Fig. 6: Imprinting status analysis of Dlk1 and DNA methylation analysis of DMRs in the Dlk1-Dio3 imprinted domain in Gtl2 polyA knock-in placentas at E12.5.

a Monoallelic expression of Dlk1 in WT and PKI placentas and biallelic expression of Dlk1 in MKI and HOMO placentas. Allelic-specific gene expression is determined by the SNP between C57BL/6 N (B6) and DBA/2 J (DBA) mice. Total RNA are extracted from Gtl2 polyA knock-in placentas and their WT control. After purifying RNA and RT-PCR, PCR products containing the SNP for Dlk1 are sequenced. Blue arrows indicate monoallelic Dlk1 expression exclusively from the paternal allele and red arrows indicate detected biallelic Dlk1 expression. b Schematic representation of DMRs (IG-DMR, Gtl2-DMR, Dlk1-DMR) in Dlk1-Dio3 domain. The inserted sequence is in orange arrow. Filled circles represent methylated DMRs and hollow circles represent unmethylated DMRs. Green vertical lines represent the CpG dinucleotides in each DMR, and the yellow vertical lines represent SNPs between B6 and DBA mice. c–e Methylation status of IG-DMR, Gtl2-DMR and Dlk1-DMR in Gtl2 polyA knock-in and WT placentas separately. Each row represents one clone. SNPs are used to distinguish paternal allele clones from maternal allele clones. Filled and hollow circles indicate methylated and unmethylated CpG sites, respectively.