Amplified fragment length polymorphism (AFLP) DNA markers were used to characterize the genetic diversity and relationships in gum Arabic tree (Acacia senegal). Twenty eight samples of Acacia senegal collected from populations distributed throughout the Gum Arabic belt were tested in comparison with samples of Acacia mellifera and Acacia leata. Nine AFLP selective primer pair combinations generated a total of 433 amplification products with an average of 89.18% detected polymorphisms. Accessions showed the least variation was found within the A. senegal accessions in contrast with mellifera and Acacia leata that presented the highest degree of polymorphism number. According to the cluster analysis two main clusters were obtained in which A. mellifera and A. leata were placed in a separate group. There were eight subgroups of A. Senegal. Three of the eight subgroups of A. senegal were clustered according to geographical origin. The variation within population might be correlated with abiotic factors of the environment.