Technical Abstract: The National Clonal Germplasm Repository in Davis, CA is a United States Department of Agriculture (USDA), Agricultural Research Service (ARS) genebank that houses national collections of various Mediterranean fruit and nut crops. The Prunus collection is the second largest in this genebank and is exceeded only by Vitis (grapes). There are more than 90 taxa and in excess of 1600 accessions of Prunus spp. in the collection that includes almonds, apricots, cherries, peaches, and plums. The mission of the genebank is to acquire, maintain, evaluate, and distribute germplasm. Acquisition is from plant collection trips and donations of germplasm to the repository and includes cultivars, breeding lines, and Prunus wild relatives. The trees are propagated and maintained in the Wolfskill Experimental Orchard near Winters, CA and in containers in screen houses in Davis. Evaluation data are compiled on phenotype and genotype and are freely available online at the Germplasm Resources Information Network (GRIN), see: http://www.ars-grin.gov/cgi-bin/npgs/html/site_holding.pl?DAV. As a part of genetic characterization of germplasm at the Davis repository, an analysis of genetic diversity and differentiation using fifteen SSR loci among five diploid Prunus spp. revealed significant variability within and among species with a number of species specific alleles of low to moderate frequencies. The average number alleles/locus across the 15 loci assayed ranged from 6.93 for peach (P. persica) to 18.27 for almond (P. dulcis) with an overall average of 11.38 alleles/locus. Among the cultivated species, the observed and expected heterozygosities were relatively higher except for P. persica with 0.3247 and 0.4351 for observed and expected levels, respectively and with the lowest average number of alleles across loci (6.93) among the cultivated species. Within population genetic variability measures indicated that P. dulcis is the most highly variable species among the cultivated Prunus followed by P. ceracifera, P. armeniaca and P. salicina. As expected, peach and almond showed close genetic affinity followed by diploid plums, and apricots. The results are summarized as applied to genetic conservation, management, and utilization of Prunus germplasm.