DNA fragments Vibrio alginolyticus chemovar iophagus, at least 7 kb in length, were ligated to Escherichia coli expression vectors. Three clones of Escherichia coli HB101 (pLCO-1, pLCO-2, pLCO-3) were obtained by the colony immunoblotting method using anti-collagenase antibody. In Escherichia coli, all these genes produced collagenase antigens which were detected with Western blotting. The amino acid sequence of chemically purified collagenase fragments was also analyzed. An approximately 2.5 kb DNA fragment of the pLCO-1 clone was sequenced, and we found that portions of the deduced amino acid sequence of the chemically analyzed fragments. Therefore, it is highly probable that the gene studied in the present experiment is truly a collagenase structural gene.