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Sample GSM6751547 Query DataSets for GSM6751547
Status Public on Apr 24, 2023
Title U2OS31_T2h_H3K27ac_Rep1
Sample type SRA
 
Source name U2OS
Organism Homo sapiens
Characteristics cell line: U2OS
integrated construct: H3.1 FLAG
hours post_infection: T2h
chip antibody: H3K27ac
Extracted molecule genomic DNA
Extraction protocol Cells were crosslinked with 1% formaldehyde for 10 min at room temperature and the reaction was quenched with glycine at a final concentration of 0.125 M. Chromatin was sheared using ultrasonication (Covaris S2). A total of 2.5 µg of antibody against H3K27ac (Abcam, ab4729), or FLAG (M2, Sigma Aldrich, F3165), was bound to 20 µl pre-washed Dynabeads Protein A (Thermo Fisher Scientific) in a total volume of 200 µl PBS containing 0.02% Tween-20 for 1h at room temperature rotating at 6 rpm. The buffer was removed on a magnet and antibody-coupled beads were resuspended in 120µl dilution buffer (20mM Tris pH7.4, 2mM EDTA, 100mM NaCl, 0.5% Triton x-100) containing proteinase inhibitors (cOmplete™, EDTA-free Protease Inhibitor Cocktail, 50x, Sigma). 80µl sonicated chromatin was added to the antibody-coupled Dynabeads (sonicated chromatin of approx. 1.7x106 cells) and incubated for 3h at room temperature rotating at 6 rpm. Beads were washed on a magnet and chromatin was eluted. After crosslink reversal, RNase A and proteinase K treatment, DNA was extracted with the Monarch PCR & DNA Cleanup kit (NEB).
Sequencing libraries were prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB) according to manufacturer’s instructions
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model NextSeq 550
 
Data processing Paired-end reads were mapped to the reference using bowtie2-aligner with following paramters: --very-sensitive-local, --no-discordant
Reads were filtered to mapping quality > 30 and only reads mapped in proper pair were kept using samtools.
CPM-normalized coverage tracks were generated using the bamCoverage script of the deepTools package
Assembly: hg19 and AY339865.1 (adapted)
Supplementary files format and content: bigwig files showing CPM normalized coverage
 
Submission date Nov 22, 2022
Last update date Apr 24, 2023
Contact name Uwe Schwartz
E-mail(s) uwe.schwartz@ur.de
Organization name University of Regensburg
Department NGS Analysis Center
Street address Universitätstraße 32
City Regensburg
ZIP/Postal code 93053
Country Germany
 
Platform ID GPL21697
Series (2)
GSE136550 Changes in adenoviral chromatin organization precede early gene activation
GSE218541 Changes in adenoviral chromatin organization precede early gene activation [ChIP-Seq]
Relations
BioSample SAMN31836799
SRA SRX18348445

Supplementary file Size Download File type/resource
GSM6751547_Rep1_U2OS31_T2h_H3K27ac.bw 55.3 Mb (ftp)(http) BW
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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