DNA barcoding coupled to HRM analysis as a new and simple tool for the authentication of Gadidae fish species

Abstract

This work aimed to exploit the use of DNA mini-barcodes combined with high resolution melting (HRM) for the authentication of gadoid species: Atlantic cod (Gadus morhua), Pacific cod (Gadus macrocephalus), Alaska pollock (Theragra chalcogramma) and saithe (Pollachius virens). Two DNA barcode regions, namely cytochrome c oxidase subunit I (COI) and cytochrome b (cytb), were analysed in silico to identify genetic variability among the four species and used, subsequently, to develop a real-time PCR method coupled with HRM analysis. The cytb mini-barcode enabled best discrimination of the target species with a high level of confidence (99.3%). The approach was applied successfully to identify gadoid species in 30 fish-containing foods, 30% of which were not as declared on the label. Herein, a novel approach for rapid, simple and cost-effective discrimination/clustering, as a tool to authenticate Gadidae fish species, according to their genetic relationship, is proposed.

Keywords: COI; Codfish; Cytb; DNA barcode; Gadoid species; High resolution melting analysis; Real-time PCR.