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GEO help: Mouse over screen elements for information. |
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Status |
Public on Jun 28, 2024 |
Title |
MDF from KI-KI mice treated with DMSO - Replicate 1 |
Sample type |
SRA |
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Source name |
Mouse tail skin
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Organism |
Mus musculus |
Characteristics |
tissue: Mouse tail skin cell type: MDF antibody: Anti-FLAG M2 (Sigma-Aldrich, F1804) strain: p53-FLAG developmental stage: adult (8-12 weeks of age) mouse pool: B Sex: male sequencing run: 26-Aug-2022; Read Length: 161 bp; Cycle Kit: P1 300
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Extracted molecule |
genomic DNA |
Extraction protocol |
500,000 MDFs were plated and left to settle for 24 h in an incubator with 10% CO2 at 37°C. Medium was replaced with medium containing nutlin-3a (10 μM, Cayman Chemicals #18585) or DMSO (vehicle control) and cells incubated for 6 h. Adherent cells were dissociated with trypsin (0.25 mg/mL, Sigma #T4174), keeping both the supernatant and the trypsinised fractions for analysis. CUT&RUN was performed using the CUTANA ChIC/CUT&RUN kit v2 (Epicypher, 14-1408) as per the manufacture’s protocol. Nuceli were isolated prior to the start of the assay. Nuclei were bound to activated conA beads. Samples were incubated with 0.5 μg of anti-FLAG antibody (Sigma, F1804) overnight. DNA was cut using pAG-MNase, and unbound fragments washed away. Bound chromatin was release from the beads, and DNA was purified. Total recovered DNA was taken into library preparation using the NEBNext Ultra II DNA library Prep Kit for Illumina (#E7645L). CUT&RUN
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Library strategy |
OTHER |
Library source |
genomic |
Library selection |
other |
Instrument model |
NextSeq 2000 |
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Data processing |
Libraries were sequenced on an Illumina NextSeq 2000 to produce paired-end reads. Reads were aligned to the mouse genome mm39 with R/Bioconductor Rsubread package and its align function. Bam files from biological replicates were merged with samtools prior to creating BigWig files for each drug-genotype combination. Assembly: mm39 Supplementary files format and content: BigWig files were created with the deepTools BamCoverage program with reads per genomic content (RPGC) normalization in 10 bp windows. Duplicated reads, reads from sex and mitochondrial chromosomes, and reads overlapping ENCODE blacklisted regions from the mm39 genome were discarded for normalization purposes Supplementary files format and content: narrowPeak files (in BED format) are peak regions called by MACS2 for each genotype-drug combination. Biological replicates were combined by the MACS2 program during the peak calling step.
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Submission date |
Sep 25, 2023 |
Last update date |
Jun 28, 2024 |
Contact name |
Gordon K Smyth |
E-mail(s) |
smyth@wehi.edu.au
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Phone |
(+61 3) 9345 2326
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Fax |
(+61 3) 9347 0852
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URL |
http://www.wehi.edu.au
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Organization name |
Walter and Eliza Hall Institute of Medical Research
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Department |
Bioinformatics
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Lab |
Smyth
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Street address |
1G Royal Pde
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City |
Parkville |
State/province |
Vic |
ZIP/Postal code |
3052 |
Country |
Australia |
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Platform ID |
GPL30172 |
Series (1) |
GSE243999 |
Mouse models to investigate in situ cell fate decisions induced by TP53 and other factors (CUT&RUN) |
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Relations |
BioSample |
SAMN37536225 |
SRA |
SRX21882213 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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