食品科学 ›› 2018, Vol. 39 ›› Issue (2): 163-169.doi: 10.7506/spkx1002-6630-201802026

• 生物工程 • 上一篇    下一篇

多基因DNA条形码鉴定6 个鳗鱼物种

陈文炳,邵碧英,缪婷玉,彭娟,陈彬,张志灯,江树勋   

  1. (福建出入境检验检疫局检验检疫技术中心,福建省检验检疫技术研究重点实验室,福建?福州 350003)
  • 出版日期:2018-01-25 发布日期:2018-01-05
  • 作者简介:陈文炳,邵碧英,缪婷玉,彭娟,陈彬,张志灯,江树勋
  • 基金资助:
    福建省科技计划农业引导性项目(2015N0016)

Identification of Six Eel Species Using Polygenic DNA Barcoding

CHEN Wenbing, SHAO Biying , MIAO Tingyu, PENG Juan, CHEN Bin, ZHANG Zhideng, JIANG Shuxun   

  1. (Fujian Provincial Key Laboratory of Inspection and Quarantine Technology Research, Inspection and Quarantine Technology Center, Fujian Entry-Exit Inspection and Quarantine Bureau, Fuzhou 350003, China)
  • Online:2018-01-25 Published:2018-01-05

摘要: 建立6?种鳗鱼的物种多基因DNA条形码精准鉴定方法。以鳗鱼DNA为模板,采用3?对通用引物对6?种鳗鱼的3?个基因(16S rRNA、Cyt b、COⅠ)部分DNA片段进行聚合酶链式反应扩增、测序,结果6?种鳗鱼各获得3?条16S rDNA(638~643?bp)、Cyt?b(464~466?bp)、COⅠ(705~707?bp)基因部分DNA序列,从中选取各物种序列同源的片段设计3 对新引物对6 种鳗鱼的16S rRNA、Cyt b、COⅠ基因部分DNA片段进行PCR扩增,其产物大小分别为504~507、400、609?bp,再从各片段中筛选出具有6?种鳗鱼物种特异性强的、碱基数分别为262、280、300?bp的3?条DNA片段序列,作为6?种鳗鱼物种的3?个基因的标准DNA条形码,应用DNAMAN?V6软件进行同源性分析,并通过GenBank数据库的比对验证,制定了供检测实践用的同源率判别指标,建立鳗鱼物种的多基因条形码检测方法。应用该方法对30?个待检鳗鱼样品进行检测,结果表明,各样品基于3?个基因DNA条形码的比对,符合同源率指标,物种判别结果互相吻合,从而精准判别样品的所属物种。该方法稳定、精准、易于操作,可应用于6?种鳗鱼的物种鉴定,值得推广。

关键词: 16S?rRNA基因, Cyt b基因, COⅠ基因, 多基因DNA条形码, 鳗鱼物种鉴定

Abstract: This paper develops an accurate method to identify six eel species including Anguilla rostrata, A. anguilla, A. japonica, A. mossambica, A. marmorata, and A. bicolor pacifica using polygenic DNA barcoding. Three pairs of universal primers were used to amplify partial fragments of the 16S rRNA, Cyt b and COⅠgenes of the eel species with the total genomic DNA from eel as a template and the amplified products were sequenced. As a result, 16S rDNA (638–643 bp), Cyt b (464–466 bp) and COⅠ(705–707 bp) gene fragments were obtained for each species. Homologous sequences of these genes were found. Three new primer pairs were designed to amplify 638–643, 400 and 609 bp fragments of the 16S rRNA, Cyt b and COⅠ genes by PCR, respectively. The specific 16S rRNA, Cyt b and COⅠfragments of 262, 280 and 300 bp were selected as standard DNA barcodes for the identification of eel species. Homology analysis was performed using DNAMAN software (version 6) and the homology for species discrimination was developed by aligning the DNA barcodes with the GenBank database. Analysis of 30 samples from six eel species by the developed method showed that the species were consistently identified using the DNA barcodes. In conclusion, the method is stable, precise and easy to operate and can be applied to the identification of six species of eels.

Key words: 16S rRNA, Cyt b, COⅠ, polygenic DNA barcode, identification of eel species

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