Genetic Diversity And Clustering Analysis Of Vaccinium Spp. Resouces By CDDP Markers

The fruit of blueberry (Vaccinium spp.) has high nutritional value and heathy benefits. The wild resources of Vaccinium species is abundant in our country, and many cultivaed breeds was introduced. But the genetic background of the introduced blueberry is not clear for a long time in our country. The genetic diversity and relationship is not clear between the wild breed and the introduced species. There are many adverse effects on collecting, protecting,and using of blueberry germplasm and breeding. Analysis of genetic diversity and phylogenetic relationship in blueberry germplasm by molecular markers have great significance for resource protection, utilization and germplasm innovation. In this study, a new molecular marker technique:CDDP (Conserved DNA-derived polymorphism) was used in genetic diversity and phyloggeetic relationship analysis between the wild Vaccinium uliginosum and 31 cultivars, screening and exploiting new CDDP primers used for identification of blueberry cultivars. The main results were as following:1. After comparing the results of single factor experiment, the optimized CDDP-PCR reaction system (20 ?L) containing 1.25 ng/?L genomic DNA,0.5 pmol/?L primer,1.5 nmol/?L Mg2+,0.375 nmol/?L dNTPs,1.5 U Taq.2. Seventeen out of 21 CDDP primers with good polymorphism and clear bands were selected in this study. Using selected primers,188 (90.82%) polymorphic and 17 (8.21%) specific bands were detected out of 207 bands among 32 blueberry accesses. At the population level, the number of effective alleles, the diversity index of Nei's gene and Shannon information index were 1.59,0.37 and 0.56, respectively. The SM similarity coefficients among different genotypes varied from 0.44 to 1.00 with an average of 0.73.3.Thirty-two blueberry were divided into three categories when the similarity coefficient was 0.69, the wild species 'Vaccinium uliginosum' separate clustered together, the lowbush species 'Blomidon' and half-highbush variety 'Northblue' were clustered into one group, the other varieties were clustered into one category. The CDDP markers in blueberries had high polymorphism and specificity, and it was also related to the chilling requirement, cold resistance and fruit color of the germplasm.4. In this study, new CDDP primers was develop based on the conserved sequence of cold resistance gene CBF and anthocyanin biosynthease gene. A total of 165 bands were amplified by 14 primers of CBF gene, which including 163 (98.79%) polymorphic bands and 25 (15.15%) specific bands. A total of 226 bands were amplified by 22 primers of anthocyanin biosynthease gene, which including 206 (91.15%) polymorphic bands and 16 (7.08%) specific bands. The polymorphism and specificity of new CDDP primers were higher than those of published primers, thus they could be used in the identification and genetic diversity analysis of blueberry genotypes with a greater advantage.