Rapid species identification of Meretrix lusoria and three other Meretrix clams using PCR and RFLP analysis of the mitochondrial COII gene.

Abstract

Meretrix clams are among the most economically important species of bivalves in Eastern Asia and Taiwan. In the past, the identification of these hard clams depended on the shell morphology; however, species identification based on shell markings and shapes confuses the taxonomy of the Meretrix genus. The DNA molecular information is useful and can easily identify them accurately and quickly, especially the sequence of mitochondrial DNA. In this study, a PCR method to analyze the mitochondrial cytochrome c oxidase II (COII) gene was developed to rapidly identify of M. lusoria, M. lamarckii, M. lyrata and M. petechialis. Four different species of hard clams were easily distinguished by examining the COII PCR gel pattern using the naked eye. The limit of COII PCR detection was 39.06 pg, and it showed good specificity of primer. Further verification using RFLP suggested that four species of hard clams could be identified on the basis of the RFLP gel pattern produced by the digestion of one restriction enzyme, and these results were consistent with the PCR results. The ML phylogenetic analysis of COII grouped the four Meretrix species into different clades with strong bootstrap support. These results are consistent with those of the phylogenetic analysis of COI gene. In summary, this study’s COII PCR and RFLP method facilitates the rapid, accurate, and inexpensive identification of Meretrix species. These results suggest that this COII PCR method can be used for routine and rapid species identification on population, aquaculture and breeding management.