|
| Status |
Public on Dec 04, 2009 |
| Title |
Tbx3_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
Embyonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
cell type: pluripotent and self-renewing cells
|
| Treatment protocol |
No treatment was performed for ESCs used for ChIP-sequencing.
|
| Growth protocol |
D3 and R1 mouse ESCs were obtained from ATCC and cultured as per manufacturer's recommendation, in the presence of inactivated MEFs supplemented with LIF.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with Tbx3 antibody. Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
Chromatin IP against Tbx3
|
| Data processing |
Alignment: Sequence reads were obtained and mapped to the mouse (mm8) genome using the Illumina Genome Analyzer II Pipeline. All reads mapping with two or fewer mismatches were retained.
Peaks: Peak detection was performed with the Model-based Analysis of ChIP-Seq (MACS) algorithm (http://liulab.dfci.harvard.edu/MACS/) using processed data file. Control input DNA sequencing library was used as background for peak calling.
|
| |
|
| Submission date |
Nov 30, 2009 |
| Last update date |
May 15, 2019 |
| Contact name |
Yuriy L Orlov |
| E-mail(s) |
orlovy@gis.a-star.edu.sg
|
| Organization name |
Genome Institute of Singapore
|
| Department |
Computational Systems Biology
|
| Street address |
60 Biopolis Street
|
| City |
Singapore |
| ZIP/Postal code |
138672 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL9250 |
| Series (1) |
| GSE19219 |
Genome-wide maps of Tbx3 binding sites in mouse ESCs |
|
| Relations |
| SRA |
SRX015118 |
| BioSample |
SAMN00007202 |
