Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-Add protease inhibitors to all lysis solutions before use.	-Keep cells on ice between the rounds of homogenisations. - Increase or decrease the homogenization step to maximize the yield of nuclei depending on cell line. -Do not place ethidium bromide in the agarose gel or the electrophoresis buffer, because of the presence of SDS.

Upstream tips
-Add protease inhibitors to all lysis solutions before use.
Protocol tips
-Keep cells on ice between the rounds of homogenisations. - Increase or decrease the homogenization step to maximize the yield of nuclei depending on cell line. -Do not place ethidium bromide in the agarose gel or the electrophoresis buffer, because of the presence of SDS.

Publication protocol

ChIP-exo was performed as previously described6,20 with chromatin extracted from 50 million cells, ProteinG MagSepharose resin (GE Healthcare), and 5 ug of antibody directed against RNA polymerase II (Santa Cruz sc899), H2A.Z (EMD Millipore 07-594), H3K4me3 (Abcam ab8580), H3K4me2 (Abcam ab7766), H3K4me1 (Abcam ab8895), H3K27ac (Abcam ab4729), H4K9Ac (Abcam ab4441), H3K27me3 (EMD Millipore 07-449), H4K20me1 (Abcam ab9051), and CTCF (EMD Millipore 07-729). Two biological replicates were prepared for each ChIP target. To enable future comparisons to drug treatment experiments, cells were treated with vehicle (final 0.03% DMSO (dimethyl sulfoxide)) for 4 h prior to harvest. Libraries were sequenced using an Illumina NextSeq500 sequencer as single-end reads 50 or 75 nucleotides in length

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Manufacturer protocol

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