The full-length cDNAs of woodchuck major histocompatibility complex (MHC) class I (MhcMamo-I or Mamo-I) genes were cloned by using cellular mRNA isolated from the peripheral blood mononuclear cells and liver tissues of woodchucks. DNA sequence analysis of Mamo-I cDNAs revealed that the coding regions of Mamo-I genes were about 1,080 bp long, encoding 359 amino acid residues. The deduced amino acid sequences of Mamo-I showed structural features like leader, alpha1, alpha2, alpha3, transmembrane and cytoplasmic domains, similar to their homologues in human and other mammals. Analysis of five full-length clones from unrelated woodchucks indicated a polymorphism within the alpha1 and alpha2 domains of Mamo-I heavy chain and a high conservation within the alpha3 and the transmembrane/cytoplasmic domains. Amino acid residues of the alpha2 and alpha3 domains that are supposed to be involved in the binding of MHC class I to CD8 molecule, were largely conserved in Mamo-I genes. Phylogenetic comparison of MHC class I genes of woodchuck and other mammals indicated a close evolutionary relationship between woodchuck and squirrel MHC class I. We tentatively named this region the locus A of Mamo-I genes (Mamo-A). Sequence analysis of 101 clones of alpha1 and alpha2 regions derived from 14 woodchucks revealed that at least 14 different alleles within Mamo-A exist. Among these 14 alleles identified so far, Mamo-A*01 and Mamo-A*09 were of the highest frequency of about 21.5% and 14.5%, respectively. Our results indicate that Mamo-I genes are of a similar molecular structure to those of human and other mammals.