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| Status |
Public on Jan 12, 2022 |
| Title |
PFC Setd1a+/- H3K4me3 CUT&RUN Sample1 |
| Sample type |
SRA |
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| Source name |
PFC
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6N
tissue: PFC
Sex: male
age: 8-10 weeks
treatment: Setd1a CRISPR-Cas9
chip antibody: H3K4me3 (Cell Signaling 9727S)
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| Growth protocol |
Adult mice were housed in our animal facility with 12 h light/dark cycles and food ab libitum. Animals were used for analysis in accordance with protocols approved by the Institutional Animal Care and Use Committee
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
FACS sorted nuclei were used for CUT&RUN experiments. Briefly, Foxp2-Cre or Foxp2-Cre/Setd1a+/- mice were euthanized by inhalation of CO2. Brains were rapidly removed and the prefrontal cortex was dissected and immediately frozen with dry ice and then stored at −80 °C until processing for nuclear isolation. Frozen brain samples were homogenized in 1 ml ice-cold homogenization buffer [320 mM sucrose, 5 mM CaCl2, 3 mM Mg(Ac)2, 10 mM Tris pH7.6, 0.1 mM EDTA, 0.1% NP40, 0.1 mM PMSF, 1 mM β-mercaptoethanol, 1% BSA, 1:250 RNasin Plus RNase Inhibitor (Clontech)] using a 1 ml Dounce homogenizer (Wheaton); 20 times with pestle A, followed by 20 times with pestle B. After 10 min on ice, the homogenate was filtered with 40 μm cell strainer (Fisher) and added 1 ml dilution buffer [50% OptiPrep density gradient medium (Sigma), 5 mM CaCl2, 3 mM Mg(Ac)2, 10 mM Tris pH 7.6, 0.1 mM PMSF, 1 mM β-mercaptoethanol] and mixed thoroughly with pipette. Loaded 0.5 ml lysate on the top of 0.5 ml 29% iso-osmolar OptiPrep solution (in PBS) in a 1.5 ml centrifuge tube and centrifuged at 6000×g for 10 min at 4 °C. After removing the supernatant, the nuclei were resuspended in wash buffer [2.5 mM MgCl2, 1% BSA in PBS, 1:500 RNasin Plus RNase Inhibitor (Clontech)] for FACS. The GFP+ nuclei were directly sorted into CUT&RUN wash buffer with Sony SH800 sorter. The sorted nuclei were immediately used for CUT&RUN experiments.
CUT&RUN libraries were constructed as previously described (Skene and Henikoff, 2017 eLife. 2017; 6: e21856, PMID: 28079019).
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Library strategy: CUT&RUN
Raw reads were first preprocessed to remove low quality reads and adaptor sequences using Trimgalore (v0.4.5) (https://github.com/FelixKrueger/TrimGalore) with the parameter “--paired”. The trimmed reads were then aligned to the mm10 genome using Bowtie2 (v2.3.4.3) [PMID: 19261174] with parameters “‘--no-unal --no-mixed --no-discordant -I 150 -X 2000”. Samtools (v1.9) [PMID: 19505943] was used to remove low mapping quality (mapq <10) and unmapped reads. Only uniquely mapped reads were kept for downstream analysis. Bigwig tracks were generated using “bamCoverage” command from deepTools (v3.0.2) [PMID: 24799436] with parameters “--binSize 20 --extendReads 250 --normalizeUsing RPKM --outFileFormat bigwig --minMappingQuality 30 --scaleFactor 1”. The bigwigs of the CUT&RUN samples with yeast spiked-in data were scaled based on the number of uniquely mapped reads to the yeast genome normalized to 100,000 reads (saling factor=10^5/[#uniquely mapped spikein reads]). The differential peaks were detected using the R/Bioconductor package Linnorm (v.2.16.0) [PMID: 28981748] for non-spikein CUT&RUN data, and we used the R/Bioconductor package csaw (v1.20.0) [PMID: 26578583] for comparison between spiked-in samples
Genome_build: mm10
Supplementary_files_format_and_content: bigwig
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| Submission date |
Jul 28, 2021 |
| Last update date |
Jan 13, 2022 |
| Contact name |
Mohamed Nadhir Djekidel |
| E-mail(s) |
djek.nad@gmail.com
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| Organization name |
Sidra Medicine
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| Department |
Translational Medicine Department
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| Lab |
Maternal and Child Health Division
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| Street address |
Gharafa
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| City |
Doha |
| State/province |
Doha |
| ZIP/Postal code |
26999 |
| Country |
Qatar |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE181025 |
Cell type-specific mechanism of Setd1a heterozygosity in schizophrenia pathogenesis [CUT&RUN] |
| GSE181027 |
Cell type-specific mechanism of Setd1a heterozygosity in schizophrenia pathogenesis |
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| Relations |
| BioSample |
SAMN20457275 |
| SRA |
SRX11583691 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5482196_PFC_Setd1aHet_H3K4me3_Rep1.bw |
65.7 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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