COI-high resolution melting analysis for discrimination of four fish species in the family Notopteridae in Thailand

Species identification is an essential and vital basis for biological research. Since several fish species have similar appearances and shapes at different developmental stages and researchers lack experience identifying these fish species, molecular techniques are needed. In Thailand, fish species of the family Notopteridae consist of two genera and four species, including Notopterus notopterus, Chitala ornata, Chitala blanci, and Chitala lopis. The combination of the cytochrome c oxidase I (COI) gene sequences with a nonsequencing approach, high-resolution melting (HRM), was developed to discriminate four target species in a rapid, reliable and cost-effective manner. One of three designed primer pair sets, Notop-HRMF1/Notop-HRMR1, was the most appropriate and provided four distinct melting curves. In addition, ten fish species of other families were used for the specificity test, and only the four target species of Notopteridae were specific for this primer pair set. The results of this study indicated that COI-HRM analysis is an efficient approach for identifying species of Notopteridae in Thailand. Applications of this technique may be used to investigate species of this family in processed fish products or larval fish species in several spawning grounds for sustainable fishery resource management.