GSM1464011: ds066 293T FOXM1 GTX 1; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
FOXM1
Cell type
Cell type Class
Kidney
Cell type
293
Primary Tissue
Kidney
Tissue Diagnosis
Normal
Attributes by original data submitter
Sample
source_name
HEK293 Flp-In cells
cell type
293 Flp
antibody
anti-FOXM1 (GTX-1000276 and GTX102170)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
IP was carried out as described in Schmidt et al. (2009) (PMID 19275939). In brief, cells were cross-linked using 1% formaldehyde for 10 min at RT, reaction was quenched with 1/20th volume 2.5M Glycine and cell pellets were collected after scraping plates. Nuclear lysates were prepared by lysing with LB1 (50mM Hepes-KOH, pH 7.5, 140mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton-X plus PI) 10 min at 4°C and LB2 (10mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5M EGTA plus PI) 5 min at 4°C, followed by sonication in LB3 (10mM Tris-HCl, pH8.0, 100mM NaCl, 1mM EDTA, 0.5M EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine plus PI) using a diagnode sonicator to give a fragment size of ~200-300bp. Sonicated lysate was combined with 100ul protein-A magnetic beads preloaded with either 10 ug anti-FOXM1 antibody (GeneTex (USA); cat no. GTX-1000276 and GTX102170) or 10 μl anti-GFP antibody (Abcam, Cambridge, UK; ab290) O/N at 4°C. Beads were washed X6 in RIPA buffer (50mM HEPES pH7.6, 1mM EDTA, 0.7% Na deoxycholate, 1% NP-40, 0.5M LiCl) and DNA was eluted for 16 hr at 65°C in elution buffer (50mM Tris-HCl, pH8.0, 10mM EDTA, 1% SDS). Extracted DNA was treated with RNase A for 30 min at 37°C, followed by Proteinase K for 1 hr at 55°C, and DNA was purified by phenol-chloroform extraction and quntified by bioanalyzer and quibit. Illumina ChIP-seq kit sample prep kit was used to prepare duplicate/triplicate samples