ChiP Seq for PU.1 in Rag Abelson Cells
All cells were treated with imatinib for 48 hours. ChIP was performed using anti-PU.1, anti-FLAG anti-HA anti-BCLAF1, control rabbit IgG and control mouse IgG antibodies DNA was cross-linked with 2% formaldehyde for 10 min at room temp (1 x 106 cells/ml). Reaction was stopped with 125 M Glycine. Cells were lysed with NP-40 and nuclei were frozen in liquid nitrogen then lysed with SDS. DNA was fragmented by sonicating with 30-second pulses for 60 cycles using a Bioruptor.DNA fragmentation was in the range of 200-500 bp and was monitored by agarose gel electrophoresis. Immunoprecipitation was performed with anti-PU.1 anti-HA anti-BCLAF1 or control rabbit IgG and Protein A Dynabeads DNA was eluted, reverse cross-linked and then purified with QIAquick PCR purification kit. For ChIP-PCR analysis, PCR was performed. For ChIP-seq analysis, fragmented DNA was quantified using 2100 Bioanalyzer and DNA libraries were prepared using Illumina TruSeq. Sequencing was performed using an Illumina HiSeq 3000 by the Washington University Genome Technology Access Center.