼物种DNA鉴别技术,根据GenBank公布的河豚鱼细胞色素b基因序列,应用软件Primer Premier5.00版设计上游引物HT1-F与下游HT1-R,对3属9种福建省搜集的常见的河豚鱼与2种未知物种名称且外观无法进行形态学判断的河豚鱼样品的细胞色素b基因序列中的部分片段进行PCR扩增,PCR产物通过琼脂糖凝胶电泳确证、纯化后,进行DNA碱基序列测定,序列长度均为423bp。应用DNA MANN软件进行样品间DNA序列同源性比对分析,建立样品间系统关系树状图,供试11个样品被划分为3个类群,第Ⅰ组与第Ⅱ组之间的同源率为89%,这2组与第Ⅲ组之间的同源率为85%。根据序列同源性分析结果,可推测2个未知种名的样品为腹刺鲀属或东方鲀属。

According to the sequence of Cytochrome b in puffer fish published in GenBank,a puffer fish-specific primer pair of HT1-F and HT1-R was designed using Primer Premier 5.00 version for establishing a DNA based method to identify puffer fish species.Partial fragments of Cytochrome b gene in puffer fish samples including 9 major species from 3 genera and 2 unknown puffer fish samples harvested from Fujian province were amplified by PCR.The PCR products confirmed by agarose electrophoresis were used for sequencing.The length of the obtained DNA fragments was 423 bp in all samples.The homology of DNA sequences among these samples was analyzed using DNA MAN software,and a phylogenetic tree among these samples was also established.Eleven samples were divided into 3 groups,89% of homology rate was observed between groups I and II;85% of homology rate was observed between two groups and group III.In addition,2 unknown puffer fish samples were probably from Gastrophysus or Takifugu genus.Collectively,these results provide a reference for the application of DNA sequencing technology to identify puffer fish components and its species present in processed food stuff.