|
| Status |
Public on Apr 08, 2022 |
| Title |
E47_vpGFP_rep1 |
| Sample type |
SRA |
| |
|
| Source name |
K562
|
| Organism |
Homo sapiens |
| Characteristics |
genotype: E47
knock down: NA
hemin treatment: no_Hemin
facs: 2xGFP_positive
viewpoint: GFP
barcode: GGGGCACAAGCTGGAGTA
|
| Treatment protocol |
For differentiation, culture medium was supplemented with porcine hemin (Sigma, 30 µM final concentration) one day after sorting. Hemin and medium were refreshed one day later.
|
| Growth protocol |
Human erythroleukemia K562 cells were grown at 37°C at 5% CO2 in RPMI 1640 (Gibco) with 10% FBS (Sigma) and 1% Penicillin-Streptomycin (Gibco). Density was kept between 1x105 and 5x105 cells per ml medium. Cells were routinely tested for mycoplasma. Prior to every experiment, cells were weekly bulk sorted for two consecutive weeks on Becton Dickinson SORP FACSAria FUSION Flow Cytometer. To obtain irreversibly silenced populations of cells, GFP negative cells where weekly sorted GFP-negative for six (E100) or nine (E407) consecutive weeks.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
4C experiments were performed as described (Krijger et al., 2020). Eight to ten million cells were cross-linked with formaldehyde. DNA was digested in situ with Csp6I (first cutter) and NlaIII (second cutter).
Indexed Illumina sequencing adapters were introduced to ligation fragments of interest with a two-step PCR strategy as described in Krijger et al., 2020.
OTHER: 4C
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Reads were mapped and processed using pipe4C (Krijger et al., 2020) (github.com/deLaatLab/pipe4C) with the following parameters: normalization to 1 million reads in cis, non-blind fragments only, window size 21, top 2 read counts removed
Genome_build: Modified versions of hg19 containing the sequences of the integrated regulatory elements at their respective integration site.
Supplementary_files_format_and_content: 4C: interaction coverage file (wig files generated using pipe4C)
|
| |
|
| Submission date |
Jul 21, 2021 |
| Last update date |
Apr 08, 2022 |
| Contact name |
Peter Krijger |
| E-mail(s) |
[email protected]
|
| Organization name |
Hubrecht Institute
|
| Lab |
Wouter de Laat
|
| Street address |
Uppsalalaan 8
|
| City |
Utrecht |
| ZIP/Postal code |
3584CT |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE180566 |
Building regulatory landscapes reveals that an enhancer can recruit cohesin to create contact domains, engage CTCF sites and activate distant genes |
|
| Relations |
| BioSample |
SAMN20342559 |
| SRA |
SRX11515275 |
