We isolated 11 new polymorphic microsatellite markers from Acacia atkinsiana, a plant species endemic to the Pilbara region in Western Australia. Next generation (454) sequencing was used to identify 28 microsatellite markers that were trialled in individuals across the species range. Characterisation of 11 of these loci in 24 individuals from a single population yielded between two and nine alleles per locus, and observed heterozygosity ranged from 0.042 to 0.958. Four loci that amplified in A. atkinsiana were also successfully amplified and polymorphic in Acacia ancistrocarpa. These loci will be valuable for assessing population genetic structure in A. atkinsiana and identifying seed sources for rehabilitation of disturbed sites.