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Series GSE103543 Query DataSets for GSE103543
Status Public on Sep 20, 2018
Title Internally Calibrated ChIP-seq (ICeChIP-seq) using a large panel of antibodies against H3K4 methylations to measure genome-wide histone modification density in mammalian cells
Organisms Homo sapiens; Mus musculus
Experiment type Genome binding/occupancy profiling by high throughput sequencing
Summary Post-translational modifications (PTMs) on histone proteins regulate genome accessibility and are frequently studied using chromatin immunoprecipitation (ChIP). In ChIP, an antibody putatively specific towards a histone PTM is used to map its genomic locations. ChIP experiments assume perfect antibody-epitope specificity, an assumption previously shown to be problematic, largely through peptide array studies. Among the most well-studied histone PTMs are the mono-, di-, and tri-methylation states of histone H3 lysine 4 (H3K4). While each state has been ascribed different biological functions, the methylation state specificity of antibodies used in these studies has not been systematically interrogated. Here, we use internally calibrated ChIP (ICeChIP) to comprehensively define the specificities of 52 commercially available antibodies marketed to distinguish the three methylation states of H3K4, allowing identification of both high- and low-specificity antibodies. We then conduct ICeChIP-seq with 18 such antibodies of varying specificity. We further find that the sum of H3K4me1 and H3K4me2 across enhancers contacting a promoter correlates strongly with gene expression for all genes, including housekeeping genes, and note that use of low-quality antibodies yields materially different biological interpretations. These results illustrate the way by which variable specificity of commercial antibodies contributes to the “reproducibility crisis” in biological research and demonstrates the need to carefully validate antibodies with techniques appropriate for the intended applications.
Overall design Semi-synthetic nucleosome standards are spiked into ChIP experiments to conduct internally calibrated ChIP-seq (ICeChIP-seq) with 17 anti-H3K4 methylation antibodies of varying specificities in K562 cells and one anti-H3K4me1 antibody in WT and dCD MLL3/4 R1 mESC cells.
Contributor(s) Shah RN, Grzybowski AT, Cornett EM, Johnstone AL, Dickson BM, Boone BA, Cheek MA, Cowles MW, Maryanski D, Meiners MJ, Tiedemann RL, Vaughan RM, Arora N, Sun Z, Rothbart SB, Keogh M, Ruthenburg AJ
Citation(s) 30244833, 33872311
NIH grant(s)
Grant ID Grant title Affiliation Name
R44 HG008907 Internally Calibrated Chromatin Immunoprecipitation Using Barcoded Nucleosomes EPICYPHER INC Michael-Christopher Keogh
R44 HG008907 Internally Calibrated Chromatin Immunoprecipitation Using Barcoded Nucleosomes EPICYPHER INC Zu-Wen Sun
R00 CA181343 Mechanisms Regulating DNA Methylation Maintenance in Chromatin VAN ANDEL RESEARCH INSTITUTE Scott Barri Rothbart
R35 GM124736 Molecular mechanisms of chromatin and epigenetic regulation VAN ANDEL RESEARCH INSTITUTE Scott Barri Rothbart
R01 GM115945 Quantitatively probing intra-nucleosomal chromatin variation and function UNIVERSITY OF CHICAGO Alexander Jackson Ruthenburg
Submission date Sep 06, 2017
Last update date Jul 25, 2021
Contact name Rohan Nishant Shah
Organization name University of Chicago
Street address CLSC 856, 920 E. 58th Street
City Chicago
State/province IL
ZIP/Postal code 60637
Country USA
Platforms (3)
GPL15520 Illumina MiSeq (Homo sapiens)
GPL16417 Illumina MiSeq (Mus musculus)
GPL18573 Illumina NextSeq 500 (Homo sapiens)
Samples (25)
GSM2773392 K562_AB-8895_H3K4me1_ICeChIP
GSM2773393 K562_AM-39635_H3K4me1_ICeChIP
GSM2773394 K562_CST-5326BF_H3K4me1_ICeChIP
BioProject PRJNA401866
SRA SRP116896

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE103543_AR16_H3K4me1-Corrected_Error_95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me1-Corrected_Error_95CI.bigwig 10.5 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me1-Corrected_HMD.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me1-Corrected_HMD_minus95CI.bedgraph.gz 6.2 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me1-Corrected_HMD_minus95CI.bigwig 10.7 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me1-Corrected_HMD_plus95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me1-Corrected_HMD_plus95CI.bigwig 10.5 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me1-Corrected_High_Confidence_Peaks.bed.gz 3.9 Mb (ftp)(http) BED
GSE103543_AR16_H3K4me1_Corrected_HMD.bigwig 10.6 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me2-Corrected_Error_95CI.bedgraph.gz 6.2 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me2-Corrected_Error_95CI.bigwig 10.5 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me2-Corrected_HMD.bedgraph.gz 6.4 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me2-Corrected_HMD_minus95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me2-Corrected_HMD_minus95CI.bigwig 10.7 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me2-Corrected_HMD_plus95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me2-Corrected_HMD_plus95CI.bigwig 10.6 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me2-Corrected_High_Confidence_Peaks.bed.gz 2.9 Mb (ftp)(http) BED
GSE103543_AR16_H3K4me2_Corrected_HMD.bigwig 10.6 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me3-Corrected_Error_95CI.bedgraph.gz 6.2 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me3-Corrected_Error_95CI.bigwig 10.5 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me3-Corrected_HMD.bedgraph.gz 6.5 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me3-Corrected_HMD_minus95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me3-Corrected_HMD_minus95CI.bigwig 10.6 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me3-Corrected_HMD_plus95CI.bedgraph.gz 6.3 Gb (ftp)(http) BEDGRAPH
GSE103543_AR16_H3K4me3-Corrected_HMD_plus95CI.bigwig 10.6 Gb (ftp)(http) BIGWIG
GSE103543_AR16_H3K4me3-Corrected_High_Confidence_Peaks.bed.gz 672.4 Kb (ftp)(http) BED
GSE103543_AR16_H3K4me3_Corrected_HMD.bigwig 10.7 Gb (ftp)(http) BIGWIG
GSE103543_RAW.tar 757.6 Gb (http)(custom) TAR (of BED, BEDGRAPH, BIGWIG)
GSE103543_README_Corrected_files.txt 1.7 Kb (ftp)(http) TXT
GSE103543_hg38_bar_Ca_1-17.fa.gz 907.2 Mb (ftp)(http) FA
GSE103543_mm9_bar_Ca_1-17.fa.gz 819.8 Mb (ftp)(http) FA
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