The Influence of DNA Extraction Methods on Species Identification Results of Seafood Products

Abstract

In terms of species identification, the ultimate aim of extracting DNA is the subsequent amplification of the selected marker; therefore, the quality and quantity of the extracted DNA must be sufficient for PCR-based methods. The purpose of this study is to compare five DNA extraction methods according to the parameters of quantity, quality and simplicity, among others, in order to determine the most suitable method for identification for Cephalopoda, Gadiformes and Pleuronectiformes. The Wizard DNA clean-up system kit (Promega), MPure-12TM automated nucleic acid purification system (MP Biomedicals), Chelex 100 resin (Biorad), DNeasy blood and tissue kit (Qiagen) and a swab method were examined. The obtained DNA quantity was determined by fluorescence, and quality was evaluated with ratios of absorbance of A260/A280 and A260/A230 by agarose gel visualization of the extracts and by analyzing the success of PCR amplifications of 720 bp fragments of cytochrome c oxidase I (COI) for Cephalopods and 465 bp fragments of cytochrome b for Gadiformes and Pleuronectiformes. Statistical results confirmed significant differences between the tested methods according to yield, efficiency and purity and no significant differences with respect to the species employed. The best yields were obtained with the Wizard kit, whereas other methods stand out in terms of their affordability (Chelex) and automation (Mpure).

Keywords

• DNA extraction methods
• Cephalopoda
• Gadiformes
• Pleuronectiformes
• polymerase chain reaction (PCR)
• sequencing