The catechol 2,3-dioxygenase is an extradiol-type dioxygenase which cleaves the C-C bond at the meta position of catechol to form 2-hydroxymuconic semialdehyde. A catechol 2,3-dioxygenase gene (tomB) in the TOM plasmid of P. cepacia G4 has been cloned and its nucleotide sequence was analyzed. The enzyme gene consisted of 945 base pairs with an ATG initiation codon and a TGA termination codon which can encode a polypeptide of molecular weight 35 kDa containing 314 amino acid residues, and a putative ribosome-binding sequence was identified at approximately 10 nucleotides upstream of the initiation codon. The deduced amino acid sequence of catechol 2,3-dioxygenase from P. cepacia G4 exhibited 79-82% homologies with those of 3-methylcatechol 2,3-dioxygenase of P. putida UCC2 and catechol 2,3-dioxygenase of P. pickettii PKO1.