Penstemon subgenus Dasanthera is a group of nine species distributed at high elevations in western North America. Hybridization is common in the subgenus when two or more species occur in sympatry. We examined evolutionary trends and the importance of hybridization to the evolution in the subgenus. Phylogenetic and biogeographic patterns among members of Penstemon subgenus Dasanthera were assessed using nuclear ribosomal ITS and chloroplast matK sequence data, and inter-simple sequence repeat (ISSR) markers. These data support previous hypotheses suggesting that the Cascade-Sierra Nevada lineage is derived from the northern Rocky Mountain lineage. Furthermore, there is a shift in growth form from suffructescent perennials to evergreen subshrubs concurrent with migration to the Cascade-Sierra Nevada mountains. Within the Cascade-Sierra lineage, P. newberryi and P. rupicola exhibit floral characters that may represent adaptations for hummingbird pollination. The ISSR data demonstrate the utility of dominant markers for reconstructing phylogenetic relationships at the interspecific level.