Abstract:
The hard clam Mercenaria mercenaria supports a $65 million-dollar aquaculture industry in the U.S.A. Sustainability of this industry depends on the development of genetically improved broodstock for high-quality seed production. Accurate identification of the species and varieties are needed to produce genetically stable families or lines. However, another hard clam species Mercenaria campechiensis is naturally distributed in the Southeast coast of the U.S.A. and sympatric with the above aquaculture species. Furthermore, these two species have no reproductive isolation, and their hybrids are fertile. The goal of this study was to develop an accurate, fast, and reliable genotypic tool to recognize M. mercenaria and M. campechiensis and their hybrids by associating with the phenotypic scoring. Clams were collected from 8 geographical locations along the east coast of the U.S.A. from Maine to Florida and the Gulf of Mexico (the Florida west coast). Phenotypic scoring revealed that clams from Maine were 100% M. mercenaria, while samples from Bradenton Beach, Florida were 100% M. campechiensis. Samples from all the other locations showed mixed phenotypic characteristics. PCR length polymorphism of partial amplification of 16S rRNA, 18S rRNA, CO!, ITSl, and ITS2 genes was not sensitive to identify the two species. All gene regions were appeared to be the same in length for both species and each gene region showed a different length: 1(approximately). RFLP-PCR analysis was proved to be an effective method to identify two species and their hybrids. For the amplified product of 16S rRNA, restriction enzyme Rsal generated two different size fragments for two clam species allowing species identification. The restriction enzyme Sty! digestion carried out for the PCR product of the ITS2 gene region generated two fragments for M. campechiensis, a fragment for M. mercenaria and three fragments for the hybrids. This study developed an accurate and efficient marker to identify clam species and their hybrids which can be used to overcome the confusions arising in clam aquaculture, assist the clams breeding program and improve the conservation implication of these species, especially M. campechiensis which are now difficult to find. The utility of RFLP-PCR marker could be considered as an effective method in species and hybrid identification because of the low cost, ease to perform in simple laboratories and low skill requirements.
