GPD genes encoding glyceraldehyde-3-phosphate dehydrogenase were isolated from the homobasidiomycetes Schizophyllum commune, Phanerochaete chrysosporium and Agaricus bisporus. All three species contain one transcriptionally active GPD gene, but A. bisporus also contains an inactive GPD gene (tandemly linked to the active gene). These genes contain 5–9 introns located at conserved positions, differing (except in one case) from intron positions in ascomycetous GPD genes. The predicted amino-acid sequences of the proteins encoded by the three active GPD genes are highly homologous. A comparison with protein sequences from filamentous ascomycetes shows a clear distinction, whereas the GPD genes from ascomycetous yeasts are quite distinct from both the filamentous ascomycetes and basidiomycetes. Promoter regions of ascomycetous GPD genes do not correspond to those of the GPD genes of basidiomycetes which may (partly) explain poor expression in basidiomycetes of introduced genes driven by an ascomycete GPD promoter.