Illumina HiSeq 2500 sequencing; Leveraging cell type-specific regulatory regions to detect SNPs associated with tissue factor pathway inhibitor plasma levels
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
No description
Antigen
NA
Cell type
Cell type Class
Cardiovascular
Cell type
Aorta endothelium
NA
NA
Attributes by original data submitter
Sample
ENA first public
2016-10-30
ENA last update
2016-07-14
ENA-CHECKLIST
ERC000011
External Id
SAMEA4079233
INSDC center alias
Program in Genetics and Genome Biology, the Hospital for Sick Children, Department of Molecular Genetics, University of Toronto
INSDC center name
Program in Genetics and Genome Biology, the Hospital for Sick Children, Department of Molecular Genetics, University of Toronto
INSDC first public
2016-10-30T17:09:23Z
INSDC last update
2016-07-14T14:45:44Z
INSDC status
public
Submitter Id
E-MTAB-4935:Input_human_rep2
broker name
ArrayExpress
cell type
endothelial cell
common name
human
ethnicity
Caucasian
individual
2366
organism part
aorta endothelium
sample name
E-MTAB-4935:Input_human_rep2
sex
male
Sequenced DNA Library
library_name
Input_human_rep2_s
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Chromatin immunoprecipitation experiments were conducted as previously described in (Schmidt et al. 2009). Antibodies used for ChIP were mouse anti-H3K27ac (Millipore, 05-1334 monoclonal) and rabbit anti-Jun (Santa Cruz Biotechnology, sc1694 polyclonal). EC were grown in supplier-recommended Endothelial Cell Growth Media (Cell Applications, catalogue #211-500) and cultured at 37° C in a 5%-CO2 humidified incubator. ChIP DNA was prepared for Illumina sequencing by blunt-end repair, dA-tailing, and ligation of Illumina adaptors using a NEBNext DNA library preparation kit (New England Biolabs, catalogue #E6040L). Total ChIP DNA (approximately 200-500ng) and 220ng of DNA input (WCE) was end repaired for 30 minutes at room temperature, and then purified using column purification with either DNA Clean and Concentrator (Zymogen, catalogue #D4014) or PCR purification columns (Qiagen, catalogue #28106) as recommended by manufacturer’s protocol. Blunt-end repaired DNA was dA-tailed for 40 minutes at 37° C, then column purified. dA-tailed DNA was ligated to Illumina adaptors (final concentration 6.67 nM) that have a T- overhang. USER enzyme was used to cleave the uracil hairpin of the Illumina adaptor, and adaptor-ligated DNA was column purified. The library was PCR amplified for 16-18 cycles using a universal primer and a barcoded primer (New England Biolabs, catalogue #E7335L). PCR-amplified DNA was purified using PCR column purification and eluted in 20 uL of elution buffer for preparation of gel extraction, or 30 uL of TE for preparation of Pippin Prep size selection. Library samples were size selected from 200-350bp using a 2% agarose dye-free automated size selection cassette from Pippin Prep (Sage Sciences, catalogue #CDF2010).