Identification And Genetic Relationships Analysis Of Blueberry Cultivars Using SRAP Markers

The modified method of extracting high quality genomic DNA from mature tissues of blueberry(Vaccinium spp.) and the optimum system of Sequence-related Amplified Polymorphic (SRAP) for blueberry were established in this thesis. Genetic relationships among 21 varieties of blueberries were evaluated through a SRAP analysis. The results will serve as reference to future identification, reservation, utilization and variety selection.Results showed that higher quantity and quality DNA was obtained from the modified CTAB method than that from methods of rapid CTAB and conventional CTAB. The genomic DNA was assessed by ultraviolet photometer with value of OD260nm/OD280nm from 1.75 to 1.85, the average DNA production was 192μg/g, the quality of genomic DNA extracted by this method can extensively meet the requirements for general molecular and biological experiments, such as SRAP, RFLP, AFLP and so on. The optimized program was:94℃for 5 min; 94℃for 1 min,47℃for 1 min,72℃for 1 min,35 cycles; 72℃for 10 min. The stable reaction system was:20μL reaction system containing 1.5-2.0 ng/μL DNA template, 0.3mmol/L dNTP,0.5μmol/L primer,1 U Taq DNA polymerase and 1.5 mmol/L Mg2+.The molecular marker system-SRAP was used to assess genetic relationship and identification in 21 different blueberry cultivars.15 primer pairs selected from 100 amplified 172 (the polymorphic rate is 83.1%) polymorphic bands with a range from 5 to 18 polymorphic bands per primer pair. Moreover, Nei's genetic similarity ranged from 0.5604 to 0.8841. Cultivar Bluecrop had the greatest genetic similarity with Blueray, and cultivar Blomidon had the greatest genetic distance with Jersey. Using arithmetic averages (UPGMA) dendrogram, the unweighted pair-group method indicated that 21 samples were classified into two major groups and four subgroups, clustered results had high consistency with the pedigree of cultivars.In addition, SRAP markers could be used for identification of blueberry varieties, and the distinguished ability per primer pair was from 33.3% to 100%.15 specific SRAP markers of 7 different cultivars were obtained respectively. One dominant SCAR marker BLG-1 was developed from SRAP marker from cultivar Legacy, and showed the same profiles as the corresponding SRAP marker.