The DNA sequence of the cytochrome b (cob) gene region of Trimorphomyces papilionaceus mitochondrial DNA has been determined. The cob gene is interrupted by an intron of 1267 bp, which has an open reading frame of 897 bp contiguous to to the upstream exon. The intron belongs to group IB and contains a reading frame with a GIY-10-YVG motif. The deduced amino acid sequence shows 52~60% homology with those of other reported fungi. The phylogenetic relationship of T. papilionaceus with Neurospora crassa, Podospora adserina, Aspergillus nidulans, Saccharomyces cerevisiae, Schizosaccharomyces pombe, and Homo sapiens were inferred through sequence alignment and homology comparison of amino acids deduced from cob genes. When Homo sapiens was set as an outgroup, two ascomycetous yeasts S. cerevisiae and S. pombe made one group and three ascomycetous fungi N. crassa, P. anserine, and a basidiomycetous fungus T. papilionaceus the other group, suggesting that the former group evolved first and then the latter group separated into ascomycetous and basidiomycetous fungi right after that.