Absolute quantification of olive oil DNA by droplet digital-PCR (ddPCR): Comparison of isolation and amplification methodologies. (15th December 2016)

• Record Type: Journal Article
• Authors: Scollo, Francesco; Egea, Leticia A.; Gentile, Alessandra; La Malfa, Stefano; Dorado, Gabriel; Hernandez, Pilar
• Is Part Of: Food chemistry. Volume 213(2016)
• Page Start: 388
• Page End: 394
• Publication Date: 2016-12-15
• Subjects: Olea europaea -- Olive oil -- Traceability -- Molecular markers -- qRT-PCR; AFLP amplified fragment length polymorphism -- ddPCR droplet digital-PCR -- PDO Protected Designation of Origin -- PGI Protected Geographical Indication -- qRT-PCR quantitative Real-Time PCR -- SCAR sequence characterized amplified region -- SNP single nucleotide polymorphism -- SSR simple sequence repeat (microsatellite); Food -- Analysis -- Periodicals; Food -- Composition -- Periodicals; 664
• DOI: 10.1016/j.foodchem.2016.06.086
• Languages: English
• ISSNs: 0308-8146

Abstract

Highlights: Defining the origin of olive oil represents a key aspect to ensure product traceability. DNA quality is often unsatisfactory and its absolute quantification may not be possible. Different DNA extraction protocols were evaluated by qRT-PCR and ddPCR. Qualitative aspects and amplified DNA's copies number were quantified. Abstract: Olive oil is considered a premium product for its nutritional value and health benefits, and the ability to define its origin and varietal composition is a key step towards ensuring the traceability of the product. However, isolating the DNA from such a matrix is a difficult task. In this study, the quality and quantity of olive oil DNA, isolated using four different DNA isolation protocols, was evaluated using the qRT-PCR and ddPCR techniques. The results indicate that CTAB-based extraction methods were the best for unfiltered oil, while Nucleo Spin-based extraction protocols showed greater overall reproducibility. The use of both qRT-PCR and ddPCR led to the absolute quantification of the DNA copy number. The results clearly demonstrate the importance of the choice of DNA-isolation protocol, which should take into consideration the qualitative aspects of DNA and the evaluation of the amplified DNA copy number.