GSM3610515: ChIP-seq GR MUT DexLPS rep1; Mus musculus; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
Nr3c1
Cell type
Cell type Class
Embryonic fibroblast
Cell type
MEF
Tissue
Embryonic Fibroblast
Lineage
primaryCells
Description
Mouse Embryonic Fibroblast
Attributes by original data submitter
Sample
source_name
mouse embryonic fibroblasts (MEFs)
strain
C57BL/6
antibody
Glucocorticoid Receptor - Proteintech 24050-1-AP
Sex
male
genotype
GR Zn / Zn
treatment
16h 1 uM Dex in EtOH + 3 100ng/ul LPS in PBS
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
ChIP was performed as previously described (Uhlenhaut et al., 2013) with the addition of a chromatin pre-clearing step. Briefly, cells were fixed with 30min DSG and 10min 1% formaldehyde. Crosslink was quenced with 0.2M glycine for 5min. Cells were lysed with Fast IP buffer (150mM NaCl, 5mM EDTA, 5mM Tris, 1% Triton X-100 and 0.5% NP40). Chromatin was sheared in shearing buffer (1% SDS, 10µM EDTA and 0.05% Tris) until majority of chromatin fragments were between 200-500bp. Chromatin pre-clearing was done by incubation with Dynabeads at 4°C rotating before IP set-up overnight. Next day, Dynabeads were added to IPs and 6h incubated at 4°C rotating. IPs were washed 6 times with Fast IP buffer. DNA was eluted with ChIP elution buffer (100mM NaHCO3 and 1%SDS). Library preperation was performed from 2 ng of ChIP DNA using the Kapa Hyper Prep Kit with PCR library amplification (#KK8504, KapaBiosystems) according to the manufactur's protocol. Shortly, ChIP DNA was end-repaired and A-tailed before adapter ligation. After adapter ligation, the ChIP DNA was purified with Agencourt AMPure XP beads (#A63880, Beckman Coulter) and size-selected for 360-600 bp using the Pippin Prep System from Saga Science. The size-selected library is amplified by PCR using the Kapa Hyper Prep Kit (#KK8504,KapaBiosystems) and purified terminally with Agencourt AMPure XP beads (#A63880, Beckman Coulter).