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| Status |
Public on May 21, 2011 |
| Title |
hESC FOXH1 ChIP-Seq pool |
| Sample type |
SRA |
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| Source name |
human embryonic stem cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: H9
cell type: embryonic stem cells
passages: 40-50
chip antibody: FOXH1, sheep polyclonal
chip antibody manufacturer: R&D Systems
chip antibody catalog #: AF4248
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| Growth protocol |
First 9 samples: Undifferentiated H9 hESCs were maintained on mouse embryonic fibroblast (MEF) feeder layers or on Matrigel (1:20 dilution; BD Biosciences) in mouse embryonic fibroblast-conditioned medium (CM). CM was produced by conditioning of MEFs for at least 24 hours in Dulbecco's modified Eagle's medium/Ham's F-12 medium (DMEM/F12) supplemented with 20% knockout serum replacement (Gibco), 1 mM L-glutamine, 0.1 mM nonessential amino acids, 0.1 mM 2-mercaptoethanol and 8 ng/ml recombinant human fibroblast growth factor-basic (bFGF; Peprotech). Cultures were routinely passed with 200 U/ml type IV collagenase (Gibco) in 1:4 ratio every 4â5 days.
Last 9: Derived endoderm was generated from hESCs as previously described (D'Amour et al., 2005, Nat Biotechnol 23, 1534-1541). Differentiation was performed in RPMI-1640 medium supplemented with glutamax, 100 ng/ml recombinant human Activin A (R&D Systems), penicillin/streptomycin, and defined fetal bovine serum (FBS; HyClone) at sequentially increasing concentrations (0, 0.2 to 2%). 2% FBS was maintained afterwards in cultures over the duration of differentiation.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and transcription factor-DNA or histone-DNA complexes were isolated with antibody. Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Catalog # FC-102-1001). 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 (3' to 5' 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 3â end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of 200~400 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 II following the manufacturer's protocols.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
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| Description |
Chromatin IP against FOXH1
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| Data processing |
Alignment: Sequence reads were obtained and mapped to the human genomes (hg18; March, 2006) using the Illumina Genome Analyzer Pipeline.
Peaks: Transcription factor ChIP-Seq reads were processed to call peaks using CisGenome (Ji et al., 2008, Nat Biotechnol 26, 1293-1300). The setting for peak-calling and sliding window size was 300 bp and the threshold number of reads required for peak to be called was 11 reads. The false discovery rate allowed was 0.01. Histone H3K4me3 and H3K27me3 peaks were called using QuEST 2.4 (Valouev et al., 2008, Nat Methods 5, 829-834). We used the âhistoneâ bandwidth setting with ârelaxedâ peak-calling parameters.
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| Submission date |
May 20, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Si Wan Kim |
| E-mail(s) |
swkimws@gmail.com
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| Organization name |
Stanford University
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| Department |
Genetics
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| Street address |
300 Pasteur Drive M313
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| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305-5120 |
| Country |
USA |
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| Platform ID |
GPL9115 |
| Series (1) |
| GSE29422 |
Chromatin and Transcriptional Signatures for Nodal Signaling During Endoderm Formation in hESCs |
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| Relations |
| SRA |
SRX064484 |
| BioSample |
SAMN00619220 |
| Named Annotation |
GSM727564_d0Foxh1.bed.gz |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM727564_d0Foxh1.bed.gz |
121.5 Kb |
(ftp)(http) |
BED |
| GSM727564_hESCFoxh1_pooled.txt.gz |
390.1 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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