|
| Status |
Public on Apr 27, 2022 |
| Title |
ChIP-seq Input (Ezh2, H3K4me3, H3K27ac) (ctrl) |
| Sample type |
SRA |
| |
|
| Source name |
Embryonic Stem Cells
|
| Organism |
Mus musculus |
| Characteristics |
cell type/genotype: mESC with conditional Smarcb1 knockdown
treatment: untreated
smarcb1 expression: unchanged
chip-seq antibody: IgG: Novusbio, #NBP2-24891
cell line: 81K1 (based on OG2)
|
| Treatment protocol |
Smarcb1 kd cells were treated with 1µg doxycycline per ml medium every day. ChIP-seq experiments were performed 72h after knockdown induction
|
| Growth protocol |
mESC were cultured on gelatine-coated plates and re-plated every other day. DMEM Glutamax-based medium was supplemented with 10% FBS superior, 1% MEM non-essential amino acids, 1% beta-Mercaptoethanol and 5µg LIF (per 500ml medium).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Ezh2, H3K4me3, H3K27ac: Cells were fixed in 1 % formaldehyde, quenched with 0.125 M Glycine, extraction was performed with ice-cold lysis buffer according to our laboratory standards. Samples were sonified, ChIP was performed with the indicated antibodies and afterwards samples were treated with elution buffer, Proteinase K and RNase A. All samples were purified using the QIAquick PCR Purification Kit. H3K27me3, Smarca4: fixation according to Active Motif proticol (formaldehyde, Glycine), snap-frozen and shipped to Active Motif.
Libraries were prepared and sequenced (~20M single read per sample) using the Next-Seq 500 sequencing platform (high-output Kit, 75 Cycles v2 Chemie) at the Genomics Core Facility (University Hospital Münster, Münster).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
S01_81K1_Input_Ctrl
|
| Data processing |
Fastq files were trimmed and prepared for further processes using TrimGalore! with automatic detection for adapter sequences, and Galaxy's default settings (Trim low-quality ends from reads in addition to adapter removal: quality score threshold 20; overlap with adapter sequence required to trim a sequence: 1; maximum allowed error rate: 0.1; discard reads that became shorter than length 20).
The resulting alignment was filtered with samtools 1.9 to include only uniquely mapped reads with less than two mismatches.
Alignement was performed using Bowtie2 using mm10 as reference genome and Galaxy's default settings.
ChIP-seq peaks were called with the algorithm MACS2 and default parameters for single samples (Histone / TF against input).
Genome_build: mm10
Supplementary_files_format_and_content: Processed data files were created with MACS2 and represent ChIP-seq peaks, including the MACS-specific score per identified peak region.
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| |
|
| Submission date |
Oct 27, 2021 |
| Last update date |
Apr 27, 2022 |
| Contact name |
Carolin Walter |
| E-mail(s) |
c_walt03@uni-muenster.de
|
| Organization name |
Westfälische Wilhelms-Universität Münster
|
| Department |
Medical Faculty of the WWU Münster
|
| Lab |
Institute of Medical Informatics
|
| Street address |
Domagkstraße 9
|
| City |
Münster |
| ZIP/Postal code |
48149 |
| Country |
Germany |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE186667 |
Smarcb1 is indispensable for the nervous system development and its loss results in a deregulation of esBAF binding [ChIP-seq] |
| GSE186669 |
Smarcb1 is indispensable for the nervous system development and its loss results in a deregulation of esBAF binding. |
|
| Relations |
| BioSample |
SAMN22601072 |
| SRA |
SRX12798067 |
