|
| Status |
Public on May 08, 2024 |
| Title |
DKO H3K27ac rep2 |
| Sample type |
SRA |
| |
|
| Source name |
K562 cells with silencer1 and silencer2 DKO
|
| Organism |
Homo sapiens |
| Characteristics |
chip antibody: H3K27ac (ab4729, Abcam)
treatment: knockout silencer1 and 2 regions
cell line: K562
|
| Treatment protocol |
EV, S1KO and DKO cells undergoes the CRISPR excision.
|
| Growth protocol |
All the cells were cultured in RPMI-1640 supplemented with 10% Fetal Bovine Serum (FBS) and 1% penicillin-streptomycin. All cultures were maintained at 37 °C, 5% CO2 in a humidified incubator.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were fixed with 1% formaldehyde (Thermo Scientific) at room temperature for 10 min, followed by quenching with glycine for 5 min at room temperature. The fixed cell pellets were lysed in 1% SDS lysis buffer supplemented with protease inhibitor cocktail tablet (Roche), and sonicated using Bioruptor (Diagenode). The cell lysate was precleared through centrifuging in dilution buffer and incubating with Protein G Dynabeads (Invitrogen) overnight at 4 °C. The precleared lysate was added into the antibody-conjugated beads, and incubated overnight at 4 °C, with rotation. The incubated beads were then washed thrice by 0.1% SDS lysis buffer, once by high salt wash buffer, once by lithium chloride wash and once by the TE buffer. After washing, the incubated beads were added into the elution buffer with RNase A (Qiagen) followed by decrosslinking with Proteinase K (Ambion) at 37 °C overnight. ChIP DNA was extracted back by using the QIAquick PCR purification kit (Qiagen), and quantitated using Qubit High Sensitivity dsDNA Assay (Invitrogen).
ChIP-seq library construction was prepared by using ThruPLEX DNA-seq 48D Kit (Rubicon) according to the instruction.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
DKO_C1_AC_Combined_peaks.narrowPeak
|
| Data processing |
ChIP-seq reads of H3K27me3, H3K27ac from EV, S1KO and DKO cells were mapped by BOWTIE2(v2.2.5) using default parameters in pair-end mode. We filtered out alignments with mapq scores smaller than 30. The two replicates were combined and peaks and bigWig files were generated by MACS2(2.1.0.20150731) using option '-q 0.01' for H3K27ac and H3K4me3 and '--broad --broad-cutoff 0.1 -q 0.05' for H3K27me3.
Genome_build: hg19
Supplementary_files_format_and_content: *.narrowPeak
Supplementary_files_format_and_content: *.broadPeak
|
| |
|
| Submission date |
Jan 11, 2022 |
| Last update date |
May 08, 2024 |
| Contact name |
KAIJING CHEN |
| E-mail(s) |
KAIJING001@e.ntu.edu.sg
|
| Phone |
98667053
|
| Organization name |
Nanyang Technological University
|
| Department |
school of biological science
|
| Lab |
Melissa Fullwood
|
| Street address |
60 Nanyang Drive
|
| City |
Singapore |
| ZIP/Postal code |
637551 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL20301 |
| Series (2) |
| GSE193481 |
Super-silencer perturbation by EZH2 and REST inhibition leads to large loss of chromatin interactions and reduction in cancer growth [ChIP-seq] |
| GSE193489 |
Super-silencer perturbation by EZH2 and REST inhibition leads to large loss of chromatin interactions and reduction in cancer growth |
|
| Relations |
| BioSample |
SAMN24848759 |
| SRA |
SRX13731641 |
