Sequences of nuclear ITS (nrDNA) regions were obtained from 22 species of Tyromyces s.l., a group of important wood-rotting, poroid holobasidiomycetes, belonging to Coriolaceae. Different reference collections of a species generally produced identical molecular sequences. Parsimony analysis of these sequences resulted in trees compatible with the currently accepted delimitation of genera, but the alignment of sequences has proven to be difficult owing to regions of uncertainty. Analyses based on inclusion and exclusion of ambiguous regions in the alignment demonstrate that closely related taxes are, however, always clustered together, especially at the generic level. It is considered that ITS sequences are reliable in distinguishing genera and species of Tyromyces s.l., but not able to provide sufficient resolution for relationships between some formerly recognized genera. The taxonomic implications of ITS variation are also discussed. Species of Oligoporus and Postia clustered in one clade, indicating a close relationship as shown from morphological research. Species of Skeletocutis are characterized by encrusted hyphal ends in the hymenophore, but the type species of Tyromyces, T. chioneus, is embedded within the Skeletocutis clade, indicating a strong link between these two genera. Species of Antrodia and of Spongipellis included in this study were not well supported as monophyletic groups, although they are clustered together within the trees.