PCR-RFLP analysis of nuclear nontranscribed spacer for mackerel species identification
PCR-RFLP analysis of nuclear nontranscribed spacer for mackerel species identification
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DOI：
10.1021/jf0484881
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发表时间：
2005-02-09
影响因子：
6.1
通讯作者：
Aranishi, F
中科院分区：
文献类型：
--
作者：
Aranishi, F
鲭鱼以新鲜和冷冻形式以及加工海鲜在世界各地销售，日本鲭鱼、太平洋鲭鱼和大西洋鲭鱼这三种鲭鱼已构成日本鲭鱼绝对消费的重要组成部分。本研究的目的是开发一种快速可靠的方法，不仅可以使用鲭鱼属特异性引物通过 PCR 扩增将鲭鱼与相关鲭鱼区分开，而且还可以通过 PCR-RFLP 分析鉴定三种鲭鱼物种。通过对鲭鱼核 5S 核糖体 RNA 基因 (5S rDNA) 的核苷酸序列进行比对，可以选择鲭鱼属特异性的寡核苷酸引物。这些引物能够扩增来自 S. japonicus、S. australasicus 和 S. scombrus 的 5S rDNA 的非转录间隔区 (NTS)，而其他鲭鱼则没有扩增。使用 ScaI 核酸内切酶对 PCR 产物进行 RFLP 分析，生成每个鲹鱼物种的独特限制模式。这种使用鲭鱼属特异性引物的简单、稳健且可重复的 PCR-RFLP 技术可以作为常规食品检验计划，以执行市售鲭鱼的标签规定。
Scomber mackerel have been marketed in fresh and frozen forms and as processed seafood worldwide, and three species of Japanese mackerel S. japonicus, Pacific mackerel S. australasicus, and Atlantic mackerel S. scombrus have constituted a significant part of absolute Scombrid consumption in Japan. The present study was undertaken to develop a rapid and reliable method not only for differentiation of Scomber mackerel from related Scombrid fish by PCR amplification using Scomber genus-specific primers but also for identification of three Scomber mackerel species by PCR-RFLP analysis. Alignment of nucleotide sequences of the nuclear 5S ribosomal RNA gene (5S rDNA) among Scombrid fish allowed the selection of oligonucleotide primers specific for the Scomber genus. These primers enabled amplification of the nontranscribed spacer (NTS) of the 5S rDNA from S. japonicus, S. australasicus, and S. scombrus, whereas no amplification was demonstrated from other Scombrid fish. RFLP analysis of the PCR products with ScaI endonuclease generated unique restriction patterns for each Scomber species. This simple, robust, and reproducible PCR-RFLP technique using Scomber genus-specific primers can serve as a routine food inspection program to enforce labeling regulations of marketed Scombrid fish.