ChIP-seq Protocol for Epigenetic Marks

ChIP was performed as previously described (Schwartz et al. 2012 (link); Davidovich et al. 2013 (link)) with the exceptions noted in the Supplemental Material. For immunoprecipitation, 5–25 µg of solubilized chromatin was used with 2 µg of α-RNA Pol II antibody (EMD Millipore, catalog no. 05-623), α-H3 (Abcam, ab-1791), α-H3K4me2/3 (Abcam, ab-6000), α-H3K27me3 (EMD Millipore, 07-449), or 4 µg of α-GABPα (Santa Cruz Biotechnology, H-180 sc-22810) and nutated overnight at 4°C.
All replicates reported in this study represent independent biological samples.

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Stern J.L., Theodorescu D., Vogelstein B., Papadopoulos N, & Cech T.R. (2015). Mutation of the TERT promoter, switch to active chromatin, and monoallelic TERT expression in multiple cancers. Genes & Development, 29(21), 2219-2224.

Publication 2015

ab-1791 ab-6000

Antibody Biological Chip Chromatin Immunoprecipitation Rna pol ii

Corresponding Organization :

Other organizations : University of Colorado Boulder, Howard Hughes Medical Institute, University of Colorado Cancer Center, University of Colorado Anschutz Medical Campus, University of Colorado Denver, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University

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Variable analysis

independent variables

Amount of solubilized chromatin used for immunoprecipitation (5–25 µg)
Antibodies used for immunoprecipitation (α-RNA Pol II, α-H3, α-H3K4me2/3, α-H3K27me3, α-GABPα)

dependent variables

Not explicitly mentioned

control variables

Experimental protocol followed as previously described (Schwartz et al. 2012 and Davidovich et al. 2013), with exceptions noted in Supplemental Material
Overnight incubation of solubilized chromatin and antibodies at 4°C with nutation

controls

Positive controls: Not explicitly mentioned
Negative controls: Not explicitly mentioned

Annotations

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