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Sample GSM1944731 Query DataSets for GSM1944731
Status Public on Feb 12, 2018
Title H3K27me3 ChIP-seq Replicate 1
Sample type SRA
 
Source name EML_C1
Organism Mus musculus
Characteristics strain: C57BL/6
passage: 4-5
antibody: H3K27me3, Millipore, cat no: 07-449, lot: 2382150
Growth protocol EML_C1 was cultured in IMDM medium supplemented with 20% FCS, 2 mM L-glutamine, 100 units/ml penicillin, 100 µg/ml streptomycin, 1.5 g/L sodium bicarbonate and 200 ng/ml mSCF.
Extracted molecule genomic DNA
Extraction protocol H3K27Me3 ChIP samples were fixed using a 1% formaldehyde (FA) fixation protocol for 10 min, while a 60 min, 2 mM DSG and a 60 min 1% FA double fixation protocol was used for Ezh2 and Runx1 antibodies. Fixed chromatin samples were fragmented using a S220 Focused-ultrasonicators (Covaris) for 10 min with 10% duty cycle, 5 intensity, 200 cycle per burst with frequent sweeping mode and constant degassing at 4°C to an average size of 200-500bps. Antibody:chromatin complexes were collected with a mixture of protein A and Protein G Dynabeads (Life Technologies) collected with a magnet, and washed 3X with a solution of 50 mM HEPES-KOH, pH 7.6, 500 mM LiCl, 1 mM EDTA, 1% NP-40, 0.7% Na-Deoxycholate. After a TE wash, samples were eluted, de-crosslinked, RNase and Proteinase K treated, and purified using a QIAGEN PCR purification kit. ChIP samples were quantified relative to inputs. A 50 μl aliquot taken from each of 1 ml of sonicated, diluted chromatin before antibody incubation serves as the input, thus the signal from the input samples represents 5% of the total chromatin used in each ChIP.
The library prep was performed on the Apollo system (Wafergen) with the kit PrepX Complete ILMN 32i DNA Library Kit (cat # 400076) and custom adapters.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2500
 
Data processing Reads were aligned with Bowtie2 (version 2.2.1) and the following parameters --maxins 1000 --no-mixed --no-discordant
Duplicate Reads were marked with Picard tools
Duplicate Reads and Secondary alignments were removed with samtools
Ezh2 and Runx1 peaks were called using MACS2 (version 2.0.10) and the following parameters --bdg -g mm --keep-dup 9999
H3K27me3 peaks were called using MACS2 (version 2.0.10) and the following parameters --bdg --broad -g mm --keep-dup 9999
Genome_build: mm10
Supplementary_files_format_and_content: narrowPeak and broadPeak peak files were generated with MACS2
 
Submission date Nov 19, 2015
Last update date May 15, 2019
Contact name Adam Mead
E-mail(s) adam.mead@imm.ox.ac.uk
Organization name University of Oxford
Department Weatherall Institute of Molecular Medicine
Lab HSCB Laboratory
Street address John Radcliffe Hospital
City Oxford
ZIP/Postal code OX3 9DS
Country United Kingdom
 
Platform ID GPL17021
Series (2)
GSE75186 Ezh2 and Runx1 Mutations Targeted to Early Lymphoid Progenitors Collaborate to Promote Early Thymic Progenitor Leukemia [ChIP-Seq]
GSE75188 Ezh2 and Runx1 Mutations Targeted to Early Lymphoid Progenitors Collaborate to Promote Early Thymic Progenitor Leukemia
Relations
BioSample SAMN04279040
SRA SRX1438364

Supplementary file Size Download File type/resource
GSM1944731_H3K27me3_rep1_peaks.broadPeak.gz 4.9 Mb (ftp)(http) BROADPEAK
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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