Multiplex PCR Method for Use in Real-Time PCR for Identification of Fish Fillets from Grouper (Epinephelus and Mycteroperca Species) and Common Substitute Species

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• 作者：Michele Trotta ; Susana Schö ; nhuth ; Tiziana Pepe ; M. Luisa Cortesi ; Antonio Puyet ; and José ; M. Bautista
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2005
• 出版时间：March 23, 2005
• 年：2005
• 卷：53
• 期：6
• 页码：2039 - 2045
• 全文大小：180K
• 年卷期：v.53,no.6(March 23, 2005)
• ISSN：1520-5118

文摘

Mitochondrial 16S rRNA sequences from morphological validated grouper (Epinephelus aeneus,E. caninus, E. costae, and E. marginatus; Mycteroperca fusca and M. rubra), Nile perch (Latesniloticus), and wreck fish (Polyprion americanus) were used to develop an analytical system for groupdiagnosis based on two alternative Polymerase Chain Reaction (PCR) approaches. The first includesconventional multiplex PCR in which electrophoretic migration of different sizes of bands allowedidentification of the fish species. The second approach, involving real-time PCR, produced a singleamplicon from each species that showed different T_m values allowing the fish groups to be directlyidentified. Real-time PCR allows the quick differential diagnosis of the three groups of species andhigh-throughput screening of multiple samples. Neither PCR system cross-reacted with DNA samplesfrom 41 common marketed fish species, thus conforming to standards for species validation. Theuse of these two PCR-based methods makes it now possible to discriminate grouper from substitutefish species.Keywords: Multiplex PCR; 16S rRNA; Epinephelus spp.; Mycteroperca spp.; Lates niloticus; Polyprionamericanus; real-time PCR; species identification