Multiple AMP deaminase (AMP-D) isoforms have been found in vertebrates, and tissue-specific inherited deficiencies of AMP-D have been described in two unrelated clinical syndromes suggesting there may be more than one AMP-D gene in higher eukaryotes. Using a newly isolated cDNA cloned from an adult rat brain library and a previously reported cDNA cloned from adult rat skeletal muscle, two linked AMP-D genes have been identified in rat and man. ampd1 is expressed at high levels in skeletal muscle of the adult rat. ampd2 is the predominant gene expressed in non-muscle tissues and smooth muscle of the adult rat, and it is also the predominant gene expressed in embryonic muscle and undifferentiated myoblasts. Both genes are expressed in cardiac muscle of the adult rat. The peptides encoded by these two genes have distinct immunological properties. The conservation of nucleotide sequence and exon/intron boundaries in these two genes suggests they arose by duplication of a common primordial gene around 150 million years ago.