FIG. 1.

DMD sequences containing tandem repeats restore maternal-specific methylation to the Ig/myc transgene. (A) Schematics of the mouse Snurf/Snrpn, Kcnq1, and Igf2r DMDs (white boxes). The Snrpn DMD contains the promoter and exon 1 of the Snurf/Snrpn locus, and the Kcnq1 and Igf2r DMDs contain promoters for untranslated antisense transcripts (bent arrows). Arrowheads or arrows within each DMD indicate tandem repeats (different for each DMD). Gray bars represent the regions analyzed in Ig/myc transgenes in this study and previously (21). The Kcnq1 region analyzed in this study included the conserved repeats examined by Mancini-Dinardo et al. plus additional DMD repeats (15). (B) Design of transgenes analyzed in this study. The Cα and Sα regions of the mouse IgA locus (Ig) and the 3′ untranslated region and c-myc exons (1-3) are shown. A black bar depicts the probe for the Southern blots. The DMD sequences depicted by gray bars in panel A were subcloned into an EcoRI site (flanking the black box). (C) Southern blot analysis of DNAs obtained from maternal and paternal carriers of each transgene. DNAs were digested with HpaII and hybridized with the probe shown in panel B. Band sizes are shown in kilobases (kb). SnrpnR, SnrpnR/Igmyc; Kcnq1, Kcnq1/Igmyc. (D) Summary of bisulfite genomic sequencing analyses performed on DNAs obtained from maternal and paternal carriers of the SnrpnR/Igmyc transgene. The top line shows the location of tandem repeats (arrows) and the CpGs analyzed (open circles). Each line below represents one sequenced allele. Filled circles indicate the positions of methylated CpGs. (E) Summary of bisulfite genomic sequencing data obtained from preimplantation stage embryos following maternal or paternal inheritance of the SnrpnR/Igmyc transgene. Mor, morulae; Blast, blastocysts. For the paternal allele only the eight-cell and blastocyst stages were examined. Each bar represents the percentage of CpGs found to be methylated in all of the subclones analyzed. M, maternal; P, paternal.