Broad H3 ChIPseq Track Settings
 
Broad Institute Chromatin State Mapping using ChIP-Seq   (All Expression and Regulation tracks)

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 ES  H3K4me1  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me1 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K4me1  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me1 ab, Embryonic Stem (ES) cells)   Data format 
 
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 NP  H3K4me1  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me1 ab, Neural Progenitor (NP) cells)   Data format 
 
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 NP  H3K4me1  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me1 ab, Neural Progenitor (NP) cells)   Data format 
 
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 ES  H3K4me2  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me2 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K4me2  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me2 ab, Embryonic Stem (ES) cells)   Data format 
 
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 NP  H3K4me2  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me2 ab, Neural Progenitor (NP) cells)   Data format 
 
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 NP  H3K4me2  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me2 ab, Neural Progenitor (NP) cells)   Data format 
 
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 Whole Brain  H3K4me2  Sites-Windowing  Broad Stem Cell Chromatin IP Sites By Window (H3K4me2 ab, Whole Brain)   Data format 
 
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 Whole Brain  H3K4me2  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me2 ab, Whole Brain)   Data format 
 
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 ES  H3K4me3  Sites-HMM  Broad Stem Cell Chromatin IP Sites by HMM (H3K4me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K4me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K4me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 MEF  H3K4me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 MEF  H3K4me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 NP  H3K4me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K4me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 NP  H3K4me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 ES-hybrid  H3K4me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me3 ab, Hybrid Embryonic Stem (ES-hybrid) cells (129/CAST))   Data format 
 
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 Whole Brain  H3K4me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites By Window (H3K4me3 ab, Whole Brain)   Data format 
 
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 Whole Brain  H3K4me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K4me3 ab, Whole Brain)   Data format 
 
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 ES  H3K9me3  Sites-HMM  Broad Stem Cell Chromatin IP Sites by HMM (H3K9me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K9me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K9me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K9me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K9me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 MEF  H3K9me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K9me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 MEF  H3K9me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K9me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 NP  H3K9me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K9me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 NP  H3K9me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K9me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 ES-hybrid  H3K9me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K9me3 ab, Hybrid Embryonic Stem (ES-hybrid) cells (129/CAST))   Data format 
 
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 ES  H4K20me3  Sites-HMM  Broad Stem Cell Chromatin IP Sites by HMM (H3K20me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H4K20me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H4K20me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K27me3  Sites-HMM  Broad Stem Cell Chromatin IP Sites by HMM (H3K27me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K27me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K27me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K27me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K27me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 MEF  H3K27me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K27me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 MEF  H3K27me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K27me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 NP  H3K27me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites by Window (H3K27me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 NP  H3K27me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K27me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 Whole Brain  H3K27me3  Sites-Windowing  Broad Stem Cell Chromatin IP Sites By Window (H3K27me3 ab, Whole Brain)   Data format 
 
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 Whole Brain  H3K27me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K27me3 ab, Whole Brain)   Data format 
 
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 ES  H3K36me3  Sites-HMM  Broad Stem Cell Chromatin IP Sites by HMM (H3K36me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  H3K36me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K36me3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 MEF  H3K36me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K36me3 ab, Embryonic Fibroblasts (MEF))   Data format 
 
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 NP  H3K36me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K36me3 ab, Neural Progenitor (NP) cells)   Data format 
 
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 ES-hybrid  H3K36me3  Density Signal  Broad Stem Cell Chromatin IP Signal (H3K36me3 ab, Hybrid Embryonic Stem (ES-hybrid) cells (129/CAST))   Data format 
 
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 ES  pan-H3  Density Signal  Broad Stem Cell Chromatin IP Signal (pan-H3 ab, Embryonic Stem (ES) cells)   Data format 
 
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 ES  RPol-II  Density Signal  Broad Stem Cell Chromatin IP Signal (RNA Polymerase II, Embryonic Stem (ES) cells)   Data format 
    
Assembly: Mouse Feb. 2006 (NCBI36/mm8)

Description

The tracks contain chromatin-state maps generated at the Broad Institute using ChIP-Seq.

Sites by HMM Enriched intervals inferred by an HMM
Sites by WindowingEnriched intervals inferred by fixed-size windows
Signal Densities ChIP-Seq fragment densities analyzed
Alignments Sequences and coordinates of uniquely aligned reads

Sites by HMM

These subtracks contain the coordinates of intervals enriched for H3K4Me3, H3K9Me3, H3K27Me3, H3K36Me3 or H4K20Me3 in mouse ES cells, as determined by a Hidden Markov Model. Prior to analysis, the fragment densities were discretized into four classes (masked, sub-threshold, near-threshold, above-threshold), which were defined for each sample by supervised training on a 10 Mb subset of the genome.

Sites by Windowing

These subtracks contain the coordinates of intervals significantly enriched for H3K4Me3, H3K4me2, H3K4me1 or H3K27Me3 in ES cells, ES-derived neural progenitors (NP), whole brain tissue and embryonic fibroblasts (MEF). Intervals were obtained by comparing the average densitity of a 1-kb window to the cumulative density distribution expected from randomly aligning reads across the unique portion of the reference genome, and merging any overlapping windows with nominal p-values of < 10-5.

The coordinates indicate the beginning of the first window and the end of the last window. The fourth column indicates the maximal fragment density within each interval.

H3K9Me3 enriched sites have been added since the initial publication of this data. These sites correspond to a somewhat more permissive threshold than the HMM-based sites analyzed in the manuscript. The maximum fragment density can be used to rank the sites by confidence.

Signal Densities

These subtracks contains the ChIP-Seq fragment densities analyzed in the paper. They represent the density of antibody enriched fragments in celltype at 25 bp resolution across the mouse genome.

A positive value indicates the number of uniquely aligned fragments oriented towards, and within 300 bp of, a particular position. Reads within 200 bp add 1 to the density, and reads within 300 bp add 0.25. (This is an approximation of the expected number of fragments overlapping the position, since the exact length of each is unknown).

Alignments

These subtracks contain alignment coordinates of the ChIP-Seq reads on the mouse reference genome (UCSC version mm8).

SAMPLE ANTIBODIES
ES cells Whole cell extract (unenriched input)pan-H3H3K4Me3H3K4Me2H3K4Me1H3K27Me3H3K36Me3H3K9Me3H4K20Me3RNA polymerase II
ES hybrid cells (129/CAST)   H3K4Me3    H3K36Me3  H3K9Me3   
MEF Whole cell extract (unenriched input) H3K4Me3    H3K27Me3H3K36Me3H3K9Me3   
NP cells Whole cell extract (unenriched input) H3K4Me3H3K4Me2H3K4Me1H3K27Me3H3K36Me3H3K9Me3   
Whole brain    H3K4Me3H3K4Me2  H3K27Me3       

Methods

Growth protocol

V6.5 murine ES cells (genotype 129SvJae X C57BL/6; male; passages 10-15) and hybrid murine ES cells (genotype 129SvJae X M. m. castaneus F1; male; passages 4-6) were cultivated in 5% CO at 37C on irradiated MEFs in DMEM containing 15% FCS, LIF, penicillin/streptomycin, L-glutamine, nonessential amino-acids and 2-mercaptoethanol. Cells were subjected to at least 2-3 passages on 0.2% gelatin under feeder-free conditions prior to analysis. V6.5 ES cells were differentiated into neural progenitor (NP) cells through embryoid body formation for 4 days and selection in ITSFn media for 5-7 days, and maintained by FGF2 and EGF2. Mouse embryonic fibroblasts (genotype 129SvJae X C57BL/6; male; E13.5; passages 4-6) were grown in DMEM with 10% fetal bovine serum and penicillin/streptomycin at 37C, 5% CO2. Primary tissues were isolated from 4-6 weeks old male 129SvJae/C57BL/6 mice.

Extraction protocol

Cells and homogenized tissues were fixed in 1% formaldehyde and resuspended in lysis buffer. Chromatin was sheared to 200-700 bp using a Branson 250 Sonifier or Diagenode Bioruptor. Solubilized chromatin was immunoprecipitated with antibodies against H3K4me3 (Abcam 8580), H3K9me3 (Abcam 8898), H3K27me3 (Upstate 07-449), H3K36me3 (Abcam 9050), H4K20me3 (Upstate 07-463), pan-H3 (Abcam 1791) or RNA polymerase II (Covance MMS-126R). Antibody-chromatin complexes were pulled-down using protein A-sepharose (or anti-IgM-conjugated agarose for RNA polymerase II), washed and then eluted. After cross-link reversal and proteinase K treatment, immunoprecipitated DNA was extracted with phenol-chloroform, ethanol precipitated, treated with RNAse and purified. One to ten nanograms of DNA were end-repaired, adapter-ligated and sequenced by Illumina Genome Analyzers as recommended by the manufacturer.

Data processing

Sequence reads from each IP experiment were aligned to the mouse reference genome (mm8) using the ARACHNE computational pipeline. First, a table was pre-computed to associate all possible 12-mers with all of their occurrences in the genome. For each ChIP-Seq read (forward and reverse complement orientation), each potential start point was then found and the number of mismatches in the corresponding gap-free alignment was computed. All uniquely aligned reads (defined as the second to best alignment having >2 mismatches more than the best alignment, and the total mismatch count being <=6) were kept. If multiple reads aligned to the same starting position, only one were kept. Fragment densities were computed by counting the number of reads (extended to 300 bp) overlapping each position in the genome (at 25 bp resolution). Non-unique positions in the reference genome were pre-computed by aligning every 27-mer in the genome to the whole genome and masking positions that did not meet the uniqueness criteria defined above.

Credits

The ChIP-Seq data were generated at the Broad Institute and in the Bradley E. Bernstein lab at Massachusetts General Hospital/Harvard Medical School, in collaboration with the Rudolf Jaenisch lab at the Whitehead Institute.

Data generation was supported by funds from the NHGRI, the NCI, the Burroughs Wellcome Fund, Massachusetts General Hospital and the Broad Institute.

References

Mikkelsen TS, Ku M, Jaffe DB, Issac B, Lieberman E, Giannoukos G, Alvarez P, Brockman W, Kim TK, Koche RP et al. Genome-wide maps of chromatin state in pluripotent and lineage-committed cells. Nature. 2007 Aug 2;448(7153):553-60. PMID: 17603471; PMC: PMC2921165

Meissner A, Mikkelsen TS, Gu H, Wernig M, Hanna J, Sivachenko A, Zhang X, Bernstein BE, Nusbaum C, Jaffe DB et al. Genome-scale DNA methylation maps of pluripotent and differentiated cells. Nature. 2008 Aug 7;454(7205):766-70. PMID: 18600261; PMC: PMC2896277