Sixteen actinomycetes capable of utilizing dibenzofuran as a sole source of carbon and energy were isolated, including Rhodococcus, Microbacterium, and Terrabacter genera. Heretofore, no dibenzofuran-utilizing strain belonging to the genus Microbacterium has been reported. Five extradiol dioxygenase genes (dfdB, and edil to 4) of the strain Rhodococcus sp. YK2 were cloned and analyzed. The nucleotide sequence of dfdB gene was almost identical to the bphC1 gene of Terrabacter sp. DPO360, which was involved in dibenzofuran metabolism in this strain. Southern and Northern hybridization analyses using these extradiol dioxygenase genes as probes suggest that the dfdB gene in YK2 was conserved in diverse dibenzofuran-utilizing actinomycetes; also, the dfdB gene was the only expressed gene among five extradiol dioxygenase genes in the medium containing DF as a sole carbon source. These results suggest that the dfdB gene is important for dibenzofuran metabolism not only in the strain YK2, but also in diverse dibenzofuran-degrading actinomycetes.