 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on May 22, 2024 |
| Title |
JN-DSRCT, assay_1, WT1, R1, ChIP-seq |
| Sample type |
SRA |
| |
|
| Source name |
JN-DSRCT-1
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: JN-DSRCT-1
cell type: Desmoplastic Small Round Cell Tumor
treatment: NA
|
| Treatment protocol |
No treatment was performed on JN-DSRCT-1 cells for assay 1. For assay 2, JN-DSRCT-1 cell line was transfected with a custom siRNA targeting EWSR1::WT1 (3’ GAT CTT GAT CTA GGT GAG A 5’), or non-targeting siRNA (Horizon Discovery «on»-TARGETplus Non-targeting siRNA#1, reference D-001810-01-05), according to manufacturer’s instructions. After cell seeding and obtention of 50% confluency, transfection was performed using LipofectamineTM RNAiMAX (InvitrogenTM reference 13778150), and the medium was replaced the day after. A 48-hour silencing time point was used for each described experiment.
|
| Growth protocol |
JN-DSRCT-1 cell line was maintained in 2D adherent culture within DMEM/F-12 (GibcoTM) supplemented with 10% FBS, 1% Penicillin-Streptomycin (GibcoTM), 1% Sodium Pyruvate (GibcoTM), 1% Sodium Bicarbonate (GibcoTM), 1% Non-Essential Amino Acids (GibcoTM) and 1% HEPES (GibcoTM). Cell passaging was performed at 1/10 twice a week. Used cells were controlled for mycoplasma-free status. For ChIP assay, JN-DSRCT-1 cells were grown to ~80% confluence in 15 cm dishes.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin was crosslinked for 15 minutes at room temperature (RT) by adding methanol-free formaldehyde (1% final) in the dishes culture media. Then, formaldehyde was quenched by adding a glycine solution, and cells were scraped and centrifuged before freezing for subsequent utilization. After thawing on ince, cells were resuspended in FL buffer before nuclei extraction according to the NEXSON protocol (Arrigoni et al., Nucleic Acids Res, 2016). Immunoprecipitation was performed on sheared chromatin using H3K9ac, H3K27ac, WT1 C-terminal (GTX15249), or rabbit isotype antibodies overnight and immune complexes were captured the next day using Dynabeads Protein G beads. Crosslink was reverted with Proteinase K and DNA was purified on Qiagen PCR purification columns.
NebNextUltra II Library prep kit for DNA
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
ChIP-seq data analyses were performed according to nf-core/chipseq pipeline
Raw reads quality control (QC) was performed using FastQC
Adapter trimming was done with Trim Galore!
Reads were further mapped to the reference genome (GRCh38) using BWA
Duplicate reads were discarded using picard
BigWig files were generated using BEDTools to allow IGV visualization of fragments
Assembly: GRCh38
Supplementary files format and content: BigWig files
|
| |
|
| Submission date |
May 18, 2024 |
| Last update date |
May 22, 2024 |
| Contact name |
Clémence Henon |
| E-mail(s) |
clemence.henon@gustaveroussy.fr
|
| Organization name |
Gustave Roussy
|
| Street address |
114 rue Edouard Vaillant
|
| City |
Villejuif |
| ZIP/Postal code |
94800 |
| Country |
France |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE263523 |
Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors |
| GSE267803 |
Single-cell multiomics profiling reveals heterogeneous transcriptional programs and microenvironment in Desmoplastic Small Round Cell Tumors [ChIP-seq] |
|
| Relations |
| BioSample |
SAMN41447729 |
| SRA |
SRX24600815 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8279064_JN1_WT1_R1.bigWig |
472.2 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
|
|
|
|
 |
