A quantitative comparison of two kits for DNA extraction from canned tuna

Abstract

The most common methods that can be used for species identification of tuna include methods based on detection of species-specific DNA via the polymerase chain reaction (PCR) method. The problem with DNA detection in processed products is the possibility of DNA fragmentation during the technological process. The quantity and quality of extracted DNA is a crucial step for species identification based on the DNA analysis. In this study, two DNA extraction methods (DNeasy Blood & Tissue Kit and DNeasy mericon Food Kit) for tuna DNA isolation were compared. Eight food products of canned tuna (three of them were declared as Thunnus albacares and five products were declared as Katsuwonus pelamis) with a different addition of various ingredients were tested. Furthermore, three different times of proteolysis (30 min, 60 min, overnight) for each sample and each extraction kit were evaluated. The DNA concentration was determined by a Qubit dsDNA HS Assay Kit fluorescence method and quantified using a Qubit fluorometer. The DNA purity was evaluated using the A260/A280 ratio of absorbances measured on a spectrophotometer. The main indicator of DNA quality and quantity was its amplifiability in the subsequent real-time PCR for Thunnus species, Thunnus albacares and Katsuwonus pelamis. Based on the results, both kits can be used for tuna species determination in highly heat-treated products with different composition, nevertheless, the DNeasy mericon Food Kit provided better statistical values in some parameters. The effect of different times of proteolysis was significant in most of the samples with regard to the crossing point values determined by real-time PCR.

Comparison of DNA Extraction Methods for the Detection of Canned Tuna Species with DNA Mini-Barcoding

“…All four extraction methods produced signifcantly diferent DNA concentration values except for DNeasy + PC and DNeasy MF, according to the Kruskal-Wallis H test with the post hoc Dunn's test (p < 0.05). Te signifcantly higher DNA concentration obtained with the DNeasy method as compared to the DNeasy MF method is consistent with a prior study comparing DNA extraction kits for canned tuna [28]. Similarly, Zahn et al [29] reported signifcantly higher DNA concentrations for shark cartilage samples extracted with the DNeasy method as compared to the DNeasy + PC method.…”

“…A previous study reported that the DNA extraction method and packing medium greatly infuenced the quality and quantity of DNA isolated from canned tuna [27]. Because of the wide variety of packing media (e.g., oil, water, salt, and other ingredients) used in the industry, canned tuna may contain varying amounts of PCR inhibitors that can negatively impact the sensitivity of the PCR reaction [28].…”

“…Te objective of this study was to compare the following four diferent commercial DNA extraction kits for use in CR mini-barcoding of canned tuna: Qiagen DNeasy Blood and Tissue Kit (current method), Qiagen DNeasy Blood and Tissue Kit + Qiagen PowerClean Pro Cleanup Kit, MP Biomedicals FastPrep-24 + Machery-Nagel NucleoSpin Tissue Kit, and the Qiagen DNeasy Mericon Food Kit. Tese kits were selected due to their success in previous studies examining species identifcation of various processed seafood products [5,21,28,29].…”

Testing of the Effect of Food Additives on the DNA Isolation from Mackerel and Fish Products

“…20 a Servusová a spol. 21 , kteří úspěšně izolovali DNA ze syrové i tepelně opracované svaloviny tuňáka žlutoploutvého (Thunnus albacares). Dosažené výsledky izolace DNA pro syrovou svalovinu byly řádově srovnatelné s našimi (51 ng μl -1 ; A 260 /A 280 = 1,8).…”

“…V práci Servusová a spol. 21 , kde byly porovnávány dva komerční kity (DNeasy ® mericon Food Kit a DNeasy ® Blood & Tissue Kit) bylo prokázáno, že DNeasy ® mericon Food Kit je vhodnější pro PCR analýzu ryb. Z výsledků experimentů bylo zřejmé, že amplifikace proběhla lépe ve vzorcích izolovaných tímto kitem, přestože koncentrace změřená spektrofotometricky byla nižší než u druhého kitu; DNeasy ® mericon Food Kit tak byl označen jako lepší pro izolace DNA z konzervovaných rybích výrobků.…”