Genes for an isoamylase-like debranching enzyme have been isolated from rice and Aegilops tauschii, the donor of the D genome to wheat. The structures of the genes are very similar to each other and to the maize SU1 isoamylase gene and consist of 18 exons spread over approximately 7.5 kb. Southern analysis and fluorescent in situ hybridization showed the Ae. tauschii gene to be located in the proximal region of the short arm of chromosome 7D, thus showing synteny with the localization of the rice isoamylase gene on rice chromosome 8. Analysis of the expression pattern of wheat sugary isoamylase genes indicates that they are strongly expressed in the developing endosperm 6 days after flowering. Three distinct Sugary-type cDNA sequences were isolated from the wheat endosperm that are likely to correspond to the products of the three genomes. The deduced amino acid sequence of rice and wheat Sugary-type isoamylase is compared with other sequences available in the database and the results demonstrate that there are three types of isoamylase sequences in plants: those containing 18 exons (the Sugary-type isoamylase gene), those containing 21 exons, and those containing only 1 exon. It is possible that different combinations of isoamylase genes are expressed in different tissues.