Through the determination of phenotypic traits of 43 wolfberry materials，combined with SSR molecular marker technology，the population structure and genetic diversity between ancient wolfberry trees and existing wolfberry varieties in Ningxia were comprehensively analyzed. The principal component analysis results of phenotypic traits determined that the principal component was 4，and the cumulative contribution rate reached 78.09%. The leaf size of the first principal component and the fruit size of the second principal component played a major role. The coefficient of variation ranged from 15% to 54%，with an average of 26.78%，and the genetic variation index ranged from 5.26 to 5.41. The phenotypic traits were divided into 4 groups by cluster analysis，3 individuals in group I，3 individuals in group II，8 individuals in group III and 29 individuals in group IV. Five materials with excellent comprehensive properties were determined by membership function method，namely L14，L16，L28，L40 and L43. The average effective alleles of SSR molecular markers were 2.018，the average expected heterozygosity was 0.398，and the average Shannon ′ s index was 0.839. The optimal K value of SSR molecular markers was 4. Among the five materials with better comprehensive evaluation，L16 showed heterozygous genotype，and L14，L28，L40 and L43 showed single genotype. In this study，a variety of methods were used to comprehensively analyze the phenotypic traits and genetic diversity of ancient wolfberry trees from different sources and ages，which provided a theoretical basis and practical guidance for the subsequent utilization of ancient wolfberry germplasm resources and the breeding of wolfberry varieties.