The first putative plant acyl-CoA oxidase cDNA has been isolated from a Phalaenopsis cDNA library constructed by poly(A)+ RNA extracted from petals 1 day after pollination. This cDNA, pOACO31, contains a 2100-bp open reading frame which encodes a polypeptide named PACO1 of 699 amino acids. The predicted isoelectric point of PACO1 is 8.74 and the molecular weight is 78,032 Da, similar to that of a monomer of predicted plant acyl-CoA oxidase. Southern blot analysis indicated that this gene occurs in one copy or a low number of copies per haploid genome. When compared with sequences in databases, PACO1 revealed significant similarity only to peroxisomal acyl-CoA oxidase particularly within 13 conserved regions and a putative FMN-binding site.