GSM3488958: Jurkat.BHLHE40 Rep.2; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
Epitope tags
Cell type
Cell type Class
Blood
Cell type
Jurkat
Primary Tissue
Blood
Tissue Diagnosis
Leukemia Acute Lymphocytic
Attributes by original data submitter
Sample
source_name
Jurkat T cell line
cell type
Jurkat T lymphocytes
label
GFP
antibody
Sigma-Aldrich F1804
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
ChIP lysates were generated from 6x10^7 cells. Cells were fixed in 1% formaldehyde for 10min. Lysate was sonicated and then immunoprecipitated with 10ug of antibody against FLAG. Antibodies were pre-bound overnight to 100ul of G protein-coupled paramagnetic beads (Dynabeads) in PBS/BSA 0.5%. Beads were then added to lysates and incubation was let to proceed overnight. Beads were washed 6 times in a modified RIPA buffer (50mM Hepes pH 7.6, 500mM LiCl, 1mM EDTA, 1% NP-40, 0.7% Na-deoxycholate) and once in TE containing 50mM NaCl. DNA was eluted in TE containing 2% SDS and crosslinks reversed by incubation overnight at 65ºC. DNA was then purified by QIAquick columns (Qiagen) and quantified using PicoGreen (Invitrogen). ChIP DNA was prepared for NextSeq 500 Illumina sequencing using a standard protocol consisting in blunting, addition of dA overhangs, ligation of Illumina adapters, PCR with index primers and purification. A mixture of T4 DNA polymerase, DNA polymerase I and T4 kinase was used according to manufacturer's instruction. Library preparation is carried out on SPRIworks Fragment Library System. The purified DNA libraries were quantified both with a TapeStation 4200 (Agilent Technologies) and Qubit (LifeTechnologies) and diluted to a working concentration of 2 nM.