|
| Status |
Public on Sep 06, 2022 |
| Title |
ATAC-LHCN-Ctrl-2 |
| Sample type |
SRA |
| |
|
| Source name |
cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: LHCN
treatment: LHCN overexpress empty vector
|
| Treatment protocol |
LHCN cells were overexpressed PAX3-FOXO1 or empty vector for 72 hours. RH30 cells were treated with QC6352 200nm or 0.1% DMSO or transduced with shRNA control or KDM4B for 72 hours.
|
| Growth protocol |
LHCN cells were cultured in medium with four parts Dulbecco's modified Eagle's medium to one part medium 199 supplemented with 15% fetal bovine serum, 0.02 M HEPES, Basic FGF 10 ng/ml , 2.5ng/ml HGF, 0.055 µg/ml dexamethasone , 0.03 µg/mlzinc sulphate (ZnSO4) and 1.4 µg/mlvitamin B12 , 60 U/ml penicillin/streptomycin on dishes coated with 0.1% porcine skin gelatin. RH30 cells were cultured with RPMI 1640 media supplemented with 100 U/mlpenicillin/streptomycin and 10% fetal bovine serum.All cells were maintained at 37 °C in an atmosphere of 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Fresh cultured LHCN or RH30 cells (100,000 per sample) with or without various treatments were harvested and washed with 150ml cold Dulbecco's Phosphate-Buffered Saline (DPBS) containing protease inhibitor (PI). Nuclei were collected by centrifuging at 500g for 10 minutes at 4°C after cell pellets were resuspended in lysis buffer (10 mM Tris-Cl pH 7.4, 10 mM NaCl, and 3 mM MgCl2 containing 0.1% NP-40 and PI. Nuclei were incubated with Tn5 transposon enzyme in transposase reaction mix buffer (Illumina) for 30 min at 37°C. DNAs were purified from transposition sample by using Min-Elute PCR purification kit (Qiagen, Valencia, CA) and measured by Qubit.
Polymerase chain reaction (PCR) was performed to amplify with High-Fidelity 2X PCR Master Mix [72°C/5mins+ 98 °C /30 s +12 × (98 °C /10 s + 63 °C /30 s + 72 °C /60 s) + 72 °C /5 min]. The libraries were purified using Min-Elute PCR purification kit (Qiagen, Valencia, CA). ATAC-seq libraries followed by pair-end sequencing on HiSeq4000 (Illumina) in the Hartwell Center at St Jude Children's Research Hospital.
|
| |
|
| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
The ATAC-seq raw reads were aligned to the human reference genome (hg19) using BWA (version 0.7.12-r1039) to and then marked duplicated reads with Picard (version 1.65), with only high-quality reads kept by samtools (version 1.3.1, parameter ‘‘-q 1 -F 1024’’). Reads mapping to mitochondrial DNA were excluded from the analysis. All mapped reads were offset by +4 bp for the + strand and -5 bp for the – strand
Subsample 150 million of reads per sample from Bam file with Samtools for peak calling. Peaks were called for each sample using MACS2 (version 2.1.1 20160309) with parameters “-q 0.01 --nomodel --extsize 200 –shift 100 ".
Genome_build: hg19
Supplementary_files_format_and_content: narrowPeak
|
| |
|
| Submission date |
Aug 28, 2020 |
| Last update date |
Sep 06, 2022 |
| Contact name |
Hongjian Jin |
| E-mail(s) |
hongjian.jin@STJUDE.ORG
|
| Organization name |
St Jude Children's Research Hospital
|
| Department |
Center for Applied Bioinformatics
|
| Street address |
262 Danny Thomas Place
|
| City |
Memphis |
| State/province |
TN |
| ZIP/Postal code |
38015 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE157092 |
Targeting KDM4B to disrupt the core regulatory transcription network governed by PAX3-FOXO1 in high-risk rhabdomyosarcoma [ATAC-seq] |
| GSE157095 |
Targeting KDM4B to disrupt the core regulatory transcription network governed by PAX3-FOXO1 in high-risk rhabdomyosarcoma |
|
| Relations |
| BioSample |
SAMN15934406 |
| SRA |
SRX9031577 |
