Fingerprinting and genetic diversity of Olea europaea L. ssp. europaea accessions from the cultivar Galega using RAPD markers

• Molecular markers helped clarifying accessions’ certification.
• RAPDs discriminated Galega genotypes from other synonymous genotypes.
• The three different Galega synonymous we studied are different cultivars.
• Somatic mutations’ accumulation probably created within group variability.
• Different genotypes’ origin also could create within group variability.

Olea europaea ssp. europaea (Oleaceae) grows in Mediterranean countries. The Galega represents one of the most common Portuguese olive tree cultivars, and is used in five out of the six PDO (Protected Designation of Origin) regions. The fruits are used for olive oil and for fruit canning, besides which the cultivar is drought tolerant. The cultivar certification is done through phenotypic characteristics, but environmental influence might blur informativeness, and molecular markers could help in clarifying cultivar accessions certification.A total of 75 putative Galega accessions considered as belonging to the Galega cultivar, three previously phenotypically identified as GGN (Galega Grada Normal), GGS (Galega Grada de Serpa) and GGE (Galega Grada de Évora), and one as a non-Galega blind control were analyzed. Twenty RAPD primers amplified 144 bands and 32% were polymorphic. A low expected heterozygosity was estimated (10%), as expected for clonal propagated plants.The Principal Component Analysis (PCA) clearly identified two groups. Group A was comprised of all the putative Galega accessions including GGN, but not individual #502. Group B included NG, GGE, GGS and #502. The analysis of molecular variance confirmed a very high differentiation between the two groups (88%), but also some within groups’ diversity (12%). Group A individuals cluster closely to GGN. Assuming that the GGN individual represents a Galega cultivar tree, we might assume that all the A group trees are Galega cultivar accessions. Nevertheless, some within group variability exists, may be due to the accumulation of somatic mutations and/or a putative origin from different, but related genotypes. Finally, the GGS, the GGE, and the #502 plot together with the individual identified as non Galega cultivar tree. Group B trees certainly had a very different genetic origin compared to the group A genotypes, which confirms some differences found in the morphologic characteristics compared to the ordinary Galega cultivar, reflected in the name differences.The pairwise matrix computed with the Dice similarity index was also used to obtain a dendrogram. Results were similar, but slightly different from the PCA. Three clusters were obtained. This indicates that the B group could be split into two sub-groups. A future study using a broader sampling and higher discriminant molecular markers, such as microsatellites, would help to reveal the phylogeny among the 3 cultivars (GGN, GGS and GGE) and to clarify if the intra-variability of those cultivars is geographically related.