Siberian apricot (Prunus sibiricaL.) is a woody tree species of ecological, economic, and social importance. To evaluate the genetic diversity, differentiation, and structure ofP. sibirica, we analyzed 176 individuals from 10 natural populations using 14 microsatellite markers. These markers generated 194 alleles in total. The mean number of alleles (13.8571) was higher than the mean number of effective alleles (6.4822). The average expected heterozygosity (0.8292) was higher than the average observed heterozygosity (0.3178). Shannon information index and polymorphism information content were separately 2.0610 and 0.8093, demonstrating the rich genetic diversity ofP. sibirica. Analysis of molecular variance revealed that 85% of the genetic variation occurred within populations, with only 15% among them. The genetic differentiation coefficient and gene flow were separately 0.151 and 1.401, indicating a high degree of genetic differentiation. Clustering results showed that a genetic distance coefficient of 0.6 divided the 10 natural populations into two subgroups (subgroups A and B). STRUCTURE and principal coordinate analysis divided the 176 individuals into two subgroups (clusters 1 and 2). Mantel tests revealed that genetic distance was correlated with geographical distance and elevation differences. These findings can contribute to the effective conservation and management ofP. sibiricaresources.