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| Status |
Public on Apr 21, 2022 |
| Title |
Sample 1_HeLa-WT |
| Sample type |
SRA |
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| Source name |
HeLa-WT
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| Organism |
Homo sapiens |
| Characteristics |
cell line background: HeLa
cell type: Epithelial cells
disease: Adenocarcinoma
chip antibody: Mouse anti-Flag (clone M2, Sigma-Aldrich, Cat# F1804)
treatment: Treated with 50 μM sodium arsenite for 6h
sirna treatment: Untreated
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| Treatment protocol |
Cells were left untreated or treated with 100 nM of either siRNA-Control or siRNA-FACT for 72h. Then, cells were incubated with 50 μM sodium arsenite for additional 6h.
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| Growth protocol |
HeLa cells (CCL-2, ATCC) and HeLa cells stably expressing TFEB-Flag were grown in DMEM, high glucose, GlutaMAX, sodium pyruvate (Gibco, 10569044) supplemented with 10% fetal bovine serum (Invitrogen, 21041-025), 100 U/ml penicillin and 100 μg/ml streptomycin (Gibco, 2114).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin was isolated from cells by adding lysis buffer (1% SDS, 10 mM EDTA and 50mM Tris-HCl, pH 8.1 containing protease inhibitors), followed by disruption with a Dounce homogenizer. Lysates were sonicated and the DNA sheared to an average length of 300-500 bp with Active Motif’s EpiShear probe sonicator (cat# 53051). Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by SPRI beads clean up (Beckman Coulter) and quantitation by Clariostar (BMG Labtech). Genomic DNA regions of interest were isolated using monoclonal antibody against Flag.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. Steps were performed on an automated system (Apollo 342, Wafergen Biosystems/Takara) following the manufacturer's protocol. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on Illumina’s NextSeq 500 (75 nt reads, single end) following the manufacturer's protocols.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
1_08RK_015NNIH_HeLa-WT_FLAG_hs_i61
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| Data processing |
Basecalls performed using Illumina bcl2fastq2 version 2.20
ChIP-seq reads were aligned to the human GRCh38/hg38 genome assembly using Burrows-Wheeler Aligner algorithm with default settings.
Data were filtered as follows: Duplicate reads were removed, and only uniquely mapped reads (mapping quality >= 25) were used for further analysis. Alignments were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library and assigned to 32-nt bins along the genome. The resulting histograms (genomic “signal maps”) were stored in bigWig files.
Peak locations were determined using the MACS algorithm (v2.1.0) with a cutoff of p-value = 1e-7 [2]. Peaks that were on the ENCODE blacklist of known false ChIP-Seq peaks were removed.
Genome_build: hg38
Supplementary_files_format_and_content: bigWIG files were generated using wigToBigWig version 4 software
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| Submission date |
Jul 18, 2021 |
| Last update date |
Apr 21, 2022 |
| Contact name |
Jose Martina |
| E-mail(s) |
[email protected]
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| Organization name |
NIH
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| Department |
NHLBI, CDBC
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| Lab |
LCB
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| Street address |
50 South Dr., Building 50, Room 3533
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| City |
Bethesda |
| State/province |
Maryland |
| ZIP/Postal code |
20892-8018 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE180322 |
The FACT complex facilitates expression of lysosomal and antioxidant genes through binding to TFEB and TFE3 [ChIP-seq] |
| GSE180324 |
The FACT complex facilitates expression of lysosomal and antioxidant genes through binding to TFEB and TFE3. |
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| Relations |
| BioSample |
SAMN20296632 |
| SRA |
SRX11490895 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5460806_1_08RK_015NNIH_HeLa-WT_FLAG_hg38_i61_uniqnorm_signal.bw |
135.4 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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