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Status |
Public on Jan 10, 2025 |
Title |
581_dTAG_H4K20me2_T0_rep3 |
Sample type |
SRA |
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Source name |
E14JU
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Organism |
Mus musculus |
Characteristics |
cell line: E14JU cell type: embryonic stem cells genotype: POLE4-dTAG, MCM2-2A clone 2 chip antibody: H4K20me2
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Treatment protocol |
Cells were incubated in EdU media (f.c. 10 µm) for 10 min and then collected (T0). If treated with DMSO or dTAG, cells were incubated for 1uM dTAG or DMSO prior to incubation with EdU for 10 min and then either collected (T0).
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Growth protocol |
ESCs were grown on gelatin-coated dishes in serum+LIF conditions at 37 °C with 5 % CO2. DMEM was supplied with fetal bovine serum (15 %), home-made LIF, non-essential amino acids, penicillin/streptomycin and beta-mercaptoethanol. Cells were passaged using Trypsin-EDTA or TrypLE. Drosophila S2-DRSC cells were cultivated in suspension in spinners using M3+BPYE media, 10% heat-inactivated FBS , and 1X penicillin/streptomycin . Cells were incubated at 25°C with 5% CO2.
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Extracted molecule |
genomic DNA |
Extraction protocol |
After the appropriate treatment, 1% formaldehyde for 10 min. Then, glycine was added to quench the reaction and cells were subsequently lysed. Sonication was performed on isolated nuclei using a Covaris E220. DNA fragments were immunoprecipitated, purified fragments were biotinylated, libraries were prepared, EdU-biotin fragments were pulled-down and sequenced. Libraries were costructed using the KAPA Hyperprep kit and Illumina-compatible indexed adapters (Pentabase) following manufacturer’s instruction. For the amplification step, 11 PCR cycles were used. Subsequently, the DNA underwent size selection using Agencourt AMPure XP beads (Beckman Coulter, A63881) to obtain fragments between 200-700 bp. Libraries were validated using Agilent Fragment Analyzer.
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Library strategy |
OTHER |
Library source |
genomic |
Library selection |
other |
Instrument model |
NextSeq 2000 |
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Description |
Library name: Sample76 581_dTAG_H4K20me2_RRPM_merged.bw
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Data processing |
adapters and low quality reads were filtered with TrimGalore (v0.0.6) reads were mapped to the genome with bowtie2 (v2.4.2) and uniquely mapped peaks kept using samtools (v1.12) duplicate reads were removed with picard (version 2.26.10) bam files were processed to bigwig for RPM by bamCoverage or RRPM in SeqMonk bigwigs: bam files were normalized to the number of million of reads (RPM) in 50 bp bins with bamCoverage Assembly: mm10 and dm6 Supplementary files format and content: bigwigs Library strategy: CHOR-seq
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Submission date |
Jan 09, 2025 |
Last update date |
Jan 10, 2025 |
Contact name |
Anja Groth |
E-mail(s) |
anja.groth@cpr.ku.dk
|
Organization name |
Novo Nordisk Foundation Center for Protein Research
|
Street address |
Blegdamsvej 3B
|
City |
Copenhagen |
ZIP/Postal code |
2200 |
Country |
Denmark |
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|
Platform ID |
GPL30172 |
Series (2) |
GSE270291 |
Disabling leading and lagging strand histone transmission results in parental histones loss and reduced cell plasticity and viability [ChOR-seq] |
GSE270297 |
Disabling leading and lagging strand histone transmission results in parental histones loss and reduced cell plasticity and viability |
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Relations |
BioSample |
SAMN46179565 |
SRA |
SRX27297577 |