|
| Status |
Public on Apr 29, 2021 |
| Title |
A673_siSA2-Dh6_72h_STAG1_spike_R1 |
| Sample type |
SRA |
| |
|
| Source name |
Ewing sarcoma cell line (A673)
|
| Organism |
Homo sapiens |
| Characteristics |
sirna: siSA2-Dh6
antibody: STAG1
timepoint: 72h
|
| Treatment protocol |
Cells were transfected for 72h with either siCT, siSA2#6 or siSA6#8 (Dharmacon). For each siRNA, we performed ChIP-seq for CTCF, cohesin members (RAD21, SMC1A, STAG1 and STAG2), EWS-FLI1 and H3K27ac. Input of A673 WT was used as normalization controls. ChIP were performed following manufacturer instructions using iDeal ChIP-seq kit for transcription factors (ref C01010170, Diagenode) and by adding Spike-in Chromatin (ref 53083, Active motif) and Spike-in Antibody (ref 61686, Active motif) to normalize the experiments. Ewing cell lines were fixed for 10 minutes with 1% of methanol-free formaldehyde (ref 28908, Thermo-Scientific). Chromatin was sonicated (Bioruptor Pico, Diagenode) for 10 minutes (30-sec on, 30-sec off) to generate Chromatin fragments with an average size around 150-300pb.
|
| Growth protocol |
A673 were cultured in DMEM supplemented with 10% fetal bovine serum. Four days before performing the ChIP experiments, cells were seeded in T150 flask at a concentration allowing to reach a cellular confluence of approximately 90% at the time of fixation of the cells with formaldehyde.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The library were constructed using TruSeq ChIP library preparation kit (Illumina) according to manufacturer recommendations
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Reads were mapped with bowtie2 against human genome (hg19)
Peaks were called with MACS2 2.1.1 with the option narrow for FLI1, CTCF and cohesin members ChIP-seq and broad for histone marks. For each cell line, ChIP-seq were normalized according to the input DNA sample.
Genome_build: hg19
Supplementary_files_format_and_content: peaks
|
| |
|
| Submission date |
Aug 21, 2020 |
| Last update date |
Apr 29, 2021 |
| Contact name |
Olivier Delattre |
| Organization name |
Institut Curie
|
| Department |
U830
|
| Lab |
Olivier Delattre
|
| Street address |
26 rue d'ulm
|
| City |
Paris |
| ZIP/Postal code |
75005 |
| Country |
France |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE133228 |
STAG2 promotes CTCF-anchored loop extrusion and cis-promoter and -enhancer interactions |
| GSE156649 |
ChIP-seq data in A673 Ewing cell line (siCT & siSA2) |
|
| Relations |
| BioSample |
SAMN15878102 |
| SRA |
SRX8984320 |
