Nucleotide sequence analysis of the cytochrome-b gene of mtDNA and univariate and multivariate statistical analyses of cranial morphology were used to determine the evolutionary status of relictual, isolated populations of pocket gophers (genus Geomys) from the Ozark Mountains of Arkansas. Results of the mtDNA sequence analysis indicated that populations from the Ozark Mountains are most similar to G. bursarius missouriensis in Missouri and not to geographically proximate populations of G. breviceps. Morphological analyses were concordant with those results. Our findings confirm earlier work comparing similarities in allozymes and species-specific ectoparasites. Nucleotide sequence and morphological divergence between the Ozark populations and G. bursarius missouriensis are typical of differentiation between other subspecies of G. bursarius. We conclude that populations of pocket gophers isolated in the Ozark Mountains represent a discrete genetic entity and should be recognized as a new subspecies.