Identification and methylation analysis of putative CTCF binding sites. (A) Alignment of the putative GTL2 CTCF binding sites with known CTCF binding sites. GTL2(a)/GTL2(b), humanGTL2 promoter; H19, human H19 upstream region; FII, chicken βglobin insulator; Myc FV, chicken c-myc promoter; Apβ, human amyloid β protein promoter. The GTL2(b) consensus sequence is present on the noncoding strand, and is therefore inverted for the purposes of sequence alignment. (B) Methylation analysis of the putative GTL2 CTCF binding sites. Genomic DNA isolated from fetal liver was bisulphite treated and amplified across putative CTCF binding sites (a) and (b). Diamonds denote the positions of unmethylated cytosines not present in CpG dinucleotides; they have been fully converted by bisulphite treatment. The presence of bands in both the T and C lanes (arrowheads) shows hemimethylation of cytosine residues within the CpG dinucleotides contained within and adjacent to CTCF binding sites (a) and (b).
