DNA fragments homologous to chitin synthase were amplified from the genomic DNA of Penicillium chrysognum by PCR. Cloning and sequencing of the PCR amplified fragments led to the identification of four different genes, designated PcCHS1, PcCHS2, PcCHS3, and PcCHS4. By comparison of the deduced amino acid sequences, PcCHS1 was identified as a gene for class I chitin synthase, PcCHS2 and PcCHS3 were for class II, and PcCHS4 was for class III. For the phylogenetic study of P. chrysogenum, the Blast P similarity search and the bootstrap analysis of the deduced amino acid sequences of the highly conserved region of chitin synthase genes were performed. The analysis of amino acid sequences implicated a close evolutionary relationship of Penicillium to ascomycetous fungi, especially to genus Aspergillus. The results from bootstrap analysis of the chitin synthase also supported the usefulness of the chitin synthase for phylogenetic study of filamentous fungi.