Rat MHC (RT.1) class I genes were analyzed by performing Southern blot analysis of digests of genomic DNA isolated from three inbred rat strains using a human derived HLA cDNA probe specific for class I genes. A large number of hybridizing restriction fragments indicated that the rat MHC is a large multigene family. Polymorphic class I DNA restriction fragments specific for the different haplotypes were found for three restriction enzymes used, indicating a high degree of restriction fragment length polymorphism of RT.1 class I genes. Analysis of the methylation state of class I RT.1 genes showed that they are hypermethylated to an extent that is haplotype specific. Furthermore, variation in number and size of Hpa II fragments were observed when comparing liver and spleen DNA from two rat strains sharing the same haplotype. This indicates methylation variation between these organs for RT.1 class I genes.