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

Francesco Scollo; Leticia A Egea; Alessandra Gentile; Stefano La Malfa; Gabriel Dorado; Pilar Hernandez

doi:10.1016/j.foodchem.2016.06.086

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

Food Chem. 2016 Dec 15:213:388-394. doi: 10.1016/j.foodchem.2016.06.086. Epub 2016 Jun 25.

Authors

Francesco Scollo¹, Leticia A Egea², Alessandra Gentile³, Stefano La Malfa⁴, Gabriel Dorado⁵, Pilar Hernandez⁶

Affiliations

¹ Instituto de Agricultura Sostenible (IAS-CSIC), Consejo Superior de Investigaciones Científicas, Alameda del Obispo s/n, 14080 Córdoba, Spain; Dipartimento di Agricoltura, Alimentazione e Ambiente (Di3A), Via Valdisavoia 5, Università degli Studi di Catania, 95123 Catania, Italy. Electronic address: fscollo@unict.it.
² Instituto de Agricultura Sostenible (IAS-CSIC), Consejo Superior de Investigaciones Científicas, Alameda del Obispo s/n, 14080 Córdoba, Spain; Dep. Bioquímica y Biología Molecular, Campus Rabanales C6-1-E17, Campus de Excelencia Internacional Agroalimentario (ceiA3), Universidad de Córdoba, 14071 Córdoba, Spain. Electronic address: z32ayegl@uco.es.
³ Dipartimento di Agricoltura, Alimentazione e Ambiente (Di3A), Via Valdisavoia 5, Università degli Studi di Catania, 95123 Catania, Italy; Horticulture and Landscape College, Hunan Agricultural University, Changsha, Hunan, China. Electronic address: gentilea@unict.it.
⁴ Dipartimento di Agricoltura, Alimentazione e Ambiente (Di3A), Via Valdisavoia 5, Università degli Studi di Catania, 95123 Catania, Italy. Electronic address: slamalfa@unict.it.
⁵ Dep. Bioquímica y Biología Molecular, Campus Rabanales C6-1-E17, Campus de Excelencia Internacional Agroalimentario (ceiA3), Universidad de Córdoba, 14071 Córdoba, Spain. Electronic address: bb1dopeg@uco.es.
⁶ Instituto de Agricultura Sostenible (IAS-CSIC), Consejo Superior de Investigaciones Científicas, Alameda del Obispo s/n, 14080 Córdoba, Spain. Electronic address: phernandez@ias.csic.es.

PMID: 27451195
DOI: 10.1016/j.foodchem.2016.06.086

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.

Keywords: Molecular markers; Olea europaea; Olive oil; Traceability; qRT-PCR.

MeSH terms

DNA / genetics*
Humans
Olive Oil / chemistry*
Polymerase Chain Reaction / methods*
Reproducibility of Results

Substances

Olive Oil
DNA