In spite of the use of chromosome number, floral anatomy, pollen morphology, and serology, substantial disagreement remains regarding the systematics of the Berberidaceae. To test competing hypotheses about intergeneric relationships in the family, sequences of the chloroplast encoded rbcL gene and ITS regions of the nuclear ribosomal repeat were examined. Analysis of rbcL sequences does not support fragmentation of the Berberidaceae into smaller families, especially the separation of Nandina as a distinct family. The large woody genera, Berberis and Mahonia, are related to the monotypic herbaceous genus Ranzania. Bongardia is nested within a clade characterized by a basic chromosome number of x = 6, and has a remote relationship to the Leontice group with which the genus has been placed in most previous classifications. The x = 6 group, the largest in the Berberidaceae, comprises two distinct lineages: Jeffersonia and the rest of the core genera (Diphylleia, Bongardia, Achlys, and Epimedium). Among the core genera, Diphylleia is sister to Epimedium/Achlys and their sister genus Bongardia. ITS sequence data provided additional support for the phylogenetic relationships of the x = 6 group observed in the rbcL gene tree.