ABSTRACT Homeobox cluster genes (Hox genes) are highly conserved and can be usefully employed to study phyletic relation ships and the process of evolution itself. A phylogenetic survey of Hox genes shows an increase in gene number in some more recently evolved forms, particularly in verte brates. The gene increase has occurred through a two-step process involving first, gene expansion to form a cluster, and second, cluster duplication to form multiple clusters. We also describe data that suggests that non-Hox genes may be preferrentially associated with the Hox clusters and raise the possibility that this association may have an adaptive biological function. Hox gene loss may also play a role in evolution. Hox gene loss is well substantiated in the vertebrates, and we identify additional possible instances of gene loss in the echinoderms and urochordates based on PCR surveys. We point out the possible adaptive role of gene loss in evolution, and urge the extension of gene mapping studies to relevant species as a means of its sub stantiation.