Development of a rapid and simple method for identification of Haliotis gigantea using species-specific PCR
Chun Mae Dong, Mi-Nan Lee, Jung-Ha Kang, Jung Youn Park, Bo-Hye Nam, Jae Koo Noh, Pil-Youn Kim, Young Ah Cho and Eun-Mi Kim
Biotechnology Research Division, National Institute of Fisheries Science, Busan 46083, Korea Jeju Special Self-Governing Province Fisheries Resources Research Institute, Jeju 697-914, Korea Dongnam Local Statistical Ulsan Office, Statistics Korea, Ulsan
This study was performed to identify rapidly Haliotis gigantea using polymerase chain reaction with species-specific primers. Around 680 bp of the mitochondrial ND5 gene region from four Haliotis species were aligned and species-specific forward primer was designed based on the single nucleotide polymorphism (SNP) from H. gigantea. The optimal PCR conditions were selected by cross reactivity using gradient PCR method from 55°C to 66°C. Species-specific PCR (SS-PCR) amplification reactions with two pairs of primers were performed for a five specimens of Haliotis species. SS-PCR leads to a species specific amplification of a 1,006 bp fragment in H. discus hannai, H. discus discus, H. madaka and 786 bp in H. gigantea, respectively. The two different sizes of each PCR products can be quickly and easily detected by single gel electrophoresis. The sensitivity of the SS-PCR was up to 1ng/μl DNA as a starting concentration in H. gigantea tested. Therefore SS-PCR technique with species-specific primer based on SNP could be a powerful tool for discrimination of H. gigantea and can contribute to the prevention of falsified labeling of this species.
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