|
| Status |
Public on May 02, 2014 |
| Title |
ECFC_Mock_ChIP |
| Sample type |
SRA |
| |
|
| Source name |
Human primary endothelial colony-forming cells
|
| Organism |
Homo sapiens |
| Characteristics |
source: ECFCs derived from human umbilical cord blood
antibody: isotype IgG control
|
| Growth protocol |
Human umbilical cord blood from normal full-term deliveries were obtained after informed consent. Light-density mononuclear cell (MNC) fractions were isolated by Ficoll density and resuspended in complete EGM-2 medium (EBM-2 Endothelial Cell basal Medium-2, Lonza, Cat# CC-3156, supplemented with EGM-2 SingleQuot Kit Supplements & Growth Factors, Lonza, Cat# CC-4176) and 10% fetal bovine serum and plated on to CellBIND 6 well plates at 1x107 cells/well. Individual ECFC colonies were isolated with 0.05% trypsin and expanded in EGM-2 medium. ECFCs at passage 3 were used.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Binding of TAL1 transcription factor was measured using a ChIP protocol as previously described (Palii, 2011). Chromatin from 0.2x10^8 cells was fragmented to a size range of 100-300 bases with a Bioruptor (Diagenode) at 4°C. Solubilized chromatin was immunoprecipitated with an antibody against TAL1 (C-21, Santa-Cruz) or an isotype IgG control. Antibody-chromatin complexes were pulled-down using Dynabeads-Protein G, washed and eluted. After cross-link reversal and proteinase K treatment, immunoprecipitated DNA was extracted with phenol-chloroform.
ChIPed DNA was amplified using the Illumina protocol with modifications described in (Palii, 2011).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Mock ChIP in ECFC cells
|
| Data processing |
Basecalling was performed with Illumina RTA 1.12.4.2
ChIP-Seq reads were aligned to hg19 genome assembly using MAQ 0.7.1 allowing a maximum of two mistmatches. Unmapped reads were truncated to 31 bp and remapped to the genome. Truncation and remapping were repeated down to 26 read size.
Reads mapping to more than two locations on the genome were removed.
Reads mapping only to one location on the genome were used to call peaks with MACS 1.3.7.1 using a minimum cutoff of 8 tags per enriched region with a p-value threshold for peak significant under 1x10^-5.
Genome_build: hg19
Supplementary_files_format_and_content: bedGraph files represent the read coverage of the ChIP-Seq over the genome. The files were generated from unique reads without duplicates from the alignment with MAQ using the UCSC genome browser standalone tools for Unix
|
| |
|
| Submission date |
Feb 20, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Marjorie Brand |
| E-mail(s) |
karthi.sivaraman@gmail.com, mbrand@ohri.ca
|
| Organization name |
Ottawa Hospital Research Institute
|
| Street address |
501 Smyth Road
|
| City |
Ottawa |
| State/province |
Ontario |
| ZIP/Postal code |
K1H 8L6 |
| Country |
Canada |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE44442 |
Genomic binding profile of TAL1 in human Endothelial Colony-Forming Cells |
| GSE44546 |
TAL1 in human Endothelial Colony-Forming Cells |
|
| Relations |
| SRA |
SRX244127 |
| BioSample |
SAMN01924255 |
