The class I alcohol dehydrogenases (ADH; EC 1.1.1.1) play a key role in hepatic alcohol catabolism. Three human class I ADH genes, ADH1, ADH2, and ADH3, which encode the alpha, beta, and gamma subunits respectively, have been isolated and mapped on chromosome 4q21-q23. Genomic cloning using a cosmid vector allowed us to obtain an 88-kb-long genomic segment, which was found to include an entire 80 kb of the class I ADH gene cluster. All three genes lie in the same transcriptional orientation and the order of genes is 5'-ADH3-ADH2-ADH1-3'. It may be of some significance that the order of transcriptional activation in the hepatic development, alpha----beta----gamma, is opposite to the order of gene arrangement. Several members of the AluI family and the KpnI (L1) family of interspersed repetitive sequences were mapped in this region. The divergence of insertional sites suggested that gene multiplication of the class I ADH genes had taken place in the earlier stages of human (or primate) evolution.