Extensive research has been conducted on the characterization of hundreds of date palm (Phoenix dactylifera L.) cultivars worldwide. However, the population genetics of date palms has never been studied. In this study, we collected 200 individuals from 19 populations of different geographical locations in Sudan. The collection sites were grouped according to the type of dates (date palm fruits) that dominates in the area. Ten microsatellite markers were used to investigate the genetic diversity within and among populations and the correlation between the genetic and geographical distances. The tested microsatellite markers showed a high level of polymorphism. A total of 261 alleles were detected at the ten loci. The overall mean value of fixation indices equalled − 0.163, which shows the presence of excess heterozygosity. However, the χ2 tests conducted for every locus in each population indicated no significant deviation from the Hardy–Weinberg equilibrium. The analyses of molecular variance exhibited that about 95% of the total genetic variation existed within populations, while significant differentiation within the type groups could be detected. Although significant isolation by distance (r2 = 0.552, P < 0.05) was detected by a Mantel test, it seems that the spatial effect has become complicated as a result from the exchange and introduction of different kinds of plant materials by date palm growers and traders, as well as seed dispersal. This complexity was clearly apparent in the weak clustering relationships among most of the tested populations.