 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jan 28, 2022 |
| Title |
Furamidine_AR-INPUT_Rep2 |
| Sample type |
SRA |
| |
|
| Source name |
LNCaP cells
|
| Organism |
Homo sapiens |
| Characteristics |
treatment: Furamidine
antibody: NA
cell line: LNCaP
|
| Treatment protocol |
For samples with PRMT1 knockdown, LNCaP cells were lentivirally transduced with pLKO.5 encoding an shRNA targeting either LacZ or PRMT1 and selected on puromycin. For samples with small-molecule PRMT1 inhibition, LNCaP cells were treated with DMSO or furamidine (8 μM).
|
| Growth protocol |
LNCaP cells were grown in RPMI supplemented with 10% FBS, 100 U/mL penicillin, 100 ug/mL streptomycin, 2 mM L-glutamine, and 100 ug/mL Normocin (Invivogen).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
10 million cells were fixed using 1% formaldehyde (Thermo Fisher Scientific, BP531-25) for 10 minutes at room temperature followed by quenching with 125 mM glycine (Sigma-Aldrich, #50046). Cells were rinsed twice with PBS and resuspended in 1 mL lysis buffer (1X PBS, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS) supplemented with protease inhibitor cocktail (Roche, #11836170001). Chromatin was sheared to 200-500 base pairs using a Covaris E220 sonicator and cleared by centrifugation for 15 minutes at 19,000 x g. Antibodies (AR, 9 μg, Abcam, ab74272; H3K27ac, 1 μg, Diagenode, C15410196) were incubated with 40 μL of protein A/G Dynabeads (Thermo Fisher Scientific, #10002D, #10003D) for at least 6 hours at 4°C before overnight incubation at 4°C with sonicated chromatin. Chromatin-bead complexes were washed 5 times with 1 mL LiCl wash buffer (100 mM Tris pH 7.5, 500 mM LiCl, 1% NP-40, 1% sodium deoxycholate) and rinsed twice with 1 mL TE buffer (10 mM Tris pH 7.5, 0.1 mM EDTA). Immunoprecipitated chromatin was resuspended in 100 μL elution buffer (100 mM NaHCO3, 1% SDS) and treated with RNase A (Thermo Fisher Scientific, #12091021) for 30 minutes at 37°C. Crosslinks were reversed in the presence of proteinase K (Thermo Fisher Scientific, #25530049) for 16 hours at 65°C, and the eluted DNA was purified using a MinElute PCR Purification Kit (QIAGEN, #28006).
ChIP-seq libraries were prepared using the NEBNext Ultra II DNA Library Preparation Kit (NEB, #E7645).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
ChIP-seq data were processed with the ChiLin pipeline (code version 2.0, db version 1.0) in simple mode using bwa to align to the hg38 human reference genome and MACS2 to call peaks, using the ‘narrow’ setting. For each condition, two replicate IP samples were processed along with their corresponding input DNA controls.
Genome_build: hg38
Supplementary_files_format_and_content: BigWig files generated for each replicate using ChiLin pipeline.
|
| |
|
| Submission date |
Jul 01, 2020 |
| Last update date |
Jan 29, 2022 |
| Contact name |
Peter Choi |
| E-mail(s) |
choip@chop.edu
|
| Organization name |
The Children's Hospital of Philadelphia
|
| Department |
Pathology and Laboratory Medicine
|
| Lab |
Choi Lab
|
| Street address |
3501 Civic Center Blvd, CTRB 4400
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
|
| Relations |
| BioSample |
SAMN15419867 |
| SRA |
SRX8651219 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4648973_Furamidine_AR-INPUT_Rep2.bw |
875.1 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
