A plasmid, pBP614 (5.647 kb), has been isolated from the New Zealand Bacillus popilliae strain 17 and characterized by physical mapping, cloning, and sequencing. An open reading frame was found which could encode a protein with homology to the replication (Rep) proteins of rolling-circle plasmids. The predicted B. popilliae Rep protein shows closest homology to Rep proteins from Bacillus plasmids of the pC194 family. Sequences homologous to plus- and minus-strand replication origins were found. The minus-strand origin shows similarities to the pal-T type family. Taken together, these observations suggest that pBP614 is a rolling-circle plasmid, the first such plasmid characterized from B. popilliae.