Genetic authentication: Differentiation of hazelnut cultivars using polymorphic sites of the chloroplast genome

High price differences between European (e.g., from Italy) and Caucasian (e.g., from Georgia) cultivars motivate food fraud in hazelnuts. In this work, we present two targeted methods for differentiation and grouping different hazelnut cultivars based on polymorphic variations in the chloroplast genome. After sequencing the chloroplast genome of 12 hazelnut cultivars two indels with lengths of 118 bp and 115 bp as well as single nucleotide polymorphisms (SNPs) were identified. Based on the indels, 109 samples from seven countries, together representing 19 cultivars, could be classified into two groups: Europe (Germany, France, Spain and Italy) and Caucasus (Georgia, Azerbaijan). Turkish cultivars show ambiguous behavior. Using capillary gel electrophoresis (CGE), a method was developed to semi-quantitatively estimate mixtures of these two groups. In addition, by using agarose gel electrophoresis (AGE) and respective analysis via grayscales a semi-quantitative estimation of adulteration was also possible. Both methods show highly effective results for detecting blends of hazelnuts with a limit of detection (LOD) of 5% and 10% admixture. Furthermore, Georgian hazelnut cultivars can be detected by restriction fragment length polymorphism (RFLP) analysis using one of the identified SNPs. In summary, both methods can be used for differentiation and semi-quantitative estimation of mixtures of hazelnuts from different growing regions in the context of authenticity control and routine analysis.