We have determined by Southern blot analysis that DNA sequences homologous to the AMG gene probe are present in the genomes of both marsupial and monotreme mammals, although adult monotremes lack teeth. In situ hybridization and Southern analysis of cell hybrids demonstrate that AMG homologues are located on autosomes. In the Tammar Wallaby, AMG homologues are located on chromosomes 5q and 1q and in the Platypus, on chromosomes 1 and 2. The autosomal location of the AMG homologues provides additional support for the hypothesis that an autosomal region equivalent to the human Xp was translocated to the X chromosome in the Eutheria after the divergence of the marsupials 150 million years ago. The region containing the AMG gene is therefore likely to have been added 80-150 million years ago to a pseudoautosomal region shared by the ancestral eutherian X and Y chromosome; the X and Y alleles must have begun diverging after this date.