This data set contains the raw genotypes that were assigned to 122 and 127 Duma florulenta and Acacia stenophylla individuals, respectively. Genotypes were identified using GeneMapper 4.0 from following fragment analysis of DNA amplified using microsatellite primers. The data set contains genotypes at 12 and 13 microsatellite loci for Duma florulenta and Acacia stenophylla, respectively. Samples were collected from sites that were located on 7 rivers in the northern Murray-Darling Basin (Balonne River, Bokhara River, Birrie River, Culgoa River, Warrego River, Paroo River and Darling River). Geographic co-ordinates for each individual are contained in the last two columns of the data for each species.