Abstract
Chromatin immunoprecipitation (ChIP), originally developed by John T. Lis and David Gilmour in 1984, has been useful to detect DNA sequences where protein(s) of interest bind. ChIP is comprised of several steps: (1) cross-linking of proteins to target DNA sequences, (2) breaking genomic DNA into 300–1000 bp pieces by sonication or nuclease digestion, (3) immunoprecipitation of protein bound to target DNA with an antibody, (4) reverse cross-linking between target DNA and the bound protein to liberate the DNA fragments, and (5) amplification of target DNA fragment by PCR. Since then, the technology has evolved significantly to allow not only amplifying target sequences by PCR, but also sequencing all DNA fragment bound to a target protein, using a variant of the approach called the ChIP-seq technique (1). Another variation, the ChIP-on-ChIP, allows the detection of protein complexes bound to specific DNA sequences (2).
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Acknowledgments
This research project was supported by PSM-2U54 CA163071-06 and MCC-2U54 CA163068-06 from the National Institutes of Health. The project was also supported by 2U54MD007587 from the PRCTRC, G12MD007579 from RCMI, 4R25GM082406-10 from RISE, The Puerto Rico Science, Technology and Research Trust, and Ponce Medical School Foundation Inc. under the cooperative agreement 2016-00026. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
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Lee, M., Gudas, L.J., Saavedra, H.I. (2018). Detection of E2F-DNA Complexes Using Chromatin Immunoprecipitation Assays. In: Santiago-Cardona, P. (eds) The Retinoblastoma Protein. Methods in Molecular Biology, vol 1726. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7565-5_13
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DOI: https://doi.org/10.1007/978-1-4939-7565-5_13
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