|
| Status |
Public on Feb 23, 2021 |
| Title |
NG-Capture C siCTL rep2 |
| Sample type |
SRA |
| |
|
| Source name |
MDA-MB-231 cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MDA-MB-231
cell type: breast cancer cell line
tumor type: triple negative - basal b
knockdown: si control
|
| Treatment protocol |
Cells were transfected with either a control siRNA (siCTL) or a mix of siRNAs directed against Fra-1 (siFra-1) (4.5nM of siRNAs for 72 hours) using INTERFERin, according to the manufacturer's instructions. Experiments were carried out in 3 independent replicates.
|
| Growth protocol |
MDA-MB-231 cells (ATCC, Rockville, MD, USA) were cultured cultured in Dulbecco’s modified Eagle’s medium containing 10% fetal calf serum and penicillin/streptomycin (100µg/ml each) in a humidified 5% CO2 atmosphere at 37°C. Cells freshly amplified and frozen after obtention from the ATCC were used every 3months and were routinely tested for the absence of mycoplasma contamination
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The experiments were performed as previously described in (Davies et al., 2016b). Briefly, 3C libraries were generated using 900 U of the Dpn II restriction enzyme (NEB: R0543M) for 11 million cells and 720 U of the T4 DNA HC ligase (Life Technologies: EL0013). 3C libraries were sonicated (Bioruptor® Pico sonication device; 5 cycles; 30s ON/OFF). Samples were indexed to allow multiplexing using Illumina paired-end sequencing adaptors (NEB: E6040, E7335) and the Herculase II polymerase (Agilent: 600677). Oligonucleotide capture was then performed using 70-mer biotinylated DNA oligonucleotides designed at each of the studied target gene promoters (http://apps.molbiol.ox.ac.uk/CaptureC/cgi-bin/CapSequm.cgi). The hybridization reaction was performed using Nimblegen SeqCap EZ kits (Roche: 05634261001, 07145594001, 06777287001). After a 72h hybridization step, streptavidin bead pulldown (Invitrogen: 65305) was performed, followed by multiple washes (according to SeqCap EZ SR user’s guide) followed by PCR amplification of the captured material. A second capture step was performed as above.
Samples were indexed to allow multiplexing using Illumina paired-end sequencing adaptors (NEB: E6040, E7335) and the Herculase II polymerase (Agilent: 600677).
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
chimeric genomic DNA - 3C libraries
|
| Data processing |
Library strategy: NG-Capture C
Data were analyzed using scripts available at https://github.com/telenius/captureC/releases
R was used to normalize data.
Differential analysis between the siCTL and the siFra-1 condition was performed using DESeq2
Peak calling was performed using the PeakC R package (Geeven et al., 2018) using the following parameters: alphaFDR=0.1, wSize=5 and qWr=1.
Adjacent restriction fragments called by PeakC were then merged into a single region using BEDTools
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files
|
| |
|
| Submission date |
Mar 11, 2020 |
| Last update date |
Feb 23, 2021 |
| Contact name |
Isabelle Jariel-Encontre |
| E-mail(s) |
isabelle.jariel@inserm.fr
|
| Phone |
00 33 (0)4 11 28 31 53
|
| Organization name |
IRCM-U1194- INSERM
|
| Street address |
208 Avenue des Apothicaires
|
| City |
Montpellier |
| State/province |
Montpellier Cedex5 |
| ZIP/Postal code |
34298 |
| Country |
France |
| |
|
| Platform ID |
GPL16791 |
| Series (2) |
| GSE146824 |
NG-Capture C analysis of 35 gene loci regulated by Fra-1 in MDA-MB-231 cells |
| GSE146825 |
Transcriptional regulation by Fra-1 in triple negative breast cancers |
|
| Relations |
| BioSample |
SAMN14360529 |
| SRA |
SRX7897904 |
