A Robust DNA Isolation Protocol from Filtered Commercial Olive Oil for PCR-Based Fingerprinting

Abstract

Extra virgin olive oil (EVOO) has elevated commercial value due to its health appeal, desirable characteristics and quantitatively limited production, and thus it has become an object of intentional adulteration. As EVOOs on the market might consist of a blend of olive varieties or sometimes even of a mixture of oils from different botanical species, an array of DNA-fingerprinting methods have been developed to check the varietal composition of the blend. Starting from a comparison between publicly available DNA extraction protocols, we set up a timely, low-cost, reproducible and effective DNA isolation protocol, which allows an adequate amount of DNA to be recovered even from commercial filtered EVOOs. Then, in order to verify the effectiveness of the DNA extraction protocol herein proposed, we applied PCR-based fingerprinting methods starting from the DNA extracted from three EVOO samples of unknown composition. In particular, genomic regions harboring nine simple sequence repeats (SSRs) and eight genotyping-by-sequencing-derived single nucleotide polymorphism (SNP) markers were amplified for authentication and traceability of the three EVOO samples. The whole investigation strategy herein described might favor producers in terms of higher revenues and consumers in terms of price transparency and food safety.

Molecular Approaches to Agri-Food Traceability and Authentication: An Updated Review

“…These panels are available in different species of agri-food interest such as grapevine, olive, pulses, cacao, and coffee [ 76 , 77 , 78 , 79 , 80 ]. Most of the works using a SNP-based traceability approach have focused on olive oil analysis [ 40 , 41 ], however, SNPs have been efficiently employed in differentiating Arabica and Robusta coffee varieties [ 81 ], in the authentication of Portuguese wine [ 42 ], and in the identification of Nebbiolo variety in musts and wines [ 43 ].…”

Microsatellite Markers in Olives (Olea europaea L.): Utility in the Cataloging of Germplasm, Food Authenticity and Traceability Studies

“…Over the last decade, different isolation protocols and kits have been tested and modified for better DNA extraction from fruits and oils, and these studies highlight the importance of DNA quality and its impact on molecular marker-based tests [116][117][118][119][120]. Recently, Piarulli et al [121] compared four DNA isolation methods referenced in the literature and came up with a modified method based on the work of Consolandi et al [122] for the extraction of DNA from extra virgin olive oil in a much smaller time frame (4 h as compared with the 30 h reported) and involving low-cost options. A washable and reusable miniaturized device has been developed as well and tested for highly efficient DNA purification from olive oil, providing an increased surface-area-to-volume ratio when compared with other approaches, allowing highly efficient DNA purification and concentration from samples with minute DNA contents [123].…”

Sustainable Palm Oil—The Role of Screening and Advanced Analytical Techniques for Geographical Traceability and Authenticity Verification

“…The contrasting results indicate that establishing reliable tools for traceability and authenticity analysis is still an important active area of research where a consensus on the most effective method(s) will take time to develop. The main challenge in using DNA markers has been establishing a DNA extraction and analysis method, mostly based on polymerase chain reaction (PCR), which, even now for olive oil, is being improved for more effective analysis of the commodity [103]. The use of DNA analysis is more complicated for some vegetable oils (e.g., palm oil) that go through a complex milling and refining process involving high temperatures, which can cause extensive fragmentation of the DNA and compromise its amplification in a PCR assay.…”