The Allegheny woodrat (Neotoma magister) is a colony-forming murid rodent inhabiting rock outcrops, cliff and talus slopes, and caves within the Central and Southern Appalachians of North America (Hall 1981). The species is currently considered threatened, endangered, or at risk throughout its range purportedly due to the direct or indirect effects of human-fragmented landscapes. Currently, Allegheny woodrat management units are arbitrarily defined as metapopulations that roughly constitute contiguous rock outcrops interconnected by forested habitat. To strengthen long-term population stability and reduce the need for further protection through the regulatory process, an effective conservation programme for this at-risk species will require unambiguous, objectively defined units of management that discern evolutionarily important lineages (Avise 1994). A prerequisite in the development of management strategies that recognize and maintain significant woodrat lineages is a thorough understanding of the levels of gene exchange among geographically proximate and distal populations. However, no information exists on population structure, levels of gene flow, or relatedness among Allegheny woodrat populations or colonies. As an initial step towards addressing this research need, we have developed a suite of polymorphic microsatellite DNA markers for this species. In this paper, we describe the isolation and characterization of these markers and demonstrate their suitability in amplifying putatively homologous products in seven additional Neotoma species.