Studies On The Application Of RAPD Molecular Markers To The Classification Of Grape Varieties

In this study, the genetic diversity of 29 grape varieties cultivated in Sichuan Agricultural University was studied by random amplified polymorphic DNA(RAPD).The relationship of the 29 grape varieties was analyzed. The results were as follows:1. The improved CTAB method developed in this study was the best one to extract vitis genomic DNA. DNA quality was high, and OD260/OD280 of DNA was 1.73-1.86. And DNA without getting rid of RNA can be used for RAPD amplification.2. The suitable reaction system and amplification procedure were found. The reaction mixture contained diluted 10 times template DNA in a 25 μL reaction volume with 2.5 μL lOxreaction buffer, 1.5mmol/L MgCl2, 250μmol/L dNTP and 1.25U Taq DNA polymerase and 1.5μmol/L primer covered with a drop of mineral oil. Amplification was performed in a PT-100 thermal cycler programmed for 40 cycles (94 ℃ for 1 min; 35 ℃ for 1 min; 72 ℃ for 2 min) followed by an extension at 72 ℃ for 8 min.3. 126 repeatable loci (including 111 polymorphic loci) were amplified using 8 random primers in 29 grape resources. The percentage of polymorphic bands was 88.1%,13.9 polymorphic bands per primer were amplified. 3-34 repeatable loci were amplified with each primer. As analyzed by NTSYSpc version 2.10, the genetic distance between 'Jinyu' and 'Ruby Seedless' was the smallest(0.1093). The distance between 'Superior Seedless' and 'Zhaohongti' was the biggest (0.8462). UPGMA clustering analysis based on genetic similarity showed that 29 materials could be classified into 5 groups. 7 materials ('Fenghou', 'Heimi', 'Nongke Ⅲ' , 'Wuhezaohong', '8612' ,'Gaoqi 'and 'Beni Fuji') belonged to group Ⅰ ; 18 materials ('Caoteng', 'Manicure Finger', 'Youxuanpiaolai', 'Huangjin Finger', 'Jinyu', 'Ruby Seedless', 'Mitsushiru', 'Flame Seedless', 'Crimson Seedless', 'Autumn Royal', 'Super Seedless', 'Venus Ⅱ' , 'Centenial Seedless', 'Victoria', 'Benitaka', 'Jinxiu', 'Nongke Ⅱ', 'Venus Ⅰ ') belonged to group Ⅱ; Nongke I was in group Ⅲ; 2 materials ('Superior Seedless' and 'Melissa Seedless') belonged to group Ⅳ; Zaohongti belonged to group Ⅴ.