Development of Rapid Identification Method for Asian Clam, Corbicula fluminea and Brackish Water Clam, Corbicula japonica Using Duplex PCR
Yong Hwi Kim, Bong Han Yun, Ho-Seop Han, Hye Jin Kim and In-Chul Bang
Department of Biology, College of Natural Sciences, Soonchunhyang University, Asan, Chungnam 31538, Republic of Korea
A duplex PCR primer set was developed for molecular biological species identification by making species-specific primers for Corbicula fluminea and Corbicula japonica distributed in Korea. The developed species-specific primers were designed to form an interspecies-specific band around C. fluminea 289 bp and C. japonica 143 bp by searching for a site of interspecies genetic variation appearing on the nucleotide sequence of the cytochrome c oxidase subunit 1 (co1) gene of mitochondrial DNA. A verification experiment was also performed on the duplex PCR primer set by measuring the amplification efficiency for each repeat number and genomic DNA (gDNA) concentration. The duplex PCR primer set finally established was found to require a minimum of 10 ng/μL of gDNA concentration when considering the sample amount and PCR reaction cycle in terms of economic feasibility, and 30 cycles were found to be suitable. Therefore, the duplex PCR primer set between C. fluminea and C. japonica developed in this study is judged to be able to perform accurate species identification quickly and conveniently by PCR band observation.
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