Identification Of Grape Cultivar And Genetic Stability Of Grape Virus Eradication Based On SSR Markers

In order to provide reference basis for the protection and utilization healthy seedlings of Ningxia wine grape variety identification and plant breeder’s rights,SSR reaction system was established,variety identification of wine grape and genetic stability of virus-free plantlets were analyzed by SSR molecular markers.10 pairs of SSR primers were used to analyze the genetic relationship of 14 wine grape resources of Ningxia,different primers and primer combinations were selected for variety identification of wine grape.110 pairs of SSR primers were used to analyze genetic stability of 4 kind virus-free plantlets.The main results from this study are as follows:1.Suitable reaction system and Program of the SSR analysis for grape genomic DNA was the lOul-reaction system with 15-30ng template DNA,3.0mmol·L-1Mg2+,0.10 mmol·L-1 dNTP,1.0U TaqDNA Polymerase,0.2 μmol·L/-1 Primer,The program was that pre-denature in 94℃for 5min, denature in 94℃ for 30s, renature in 54-64℃ for 15s, elongate in 7 2℃for 1 min,cycling number in 35,holding in 72℃for 10 min.2. The system was used for the amplification reaction in the experiment in all the material test,10 primers showed high efficient amplification were selected from 110 random primers and used for all materials in amplification tested.The number of detceted allele per primer is 7.4,the most are SP0102,for 11.The 14 materials were clustered into three groups by using genetic similarity coefficient.The fist group only include Traminette,the second include two materials,the others all belong to the third.3.The results of SSR technique was processed by using the NTSYSpc Version 2.10e software. According to the identification efficiency of primers,the highest identification efficiency primer SP0097 could differentiate the 14 materials separately.23primer combinations selceted from the other 9 primers also could differentiate the 14 materials.4.110 pairs of SSR primers were used to analyze genetic stability of 4 kind virus-free plantlets by Cryotherapy.The results showed that the genetic stability of regenerated plantlets were good.