GSM4648990: shPRMT1 AR-IP Rep1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
AR
Cell type
Cell type Class
Prostate
Cell type
LNCAP
Primary Tissue
Prostate
Tissue Diagnosis
Carcinoma
Attributes by original data submitter
Sample
source_name
LNCaP cells
treatment
shPRMT1
antibody
AR, Abcam, ab74272
cell line
LNCaP
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_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).