FIG. 4.

ChIP assays confirm the proximity of the insulator and promoter and enhancer elements on the maternal chromosome. (A) Schematic representation of the Igf2-H19 locus including the three Igf2 promoters, the ICR, the H19 promoter, and the shared endodermal (open circle) and mesodermal (closed circle) enhancers. Numbers above the line indicate the relative positions of the corresponding elements. Arrowheads indicate the locations and orientations of primers used the ChIP analysis. (B and C) Cross-linked extracts were prepared from muscle (B) and liver (C) cells and analyzed by ChIP using polyclonal antisera specific to CTCF protein. ICR, enhancer, and Igf2 promoter 2 sequences were identified using the primers indicated and quantitated by testing PCR products after 41, 43, and 45 cycles. In addition, primers specific to the CD3 locus were tested as a nonspecific control. + Ab, with antisera; − Ab, without antisera.