Amelanchier sinica has important ornamental, nutritional, and ecological value. Genetic diversity studies could lay a foundation for the protection and utilization of its resources. We analyzed the genetic diversity of 96 individuals from 6 natural populations in Shaanxi Province using amplified fragment length polymorphism (AFLP) markers, so as to clarify the genetic relationship of A. sinica and provide theoretical basis of the protection, breeding and utilization for A. sinica resources. The results showed: (1) eight primers with strong polymorphism were identified from 64 AFLP primers, generating a total of 277 bands, of which 116 (42.86%) showed polymorphism. (2) The results of UPGMA clustering, principal coordinate analysis (PCoA) and structure analysis indicated that the 6 wild populations of A. sinica were divided into two groups. (3) There was obvious genetic differentiation between north and south in the Qinling Mountains, and there was some gene flow among the populations. The results of molecular variance analysis (AMOVA) indicated that 37% of genetic variation is derived from among populations and that 63% is derived from within populations. The results of the Mantel test (r = 0.192, P = 0.220) of the wild populations of A. sinica in Shaanxi Province indicated that the correlation between geographical distance and genetic distance was not significant. The results showed that AFLP molecular markers could be used to analyze the genetic diversity of A. sinica accurately and effectively; more genetic variation in A. sinica existed within populations, while gene exchanged among populations was limited. The overall genetic diversity of A. sinica in Shaanxi Province was relatively low, while some populations had higher genetic diversity. These results provide a theoretical basis for the protection of wild germplasm resources, breeding of elite varieties and development and utilization of the resources of wild populations of A. sinica.