Hypothetical evolutionary pathways leading to tetrapod ADH multiplicity. At the base of vertebrate radiation, an initial tandem duplication of the ancestral ADH3 led to a two-gene cluster. Actinopterygia (ray-finned fish) and sarcopterygia (lobe-finned fish and tetrapods) acquired ADH1 activity by the most 5′ member of the cluster . Before the amniota/amphibian split (360 Mya), ADH2 and ADH7 would have arisen in tetrapods as a consequence of gene duplication events. In reptiles and birds, no additional ADH classes have been found. In contrast, ADH1 tandem duplications led to further class multiplicity in the amphibian lineage; thus, ADH8, ADH9, and more recently ADH10 forms would derive from the ancestral ADH1. Close to the origin of mammals, ADH7 was lost while gene duplications generated ADH5 and ADH6, and tandem duplication of ADH1 gave rise to ADH4. Only in primates, ADH6 was lost simultaneously or close to ADH1 duplications generating ADH1A-C isozymes . Likewise, additional duplications occurred in other vertebrate lineages, and those ADH genes leading to isoenzyme multiplicity in at least one member of that lineage are underlined (in reptiles, multiple ADH1 and ADH7 are found in lizards, but not in turtles). In some organisms, ADH pseudogenes are also observed.