The isolation of Hox genes from two cnidarian groups, the Hydrozoa and Anthozoa, has sparked hypotheses on the early evolution of Hox genes and a conserved role for these genes for defining a main body axis in all metazoan animals. We have isolated the first five Hox genes, Scox-1 to Scox-5, from the third cnidarian class, the Scyphozoa. For all but one gene, we report full-length homeobox plus flanking sequences. Four of the five genes show close relationship to previously reported Cnox-1 genes from Hydrozoa and Anthozoa. One gene, Scox-2, is an unambiguous homologue of Cnox-2 genes known from Hydrozoa, Anthozoa, and also Placozoa. Based on sequence similarity and phylogenetic analyses of the homeobox and homeodomain sequences of known Hox genes from cnidarians, we suggest the presence of at least five distinct Hox gene families in this phylum, and conclude that the last common ancestor of the Recent cnidarian classes likely possessed a set of Hox genes representing three different families, the Cnox-1, Cnox-2, and Cnox-5 families. The data presented are consistent with the idea that multiple duplication events of genes have occurred within one family at the expense of conservation of the original set of genes, which represent the three ancestral Hox gene families.