BGL3 is preferentially expressed in cells containing high HbF. (A) Map of the human β-globin locus showing the positions of primer pairs used to determine RNA transcript levels and factor occupancy by ChIP and real-time quantitative PCR (vertical lines). The globin genes and LCR DNase I–hypersensitive sites are represented by blue and red rectangles, respectively. The BGL3 region is depicted by a green rectangle. (B) Globin gene and intergenic transcription in K562 cells was evaluated using reverse-transcribed total RNA and quantitative RT-PCR with primers specific to the regions indicated on the x-axis. White bars indicate no reverse transcriptase. Results were normalized to GAPDH and error bars indicate SD. (C) Globin gene and intergenic transcription was evaluated for low-HbF (black bars) and high-HbF cells (white bars) as in panel B. Dark and light gray bars indicate values for reactions with no reverse transcriptase for low- and high-HbF cells, respectively. (D) ChIP was performed using an RNA Polymerase II Ab for low- and high-HbF cells. The locations of primers for PCR are indicated on the x-axis. Results were calculated against an input sample and normalized to GAPDH. Error bars indicate SD; black bars, low-HbF cells; white bars, high-HbF cells. Dark and light gray bars indicate values for an isotype matched control Ab in low- and high-HbF cells, respectively. The dotted line indicates the average of IgG control precipitations across all amplicons. (E) ChIP was performed using an H3K4me3 Ab to determine occupancy in low- and high-HbF cells. All data were calculated and plotted as described in panel D. *P < .05.