Species-specific mitochondrial DNA marker, length polymorphism and heteroplasmy in the clam Macridiscus

In Genus Macridiscus there are three morphologically very similar species: M .melanaegis, M .multifarious and M .semicancellata, the first 2 are sympatric. It is difficult to distinguish these species solely by morphological characters. In this article, genomic DNA was extracted from ethanol preserved foot tissues by a CTAB extraction. A DNA fragment from mitochondrial control region of cytochrome b gene(Cyt b) was used for polymerase chain reaction(PCR) amplification with primers Macr_CO1_F16834 and Cytb_Macr_R2. Sequencing of PCR products revealed a 363 bp variable number of tandem repeat (VNTR) close to 3′end of the control region leading to length polymorphism in 44% heteroplasmic individuals in M .melanaegis. While in the control region of M .semicancellata, two short VNTRs,14bp and17bp each, were found close to 3′ end with length polymorphism between individuals. M .multifarious demonstrated a length conserved control region. These data proved that the three species of Macridiscus could be efficiently and accurately identified via PCR with primer pair Macr_CO1_F16834 and Cytb_Macr_R2.