Development Of Microsatellite Markers And Analysis Of Genetic Diversity Of Populations From Yan Mountains In Prunus Sibirica L.

Prunus sibirica L., an important ecological and economic tree species endemic to Asia, is widely distributed in the mountain area of North and Northeast of China. There exist abundant genetic variation for P. sibirica in the nature; however, few is known about the genetic diversity and population structure. Therefore, we developed SSR markers of P. sibirica and studied genetic diversity of P. sibirica populations in Yan Mountains, this study would provide important basis to the conservation and further utilization of P. sibirica germplasm resources. The main research work and the results are as follows:1. Development of microsatellite markers in P. sibiricaWith biotinylated (AG)15 oligoprobes hybridization to fragments of genomic DNA digested by restriction enzyme Alu I and HaeⅢ, two libraries were constructed. A total of 384 single white clones were selected, among which 166 clones were confirmed to containe microsatellite loci by the method of three primers PCR. After the positive clones sequenced,151 high quality sequences were obtained. We finally identified 136 non-redundency sequences, containing 200 SSR loci. Further analysis showed that the motif of AG is the main motif type. In total,141 primer pairs were designed by Primer3. In order to screen the primers designed, eight P. sibirica individuals randomly selected from eight wild populations in the north of Hebei Province were selected.71 primer pairs amplificated successfully and produced band with expected size, among which 19 primer pairs could produce the products in all 3 P. sibirica populations and 6 P. armeniaca individuals. These microsatellite primer pairs should be useful for population genetics, germplasm identification and marker assisted selection in the breeding of P. sibirica.2. Genetic diversity and genetic structure of P. sibirica populationsIn order to evaluate the genetic diversity and genetic differentiation of populations and present a scientific framework for the conservation and utilization of P. sibirica, the genetic diversity and population structure of 17 populations of Siberian apricot in Yan Mountains area were investigated by using nine microsatellite markers. A total of 203 alleles were identified in 533 individuals, with a mean of 22.556 alleles per locus. Effective number of alleles per locus (Ne) was 5.714, percentage of polymorphism loci (PPL) was 100%, the mean expected heterozygosity (He) was 0.788, suggesting a high genetic diversity in natural populations of P. sibirica. The highest genetic diversity was found in Badaling population, the second was Yushulinzi population, while the lowest was found in Yimatu population. The population genetic differentiation (FST) was 0.065 and gene flow (Nm) was 3.836. AMOVA analysis showed that genetic variation within the populations accounted for 95.62% of the total. Mantel test revealed that genetic distance and geographic distance were positively correlated (r= 0.5894, P< 0.0001). The UPGMA cluster analysis showed that populations with close physical distance clustered into the same clade, which was consistent with results of Mantel test. Based on the present result, some suggestion on further collection of P. sibirica genetic resoures were put forward.