|
| Status |
Public on Oct 05, 2010 |
| Title |
m_Escell_H3K4me3_M |
| Sample type |
SRA |
| |
|
| Source name |
ES cells
|
| Organism |
Mus musculus |
| Characteristics |
cell type: ES cells
cell line: E14 TG2a
age: NA
chip antibody/mbd affinity column: anti-H3K4me3 (ab8580; Abcam)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ES cells were crosslinked using 1% formaldehyde. Chromatin was sheared using sonication. Protein-DNA complexes were immunoprecipitated with an H3K4me3 antibody and the DNA was subsequently extracted using the genomic tip 500/G kit from Qiagen (cat no. 10262).
Libraries were prepared as follows using enzymes from New England Biolabs (NEB). End Repair was carried out by incubating immunoprecipitated DNA with T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase for 30 minutes at 20 C. The blunt, phosphorylated ends were treated with Klenow fragment (exo-) and dATP for 30 minutes at 37 C to yield a protruding 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. Ligated DNA was extracted using MiniElute PCR clean up columns (Qiagen) and PCR amplified with Illumina primers for 10-12 cycles. Library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were isolated by gel extraction. 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. Sequencing was performed at the Wellcome Trust Sanger Institute; Hinxton, Cambridge, UK.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer |
| |
|
| Description |
Chromatin IP against H3K4me3
|
| Data processing |
Alignment: Single-end sequence reads were mapped to the mouse genome (NCBIm37) using MAQ (http://maq.sourceforge.net/). Reads with a mapping score greater or equal to 30 where retained. Alignment was performed at the Wellcome Trust Sanger Institute; Hinxton, Cambridge, UK.
|
| |
|
| Submission date |
Apr 21, 2010 |
| Last update date |
May 15, 2019 |
| Contact name |
Adrian P Bird |
| E-mail(s) |
A.Bird@ed.ac.uk
|
| Phone |
0131 6508695
|
| Fax |
0131 6505379
|
| Organization name |
The University of Edinburgh
|
| Department |
WTCCB
|
| Lab |
Bird Lab
|
| Street address |
Mayfield Road
|
| City |
Edinburgh |
| State/province |
Midlothian |
| ZIP/Postal code |
EH9 3JR |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL9185 |
| Series (1) |
| GSE21442 |
Comprehensive analysis of orphan CpG islands identifes novel promoters with conserved DNA methylation dynamics |
|
| Relations |
| SRA |
SRX019618 |
| BioSample |
SAMN00011950 |
