By screening Caenorhabditis elegans cDNA libraries, overlapping cDNA clones encoding DNA topoisomerase I were obtained. An open reading frame of 751 amino acids was found in 3.2-kb cDNA sequence. The open reading frame has 54% and 50% identities to the amino acid sequences of human and Drosophila melanogaster DNA topoisomerases I, respectively. Northern blot analysis showed the presence of an mRNA of 3.4 kb which suggests that the cDNA sequences is close to full length. The 72-kDa C-terminal polypeptide expressed in Escherichia coli cells showed catalytic DNA topoisomerase I activity. The DNA topoisomerase I gene was mapped to position 18 of chromosome I by screening polytene YAC plasmid DNAs.