The development of species-specific AFLP-derived SCAR and SSCP markers to identify mantis shrimp species

Mantis shrimp has become commercially valuable in many countries, while the commercially aquaculture still unsuccessful. The stable supply of the species-specific markers for precise identification can play a key role of foods authentication as well as restoring/enhancing mantis shrimp stocks in future. The aim of this research was to identify species-specific markers for Squillid and Harpiosquillid mantis shrimp taxa using Amplified fragment length polymorphism-Single strand conformation polymorphism (AFLP-SSCP) approaches. Selective amplification would be substituted as a total of 40 primer combinations was performed using either three-base (i.e., EcoRI+3 and MseI+3 in 20 primer combinations) or two-base (i.e., EcoRI+2 and MseI+2 in 20 primer combinations) selective primers. These had been size-fractionated via 6% denaturing polyacrylamide gel electrophoresis, ten AFLP fragments exhibiting species or genus-specific characteristics were cloned, sequenced, and GenBank interrogated. A primer pair was designed and their specificity was tested versus the genomic DNA of various species. Results show that the primer E₊₂-13/M₊₂-13Hr₁₅₈ generated PCR products for just H. harpax, while E₊₃-14/M₊₃-2HhHr₁₅₁ and E₊₂-13/M₊₂-13Hh₁₅₀ generated PCR products for both H. harpax and H. raphidea and not others (i.e., M. nepa, O. oratoria, and E. woodmasoni). SSCP was then applied in order to differentiate between H. harpax and H. raphidea. These SSCP results indicate that species can be differentiated based on polymorphic fragment nucleotides. Indeed, primers E₊₂-13/M₊₂-13Hr₁₅₈, E₊₃-14/M₊₃-2HhHr_151, and E₊₂-13/M₊₂-13Hh₁₅₀ were all successfully confirmed as present in processed mantis shrimp samples (i.e., saline-preserved and heat-dried). These results provide new species-specific markers for mantis shrimp identification.