One partial and two complete genomic clones of the three loci specifying alcohol dehydrogenase (ADH) in barley were isolated by screening libraries with a maize Adh1 cDNA probe. Each gene is characterised by an intron arrangement similar to that of both maize Adh1 and Adh2, although two genes show an exon fusion. A comparison with the maize coding sequences unambiguously assorts the barley loci into an Adh1-like gene and two Adh2-like genes, indicating that an ancient gene duplication underlies the widespread occurrence of two Adh loci in higher plants. In the barley lineage there has been a further duplication-transposition of a progenitor Adh2 locus to give rise to the extant three-gene system, with gene copies of different ancestry being closely linked. An Adh1 null-allele, Adh1-M9, has been cloned; the available sequence includes an intron with a missing acceptor splice signal. Two independent clones of one of the barley Adh2-like genes have an 18 bp in-frame deletion towards the 3 end of the coding sequence. The barley Adh2-like genes are extensively diverged in their 5 sequences apart from a conserved 15 bp motif in the mRNA leader region and sequences at the start of transcription. A sequence related to the hexanucleotide core of a regulatory element found in maize Adh1 and in other anaerobically induced plant genes is present in the 5 region of barley Adh2.