Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
VDR

Cell type

Cell type Class
Prostate
Cell type
RWPE-1
Primary Tissue
Prostate
Tissue Diagnosis
Normal

Attributes by original data submitter

Sample

source_name
RWPE1 cells
cell line
RWPE1
cell type
HPV-18 immortalized, non-tumorigenic prostate epithelial cell line
agent
vehicle
chip antibody
anti-VDR polyclonal antibody (C-20, sc-1008, Santa Cruz Biotechnology)
number of reads
9,001,958

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Immune complexes from immunoprecipitations of formalin-fixed, sonicated samples were collected with a 50% slurry of protein G for 1 h at 4oC. Cross-links were reversed overnight in 1% SDS, 0.1 M NaHCO3, and 0.2 M NaCl at 65oC. DNA was purified using Qiagen QIAquick Spin Kits. We used methods originally reported in Robertson G. et al. (2007) Nature Methods 4:651-7. Briefly, 100-300 bp fragments of the immunoprecipitated were eluted from 12% PAGE gels and the DNA was recovered using a QIAquick PCR purification kit. DNA ends were repaired using a 1:5 mix of T4 and Klenow DNA polymerases, phenol–chloroform–isoamyl alcohol (pH 8.0) extracted, and precipitated with 100% EtOH, mussel glycogen (Invitrogen) and 7.5 M NH4OAc. We recovered the precipitate by centrifugation at 20,200g for 15 min at 4 °C. We added a single adenine base to the DNA using Klenow exo– (3′ and 5′ exo minus; Illumina), phenol–chloroform–isoamyl alcohol extracted, and precipitated as above.. The DNA pellet was washed twice with 70% EtOH and resupended in 10 ml of LoTE buffer. Adapters (Illumina) were ligated to ends of the single adenine–tailed DNA following the manufacturer's instructions with the exception that we diluted the adaptor oligonucleotide mix (Illumina) 1/10 before use. We recovered the DNA using a QIAquick PCR Purification Kit (Qiagen) and we enriched the adaptor-modified DNA fragments by PCR using Phusion polymerase (Finnzymes) and PCR primer 1.1 and 2.1 (Illumina) following the manufacturer's instructions (separate 10- and 15-cycle reactions were run). After cycling, we purified the reactions using a QIAquick MiniElute kit (Qiagen) according to the manufacturer's instructions, with 15 μl of elution buffer heated to 50 °C. We quantified the DNA quality with an Agilent DNA 1000 series II assay and a Nanodrop 7500 spectrophotometer using a 1.5-μl aliquot that was diluted to 10 nM.

Sequencing Platform

instrument_model
Illumina Genome Analyzer II

hg19

Number of total reads
9001958
Reads aligned (%)
59.7
Duplicates removed (%)
8.8
Number of peaks
338 (qval < 1E-05)

hg38

Number of total reads
9001958
Reads aligned (%)
60.9
Duplicates removed (%)
8.1
Number of peaks
74 (qval < 1E-05)

Base call quality data from DBCLS SRA