Molecular and histochemical analysis of Vitis vinifera L. cultivars

The genotypic characterization of grapevine cultivars was carried out on 20 wine grape cultivars, 10 with white and 10 with black berries using RAPD analysis. DNA was extracted from young leaves harvested in a collection field located in Grugliasco (Piemonte). A total of 25 oligonucleotide primers (10-mers) from the kits A, M, P, O from Operon Technologies Inc. (USA) were used. Almost all of the primers tested produced polymorphic bands in the size range between 450 bp and 2000 bp. Using polymorphic markers the genotypes were compared and a phylogenetic tree was generated by cluster analysis using Statistica software. Moreover, pericarp structures were studied on two black berried and two white berried cultivars using the new technique of ethylene glycol cryostabilization and inclusion in hydrophilic resin (GMA: Technovit 7100, Kulzer). This technique preserves the cellular structures and nuclei even in ripe berries. The aim was to investigate, under the same cultural growth conditions, the differences between white berried and black berried cultivars: examining in particular wax thickness and epicarp structure, all factors that are correlated to Botrytis cinerea resistance. Slices of 2.5 mm thickness were stained with PAS reaction, Toluidine blue, Nile red and DAPI, and were observed by LM.