ChIP was performed by standard protocols for ChIP-qPCR and ChIP (12 (link),26 (link)). Antibodies were specific for RNAPII (Abcam ab76123 for ChIP-qPCR or Cell Signaling Technology 14958S for ChIP-seq), CEBPβ (Santa Cruz sc-7962), H3K27Ac (Millipore 07-360), CTCF (Millipore 07-729) or rabbit IgG (Millipore 12-370). All ChIP-seq experiments were performed twice and irreproducible discovery rate (IDR) data are shown. Raw reads were processed using the AQUAS Transcription Factor and Histone ChIP-Seq processing pipeline (https://github.com/ENCODE-DCC/chip-seq-pipeline2 ) using the ENCODE (phase-3) guidelines on the hg19 reference genome. This includes mapping using BWA and peak calling with MACS2. Primer sequences used for qPCR are shown in Supplementary Table S2 .
Protocol detail
ChIP-seq and ChIP-qPCR Protocols for RNAPII, CEBPβ, H3K27Ac, CTCF
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Corresponding Organization : Case Western Reserve University
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Variable analysis
independent variables
- ChIP protocols (ChIP-qPCR and ChIP-seq)
- RNAPII binding
- CEBPβ binding
- H3K27Ac levels
- CTCF binding
- Antibodies used for ChIP (RNAPII, CEBPβ, H3K27Ac, CTCF, rabbit IgG)
- Processing of raw ChIP-seq reads (using AQUAS Transcription Factor and Histone ChIP-Seq processing pipeline)
- Mapping using BWA and peak calling with MACS2
- Experiments performed in duplicate and IDR data shown
- Reference genome (hg19)
- Positive control: None specified
- Negative control: Rabbit IgG
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