Scallops (Bivalvia, Pectinidae) are among the most valuable source of marine food. With about 350 extant species distributed worldwide and a total global production comprising 18 species, the development of proper species-level identification assays is imperative. DNA barcoding has proven to be a useful tool in species identification. A partial region at the 5′ end of the mitochondrial cytochrome c oxidase subunit I (COI) gene, known as the “Folmer region,” was proposed as the most suitable DNA barcoding marker. However, Folmer primers have failed to amplify polymerase chain reaction (PCR) products in different organisms, including scallops. Searching for an alternative barcoding gene region, we analyzed the complete mitochondrial 16S rRNA gene in 15 scallop species. We found that the interspecific variation at the 5′ end is twice as high as that at the 3′ end. Based on that evidence, we designed a novel Pectinidae family-specific primer set, aiming to amplify a partial region at the 5′ end of the 16S rRNA gene, and tested its suitability as a barcoding tool. A neighbor-joining analysis identified correctly 100 % of the scallop specimens analyzed, with high bootstrap support. Our new primers are well suited for DNA barcoding analysis and may contribute to scallop food industry surveys, as well as routine taxonomic surveys.