GSM2467511: DE H3K4me3 rep3; Mus musculus; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
Histone
Antigen
H3K4me3
Cell type
Cell type Class
Pluripotent stem cell
Cell type
mESC derived endodermal cells
NA
NA
Attributes by original data submitter
Sample
source_name
Definitive Endoderm
cell type
ES_derived Definitive Endoderm
strain
129/Sv//EV
day of differentiation
Day 5
chip antibody
anti-H3K4me3 (07-473; Millipore)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Fixed cells were lyzed on ice for 20 minutes (5 mM PIPES, 85 mM KCl, 0.5% Igepal-CA 630) before pelleting and nuclear lysis (50 mM Tris-HCl, 10 mM EDTA, 1% SDS), chromatin was then sonicated to 200bp in a Covaris S220 and incubated overnight with 2 µl of anti-H3K4me3 (07-473; Millipore) and Protein A and Protein G Dynabeads (Invitrogen). Beads were washed seven times with RIPA buffer variants (10 mM Tris-HCl, 1mM EDTA, 0.5mM EGTA, 1% Triton X-100, 0.1% SDS, 0.1% Sodium Deoxycholate): RIPA (twice), High Salt RIPA (500 mM NaCl; twice), RIPA with 250mM LiCl (once) and T.E. Buffer (twice) before treatment with RNAse A at 37ºC for 1 hr (Roche) and Proteinase K 65ºC overnight (Thermo Fisher). DNA was recovered by phenol-chloroform extraction. Libraries were indexed for sequencing using NebNext Ultra II (New England BioLabs); library profiles were visualized using D1000 tape on a TapeStation (Agilent Technologies) and quantified using a universal library quantification kit (KAPA Biosystems).