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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Nov 16, 2020 |
| Title |
HA36_TKO_DNMT3B1_r2 |
| Sample type |
SRA |
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| Source name |
Mouse Embryonic Stem Cells
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| Organism |
Mus musculus |
| Characteristics |
cell type: embryonic stem cells
genotype: Dnmt1,3a,3b,-tripleKO
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| Growth protocol |
ES cells were 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.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA extraction, followed by phenol/chloroform extraction and ethanol precipitation. RNaseA treatment was performed for 30 min at 37C
Prior library preparation unmethylated Lambda phage DNA and SssI-methylated T7 DNA were spiked-in to genomic DNA samples to control bisulphite conversion rates. For WGBS, 10ug of DNA was sonicated to ca 500bp using a Bioruptor Pico. DNA was end-repaired and A-tailed using the NEB-Ultra Illumina library preparation kit (E7370). DNA fragments were ligated to methylated adapters (NEB E7535) following the NEB Ultra protocol. After adapter removal using Ampure xp beads (Beckman Coulter), DNA was converted using the Qiagen Epitect bisulphite conversion kit following the FFT sample protocol. After conversion library PCR amplification was performed with 10 cycles following NEB Ultra kit protocol, and cleaned up using Ampure xp beads.
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
WGBS
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| Data processing |
Sequencing reads were pre-processed and trimmed using cutadapt v1.16 and sickle v1.33
Trimmed reads were mapped against the mm9 mouse assembly using bwa-meth v0.2.0 running bwa v0.7.12-r1039
Duplicates were removed with Picard MarkDuplicates (picard-tools v1.96, http://broadinstitute.github.io/picard/) with `REMOVE_DUPLICATES=TRUE and REMOVE_SEQUENCING_DUPLICATES=TRUE` flags.
Alignments with MAPQ > 40 were used as input to methylation calling with methyldackel v0.3.0-3-g084d926 (https://github.com/dpryan79/MethylDackel) (using HTSlib v1.2.1) in `cytosine_report` mode for all cytosines in the genome both CpG and CpH contexts (`extract -q 40 --cytosine_report --CHH –CHG`).
Genome_build: mm9
Supplementary_files_format_and_content: Processed tab delimited file contains cytosine report with following information: chromosome, nucleotide position, reported methylated cytosines, reported unmethylated cytosines, sequence context, exact sequence
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| Submission date |
Oct 20, 2020 |
| Last update date |
Nov 17, 2020 |
| Contact name |
Tuncay Baubec |
| Organization name |
Utrecht University
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| Department |
Department of Biology
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| Lab |
Genome Biology and Epigenetics
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| Street address |
Padualaan 8
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| City |
Utrecht |
| ZIP/Postal code |
3584 CH |
| Country |
Netherlands |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE151992 |
Flanking sequence preference modulates de novo DNA methylation in the mouse genome |
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| Relations |
| BioSample |
SAMN16484365 |
| SRA |
SRX9317309 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4837324_20200826.A-Sample2_pool1_bwameth_default.cytosine_report.txt.gz |
4.1 Gb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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