Figure 4.

Disrupted polyadenylation in maternal imprint-free embryos. (A) Methylation status of the Mcts2 CpG island in pooled embryos (n = 5) derived from Dnmt3l^−/− dams and wild-type cast sires (Dnmt3l^−/+) and control embryos, determined by sequencing bisulphite-modified genomic DNA. Parental strands were distinguished on the basis of a SNP between Mus domesicus (B6) and Mus musculus (cast) alleles. Filled circles are methylated, and closed circles are unmethylated. Strands derived from the same PCR amplification are joined by braces on the right. (C) Allele-specific RT–PCR sequencing assay for Mcts2 in Dnmt3l^−/+ hybrid embryos (pooled, n = 5). (C) Qualitative 3′ RACE in cDNA from pooled Dnmt3l^−/+ hybrid embryos. (D) 3′ RACE products from C were excised and sequenced over regions containing SNPs between the two parental strains.