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| Status |
Public on Apr 15, 2024 |
| Title |
TC-32 siNEG, H3K27ac, rep 1a |
| Sample type |
SRA |
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| Source name |
TC-32
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| Organism |
Homo sapiens |
| Characteristics |
cell line: TC-32
cell type: Ewing sarcoma
chip antibody: H3K27ac (ab4729, #39133)
sirna: siNeg
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| Treatment protocol |
RNAi-based studies, cells were reverse transfected with 20 nM siRNA complexed with Lipofectamine RNAi-Max (Invitrogen). siNeg (SI03650318, Qiagen) and siFLI1 (s5266, Thermo Fisher Scientific).
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| Growth protocol |
Cells were cultured in RPM1 1640 supplemented with 10% FBS and Plasmocin Prophylactic (Invivogen, San Diego, CA), and grown at 37°C, 5% CO2. We confirmed the identity of cell lines using short tandem repeat (STR) analysis (ATCC) at intervals throughout experimentation, and we monitored for mycoplasma contamination using the MycoAlert Plus system (Lonza, Walkersville, MD).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP assays were performed using SimpleChIP plus Enzymatic Chromatin IP kit (9005S; Cell Signaling Technology, Danvers, MA). Briefly, cells were crosslinked with final concentration of 1% formaldehyde for 10 minutes. After quenching, cell lysis and enyzmatically digested, chromatin was sonicated using Branson SFX250 Digital Sonifier (Branson Ultrasonics) to obtain a fragment size of 200 - 900 bp. Solubilized chromatin was immunoprecipitated with the indicated antibodies and Protein G-Dynabeads (Thermo Fisher Scientific).
ChIP-seq libraries were using SMARTer ThruPlex TAKARA Library Prep kit following manufacturer's instructions. 10 ng of DNA was used as starting material for input and IP samples. Libraries were amplified using 8 cycles on the thermocycler.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
NextSeq 2000 |
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| Data processing |
ChIP-seq analysis were carried out using center for cancer research bioinformatics resource (CCBR) pipeline (https://github.com/CCBR/Pipeliner/wiki/Pipeline-Documentation - chip-seq).
Sequence reads were trimmed for adaptor sequence using cutadapt v4.4.
Trimmed reads were aligned to GRCh38/hg38 human genes and Drosophila melanogaster dm6 genes using BWA mem v.0.7.17.
PCR duplicates were removed with the Picard v.2.17.11 SamToFasq (for blacklist read removal) and MarkDuplicates (to remove PCR duplicates).
Peak calling was performed against input control using model-based MACS2 v2.1.1, with the cut-off q-value < 0.02.
Assembly: GRCh38/hg38
Supplementary files format and content: narrowPeak (except for Input sample)
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| Submission date |
Dec 18, 2023 |
| Last update date |
Apr 15, 2024 |
| Contact name |
Natasha J Caplen |
| Organization name |
National Institutes of Health
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| Department |
National Cancer Institute
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| Lab |
Functional Genetics
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| Street address |
37 Convent Drive, Bldg. 37, Rm 6128
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| City |
Bethesda |
| State/province |
Maryland |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL30173 |
| Series (2) |
| GSE243171 |
EWSR1::FLI1 fusion oncoproteins’ regulation of ETS1 (ChIP-seq) |
| GSE243184 |
ETS1, a target gene of the EWSR1::FLI1 fusion oncoprotein, regulates the expression of the focal adhesion protein TENSIN3 |
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| Relations |
| BioSample |
SAMN38913963 |
| SRA |
SRX22955359 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7980901_TC32_WT_siNEG_H3K27ac_1_S6_peaks.narrowPeak.gz |
717.4 Kb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
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