Chromatin Immunoprecipitation Assay Protocol
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Corresponding Organization : The Ohio State University Wexner Medical Center
Other organizations : Texas Biomedical Research Institute, Columbia University Irving Medical Center, University of Alabama at Birmingham
Variable analysis
- Antibodies recognizing the targeted proteins
- Protein-DNA complexes
- 1% paraformaldehyde (PFA) for cell cross-linking
- 125 mM glycine to quench the reaction
- CE buffer to re-suspend cells
- SDS lysis buffer and sonication to generate DNA fragments
- CHIP dilution buffer
- Protein A/G magnetic beads pre-blocked with 0.5 mg/ml BSA and 0.125 mg/ml herring sperm DNA
- Low-salt buffer, high-salt buffer, LiCl buffer and TE buffer for washing the beads
- Elution buffer to elute the IPed protein-DNA complexes
- 0.2 M NaCl and 65°C overnight incubation to recover DNA samples
- EDTA, Tris–HCl (pH 6.5), and proteinase K treatment
- Phenol/chloroform/isoamyl alcohol (25:24:1) extraction
- Nuclease-free water to re-suspend DNA pellets
- Control mouse/rabbit IgG
- Input (5%)
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