Mouse hepatoma × rat hepatocyte hybrids that segregate rat chromosomes were used to determine the chromosomal location of the rat genes encoding 11 β-hydroxylase and aldosterone synthase (Cyp11b1 and Cyp11b2 respectively). By means of species-specific restriction fragments and microsatellite markers both genes were mapped to rat chromosome 7. The Cyp11b1 microsatellite marker was subsequently found to vary in length between and within rat strains. Furthermore, we compared the sequences of Cyp11b1 markers in two genetically hypertensive strains of rat with their normotensive counterparts. Previous studies have indicated that 11β-hydroxylase activities in Milan and Lyon hypertensive strains are different from their respective genetic controls. The Cyp11b1 microsatellite regions from Lyon hypotensive and normotensive strains of rat were similar and were both shorter by 15 bases than that of the Lyon hypertensive strain. The Cyp11b1 marker in Milan hypertensive (MHS) and normotensive (MNS) strains differ from all the Lyon strains and from each other. The MHS marker is 12 bases shorter than that of MNS rats. These differences in microsatellite length may provide useful polymorphic markers in co-segregation studies of genetic hypertension in rats.