|
| Status |
Public on Feb 06, 2023 |
| Title |
ChOR_T480_H2BK120ub_r1 |
| Sample type |
SRA |
| |
|
| Source name |
Embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
cell line: E14JU
strain: 129/Ola
genotype: wildtype
treatment: None
|
| Treatment protocol |
Cells were incubated in EdU media (f.c. 10 µm) for 10 min and then either collected (T0) or cultured for indicated maturation intervals (T15-T480, in minutes) in media without EdU before collection. If treated with DMSO (volume equivalent), Triptolide (10uM), Indole-3-acetic acid (500uM) and/or dTAG-13 (1uM), cells were incubated for indicated times in the description and then collected.
|
| Growth protocol |
ESCs were grown on gelatin-coated dishes in serum+LIF conditions at 37 °C with 5 % CO2. DMEM was supplied with fetal bovine serum (15 %), home-made LIF, non-essential amino acids, penicillin/streptomycin and beta-mercaptoethanol. Cells were passaged using Trypsin-EDTA or TrypLE.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
After the appropriate treatment, cells were washed with cold PBS, crosslinked for 5min with 1% Formaldehyde at room temperature and subsequently quenched with 0.1M Glycin (final concentration) for 5min before harvesting. Then, nuclei were isolated and sonicated using the Covaris Chromatin Shearing Kit. ChIP-Immunoprecipation of chromatin was done using antibodies according to the manufacturers recommendation with EdU-labelled Drosophila S2 chromatin serving as Spike-In control and, where the epitope of the antibody does not recognize the Drosophila homolog (i.e. H2A.Z) , H2AV antibodies to immunoprecipitate Spike-In chromatin as well. Purified fragments were biotinylated, libraries prepared, EdU-biotinylated fragments pulled down using Streptavidin and sequenced
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
EdU-labelled genomic DNA
ChOR_T480_H2BK120ub_RRPM_MEAN.bw
|
| Data processing |
adapters and low quality reads were filtered with TrimGalore (v0.0.6)
reads were mapped to the genome with bowtie2 (v2.4.2) and uniquely mapped peaks kept using samtools (v1.12)
duplicate reads were removed with picard (version 2.26.10)
bam files were read into Seqmonk (v1.47.1) with MAPQ>20, ignoring duplicates, and reads extended by 250bp. Samples were normalized based on Drosphila spike-In abundance and corresponding ClickedInputs. Replicates were merged, exported as bedgraphs and converted to bigwig using UCSC BedGraphtoBigWig
Assembly: mm10
Assembly: dm6
Supplementary files format and content: bigwig files for merged replicates (RRPM-adjusted) for the mouse genome
Library strategy: ChOR-seq
|
| |
|
| Submission date |
May 27, 2022 |
| Last update date |
Feb 06, 2023 |
| Contact name |
Anja Groth |
| E-mail(s) |
anja.groth@cpr.ku.dk
|
| Organization name |
Novo Nordisk Foundation Center for Protein Research
|
| Street address |
Blegdamsvej 3B
|
| City |
Copenhagen |
| ZIP/Postal code |
2200 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE204981 |
Recycling of H2A-H2B provides short-term memory of chromatin states [ChOR-Seq] |
| GSE204988 |
Recycling of H2A-H2B provides short-term memory of chromatin states |
|
| Relations |
| BioSample |
SAMN28698725 |
| SRA |
SRX15725689 |
