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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jan 20, 2015 |
| Title |
TKO_DNMT3B1 |
| Sample type |
SRA |
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| Source name |
embryonic stem cells, mock treatment
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| Organism |
Mus musculus |
| Characteristics |
cell type: embryonic stem cells
genotype: DNMT1,3a,3b-triple-null
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| Growth protocol |
Mouse embryonic stem cells were cultivated on feeder cells or 0.2% gelatine coated dishes. ES cell growth medium consisted of DMEM (Invitrogen) supplemented with 15% foetal calf serum (Invitrogen), 1x non-essential amino acids (Invitrogen), 1mM L-glutamine, LIF and 0.001% beta-mercaptoethanol. Differentiation was performed as previously described (Bibel et al).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For cross-linking and chromatin extraction, cells were fixed for 10 minutes with 1% Formaldehyde at room temperature and incubated for 10 min on ice in presence of 1.2M Glycine. Cells were harvested and treated for 10 min with 10mM EDTA, 10mM TRIS, 0.5mM EGTA and 0.25% Triton X-100 and 10 min in 1mM EDTA, 10mM TRIS, 0.5mM EGTA and 200mM NaCl with subsequent lysis in 50mM HEPES, 1mM EDTA, 1% Triton X-100, 0.1% deoxycholate, 0.1% SDS and 150 mM NaCl for 2 hours on ice. Crosslinked chromatin was subjected to sonication in a Bioruptor instrument (Diagenode). ProteinA-dynabeads pre-cleared 150-250 µg chromatin were incubated with 40 µl blocked (1%CFSG, 100ng tRNA) Streptavidin-M280 magnetic beads over night at 4°C. Beads were washed with two rounds of 2% SDS, 1x high salt buffer, 1x LiCl Buffer and two rounds of TE. Beads were treated with RNaseA for 30 min at 37°C, Proteinase K for 3 hours at 50°C then de-crosslinked over night at 55°C. DNA was purified with Phenol-Chloroform extraction and EtOH precipitation.
ChIP-seq library using Illumina TruSeq library protocol with index adapters. Four to five libraries with different barcodes were pooled at equimolar ratios per lane
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
DNMT3B1_ChIPseq
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| Data processing |
basecalls using CASAVA
fastq to fasta conversion
reads were aligned to mm9 mouse genome using BOWTIE, allowing up to 2 missmatches
coordinates of unique reads that map only once to the mouse genome were converted to WIG format
Genome_build: mm9
Supplementary_files_format_and_content: wig track format: read coverage per consecutive 100bp windows
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| Submission date |
May 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Dirk Schuebeler |
| Organization name |
Friedrich Miescher Institute for Biomedical Research
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| Street address |
Maulbeerstrasse 66
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| City |
Basel |
| ZIP/Postal code |
4058 |
| Country |
Switzerland |
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| Platform ID |
GPL13112 |
| Series (2) |
| GSE57412 |
Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation [ChIP-Seq] |
| GSE57413 |
Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation |
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| Relations |
| BioSample |
SAMN02767326 |
| SRA |
SRX535254 |
| Named Annotation |
GSM1382262_MmTKO_CAGbioDnmt3b1_12_1.mm9-mmV02-aln2.w100.m1.i.wig.gz |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1382262_MmTKO_CAGbioDnmt3b1_12_1.mm9-mmV02-aln2.w100.m1.i.wig.gz |
21.9 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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