Use of mtDNA Direct Polymerase Chain Reaction (PCR) Sequencing and PCR−Restriction Fragment Length Polymorphism Methodologies in Species Identification of Canned Tuna

Abstract

Identification of six canned tuna species using DNA-based methodology was studied. DNA was degraded during the canning process of fish muscle: DNA fragment sizes that ranged from <100 up to 200 bp were obtained from canned tuna muscle, whereas DNA sizes for frozen tuna muscle ranged from <100 up to 20 000 bp. Amplification of DNA from canned tuna muscle was carried out using primers flanking a region of cytochrome b gene of 126 bp. Sequences from PCR-amplified DNA of six tuna species were studied for polymorphic sites; seven diagnostic positions were identified in this fragment for the species studied. The suitability of a genetic distance measurement with phylogenetic tree construction method for the identification of canned tuna species using two cytochrome b sequences (299 and 126 bp) was studied. PCR-amplified DNA from canned tuna was also analyzed by using three restriction endonucleases, BsiYI, MboI, and MnlI. The restriction fragments allowed for the identification of the six tuna species studied. Keywords: Canned tuna; species identification; cytochrome b; genetic distance; RFLP−PCR

Development of a genetic method for the identification of salmon, trout, and bream in seafood products by means of PCR–RFLP and FINS methodologies

“…This was the case of cans or smoked products, where fragments of little sizes were ampliWed. Quinteiro et al [10] established a maximum fragment size in canned products of 176 bp to ensure the ampliWcation. Other authors, under certain conditions, ampliWed fragments higher than 200 bp from canned products [28][29][30][31].…”

“…The main disadvantage of all the previous studies is that these cannot be applied to highly processed products such as canned foods, because the ampliWed PCR product is longer than 200 bp. In this kind of products it is not possible to amplify DNA fragments of such a size [10][11][12][13]. Moreover, these methods do not include all the possible substitute species, and this fact can produce incorrect or not assignations.…”

A Comparison of Eight Methods for DNA Etraction from Processed Seafood Products

“…Furthermore, the DNA size in these seafood products was smaller than 650 bp, with parts of the fragments being larger than 358 bp. The different result on the fragment size in the aforementioned study of Quinteiro et al (1998) and Chapela et al (2007) may be partly caused by the sample tested in these studies being a different species, with different matrices and different processing conditions.…”

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“…In recent years, species-identification techniques based on stable DNA molecules carrying the specific genomic information have gained recognition in food sciences, particularly fish origin. In view of its reliability, RFLP is standard technique used for genera, species or subspecies discrimination as studied in fishes by Quinteiro et al (1998); Wolf et al (2000); Russell et al (2000).…”