Genetic variability and population structure in the endangered peregrine falcon (Falco peregrinus) were studied using DNA microsatellite markers. Special emphasis was placed on the subspecies F. p. peregrinus living in Scandinavia and Scotland. For comparative purposes, three North American peregrine subspecies (F. p. tundrius, F. p. pealei and F. p. anatum) and one Tasmanian subspecies (F. p. macropus) were included in this analysis. Twelve DNA microsatellite loci were analysed across a total of 146 individuals. The amount of genetic variation did not differ in peregrine populations for which adequate sample sizes were obtained. Within F. p. peregrinus, significant genetic differentiation was found between populations in northern and southern Scandinavia and between the Scandinavian and Scottish populations, while the populations in south-eastern Norway and south-western Sweden did not differ significantly. Analysis of cliff-nesting peregrines in northern Sweden and bog-nesting peregrines in northern Finland/Sweden suggests that the difference in nesting habitat is not associated with genetic differentiation. Population structuring in F. p. peregrinus is further supported by an assignment test, wherein simulated genotypes are correctly assigned to the northern and southern Scandinavian and Scottish populations with relatively high probabilities, and by analysis of allele-sharing among individuals. Cluster analysis of genetic distances grouped populations of peregrines in accordance with their subspecific designation, and peregrines on either side of the Atlantic Ocean were clearly separated.