Intrav arietal DNA polymorphisms in grapevine (Vitis vinifera L.)

The high genetic variability of grapevines enabled the clear distinction of varieties by using different molecular markers as RFLPs, RAPDs and STRs. These markers, however show some limits in the differentiation of clones belonging to the same variety of Vitis vinifera. In this study a different approach was considered, the AFLP (Amplification Fragment Length Polymorphism) method was used in order to find intravarietal genetic diversity and to develop a method for the identification of clones. About sixty primer pairs on nine different international grape cultivars such as Chardonnay, Cabernet Sauvignon, Merlot were chosen, and in each reaction an average of 9 fragments were polymorphic. A number of clones belonging to these varieties were typed using the more informative primer combinations. Finally, only clones belonging to the varieties Pinot blanc, Pinot noir and Pinot gris were analysed using about further fifty primer pairs in order to investigate their genetic diversity. Amplification products were analysed with different techniques: by loading on 4.5% acrylamide gel and isotope detection, by loading on 8% acrylamide gel and detection by LI-COR (MWG-Biotech), and by capillary electrophoresis on the ABI PRISM 310 (Perkin Elmer) automated fluorescent sequencer using GeneScan 3.1 analysis. Some of the polymorphisms detected among clones were not reproducible when using newly digested DNA samples. Nevertheless, other polymorphisms have been confirmed using the different electrophoresis systems, and will be considered in the development of SCARs. All the data were used to estimate the genetic distance among the grapevines. The possible application of AFLPs as molecular markers for clonal selection, identification, certification and protection was discussed.