Genomic imprinting in mammals is an epigenetic process that results in differential expression of the two parental alleles. The tightly linked murine H19 and Igf2 genes are reciprocally imprinted: H19 is expressed from the maternal chromosome while Igf2 is expressed from the paternal chromosome. A single regulatory region in the 5' flank of the H19 gene has been implicated in silencing both genes. On the paternal chromosome, this region is heavily methylated at CpG residues, leading to repression of the H19 gene. The mechanism by which the same region in an unmethylated state on the maternal chromosome silences Igf2 is less well understood. We have probed the chromatin structure of the region by assessing its sensitivity to nuclease digestion. Two regions of nuclease hypersensitivity that are specific to the maternal chromosome were identified. These coincide with the region that is most heavily methylated on the paternal chromosome. As is the case with paternal methylation, hypersensitivity is present in all tissues surveyed, irrespective of H19 expression. We suggest that the chromatin structure of the maternal 5' flank of the H19 gene may represent an epigenetic mark involved in the silencing of Igf2.