The Hox genes have been found to encode transcription factors, which specify the morphological identity of structures along the anteroposterior axis of animals ranging from worms to mice. The canonical set of nine genes is organized in a cluster in the genome of several protostomes and deuterostomes. However, within insects, whereas the Hox genes are organized in a single cluster in the beetle Tribolium castaneum, they are split into two separate groups in the flies Drosophila melanogaster and Drosophila virilis. The significance of a split Hox cluster is unknown and has been observed in only one organism outside the Drosophila lineage: the nematode Caenorhabditis elegans. We have cloned a majority of the Hox genes from the mosquito Anopheles gambiae (Diptera: Culicidae) and compared their genomic organization with that of Tribolium and Drosophila to determine if a split Hox cluster is found in dipterans aside from the Drosophilidae. We find that the Hox genes in Anopheles, as in Tribolium, are organized in a single cluster that spans a genomic region of at least 700 kb. This finding suggests that, within the insect genome, the partition of the Hox cluster may have evolved exclusively within the Drosophila lineage. The genomic structures of the resident genes, however, appear to be largely conserved between A. gambiae and D. melanogaster.