The imprintedU2af1-rs1gene that maps to mouse chromosome 11 is predominately expressed from the paternal allele. We examined the methylation of genomic sequences in and around theU2af1-rs1locus to establish the extent of sequence modifications that accompanied the silencing of the maternal allele. The analysis ofHapII orHhaI sites showed that the silent maternal allele was hypermethylated in a block of CpG sequences that covered more than 10 kb. By comparison, the expressed paternal allele was unmethylated from a CpG island upstream of the transcribed region through 2 kb. An analysis of DNaseI hypersensitivity of a putative promoter ofU2af1-rs1showed an open chromatin conformation only on the unmethylated, expressed paternal allele. These results suggest that allele-specific hypermethylation covering the gene and its upstream CpG island plays a role in maternal allele repression ofU2af1-rs1,which is reflected in altered chromatin conformation of DNaseI hypersensitive sites.