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. 2018 Aug 8;27(23):3999–4011. doi: 10.1093/hmg/ddy274

Figure 3.

Figure 3

Loss of methylation at the maternal PWS-IC in Ctrl B iPSC results in loss of imprinting of genes at the chr15q11-q13. (A) PWS-IC and IG-DMR COBRA for Ctrl and AS FIBs and several iPSCs (Ctrl A, Ctrl B, Ctrl E; AS A and AS E). White circle, unmethylated band; half black circle, partially methylated band; black circle, fully methylated band. (B) Bisulfite sequencing analysis for PWS-IC in Ctrl A, Ctrl B and AS A iPSCs. Each line represents the methylation profile of an independent PCR amplicon as determined by Sanger sequencing and BiQ Analyser; white dots, unmethylated CpGs; black dots, methylated CpGs; Bsh1236I recognition sites used for PWS-IC COBRA are indicated; the numbers in brackets represent the number of times the same amplicon was sequenced; estimated number of bacterial clones obtained in the LB agar plate is indicated. (C) Allelic-specific SNRPN expression analysis assayed by Sanger sequencing for the Ctrl A, Ctrl B and AS E iPSCs. Yellow stripe indicates the rs705 SNP. (D) Nascent-transcript RNA FISH for UBE3A and SNORD116 in Ctrl E, Ctrl B and AS E iPSCs. Percentage of biallelic/monoallelic UBE3A and SNORD116 FISH signals was calculated by counting a minimum of 100 cells per sample.