Dataset features

Specifications


Application: ChIP-seq analysis
Number of samples: 20
Release date: Feb 13 2017
Last update date: Jun 21 2017
Access: Public
Computational protocol: ELAND, BaseSpace, ComBat, DAVID, GEO
Dataset link Histone modification (H3K4me3 and H3K27me3) during vascular endothelial cell differentiation from mouse embryonic stem cells

Experimental Protocol


We prepared Flk-positive mesoderm cells, and 6 and 48 hours after with or without VEGF treatment cells. Cells were collected and cross-linked with 1% formaldehyde for 10min. After neutralization by using 0.2M glycine for 5min, cells were collected, re-suspended in SDS lysis buffer (10 mM Tris-HCl, 150 mM NaCl, 1% SDS, 1mM EDTA; pH 8.0, protease inhibitor cocktail), and fragmented by sonication (Sonifer 250, Branson; 10min, 60% duty, output level 4). The sonicated solution was diluted in ChIP dilution buffer (20 mM Tris-HCl (pH 8.0), 150 mM NaCl, 1 mM EDTA, 1% Triton X-100) to a volume of 10.3ml, and 10ml was used for IP,whereas 300 ul was used as INPUT. Specific antibodies were bound with Dynabeads Magnetic beads (Life technologies, Madison, WI, USA) and applied to the diluted sonicated solution for immunoprecipitation. Antibodies against H3K4me3 (kindly gifted by Dr. Kimura (Tokyo Institute of Technology, Japan)) and H3K27me3 (07-449, Millipore, Billerica, MA, USA) were used. The prepared DNA was quantified using Q-bit (Life Technologies) and more than 10 ng of DNA was processed for the ChIP-seq assay. ChIP-DNA was prepared for sequencing according to a modified version of the genomic DNA protocol (Illumina, San Diego, CA, USA).

Repositories


GEO

GSE94828

ENA

SRP099437

BioProject

PRJNA374538

Download


Contact


Yasuharu Kanki

Dataset Statistics

info

Citations per year

Dataset publication