Peru is one of the world’s leading fishing nations and its seafood industry relies on the trade of a vast variety of aquatic resources, playing a key role in the country’s socio-economic development. DNA barcoding has become of paramount importance for systematics, conservation, and seafood traceability, complementing or even surpassing conventional identification methods when target organisms show similar morphology during the early life stages, have recently diverged, or have undergone processing. Aiming to increase our knowledge of the species diversity available across the Peruvian supply chain (from fish landing sites to markets and restaurants), we applied full and mini-barcoding approaches targeting three mitochondrial genes (COI, 16S, and 12S) and the control region to identify samples purchased at retailers from six departments along the north-central Peruvian coast. DNA barcodes from 131 samples were assigned to 55 species (plus five genus-level taxa) comprising 47 families, 24 orders, and six classes including Actinopterygii (45.03%), Chondrichthyes (36.64%), Bivalvia (6.87%), Cephalopoda (6.11%), Malacostraca (3.82%), and Gastropoda (1.53%). The identified samples included commercially important pelagic (anchovy, bonito, dolphinfish) and demersal (hake, smooth-hound, Peruvian rock seabass, croaker) fish species. Our results unveiled the marketing of protected and threatened species such as whale shark, Atlantic white marlin, smooth hammerhead (some specimens collected during closed season), shortfin mako, and pelagic thresher sharks. A total of 35 samples (26.72%) were mislabeled, including tilapia labeled as wild marine fish, dolphinfish and hake labeled as grouper, and different shark species sold as “smooth-hounds”. The present study highlights the necessity of implementing traceability and monitoring programs along the entire seafood supply chain using molecular tools to enhance sustainability efforts and ensure consumer choice.