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| Status |
Public on Apr 07, 2020 |
| Title |
Nsd1KO_none_Maintainance_Input_rep2 |
| Sample type |
SRA |
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| Source name |
Cultured erythroblast from bone marrow
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| Organism |
Mus musculus |
| Characteristics |
cell type: Cultured erythroblast from bone marrow
add-back of nsd1: none
genotype: NSD1-/-
growth_medium: Maintainance
time: 0h
antibody: Input
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| Treatment protocol |
Cell are NSD1 KO transduced with either wildtype Nsd1 or Nsd1N198Q with disrupted SET-domain
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| Growth protocol |
Either maintainance or differentiation medium, as indicated. Maintenance medium: StemSpan SFEM (StemCell Technologies, Vancouver, Canada), supplemented with 1%Pen/Strep, 0.4%cholesterol (Gibco, Thermo Fisher Scientific, Reinach, Switzerland), 2U/ml hEpo (Eprex 4000, 9096976, Pharmacy of University Hospital Basel), 100ng/ml mScf(Peprotech, London, UK), 10-6M dexamethasone (Calbiochem, Sigma Aldrich, Buchs, Switzerland) and 40ng/ml hIGF-1 (Peprotech, London, UK). Differentiation medium composed of IMDM (Gibco, Thermo Fisher Scientific, Reinach, Switzerland), 1%P/S, 10%FCS, 10%PFHMII (Gibco, Thermo Fisher Scientific, Reinach, Switzerland), 5%hPDS (0.45µM filtered, Blood donation service, University Hospital Basel), monothioglycerol (Sigma Aldrich, Buchs, Switzerland), 100ng/ml mSCF and 2U/ml hEpo
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Pull-down beads were Protein G sepharose (Sigma-Aldrich, P3296). Washes were done with the low salt, high salt, li-Cl wash buffers according to EZ-Magna CHIP A/G kit protocol from Millipore. DNA elution was done with the IPURE Kit V2 (Diagenode, Cat.No. C03010012). Antibodies: GATA1 ab11862 10 ug per ChIP Abcam; H3K36me3: ab9050-11 10 ug per ChIP Abcam; H3K27ac: ab4729 10 ug per ChIP Abcam
ChIP protocol was adapted from the EZ-Magna ChIPTM A/G kit protocol (Millipore, Merck KGaA, Darmstadt, Germany). Cells (Nsd1 WT and Nsd1 Set in maintenance as well as 24h differentiation) were fixed with 1% formaldehyde, lysed with Cell Lysis and then Nuclear Lysis buffers to a concentration of 20^106 cells per mL, and finally sonicated (20-min cycle on Covaris apparatus; KBioscience). Sheared chromatin was immunoprecipitated overnight using the following antibodies: anti-GATA1 (Abcam, ab11852), anti-H3K36me3 (Abcam, ab9050-100), anti-H3K27ac (Abcam, ab4729). 1/10 of the sheared chromatin was used as a reference (Input). Immune complex collection was performed using Protein G Sepharose (Sigma-Aldrich, P3296) for 1h30 at 4°C. Rinses were done according to Magna ChIPTM kit protocol with Low salt, High salt and LiCL immune complex wash buffers. Finally, elution was performed according the IPure Kit protocol (Diagenode, Cat.No. C03010012) following manufacturer’s instructions. Two independent ChIP-seq experiments were conducted for Gata1, H3K27ac and H3K36me3.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Reads were aligned with bowtie2 to the mouse genome (UCSC version mm10). The output was sorted and indexed with samtools.
Duplicated reads were marked with picard. Coverage tracks per sample were generated by tiling the genome in 20bp windows and counting 5'end of reads per window using the function bamCount from the bioconductor package bamsignals.
These window counts were exported in bigWig format using the bioconductor package rtracklayer.
Genome_build: mm10
Supplementary_files_format_and_content: BigWig file with FPKM counts per bin
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| Submission date |
Sep 03, 2019 |
| Last update date |
Nov 17, 2021 |
| Contact name |
DBM Bioinformatics Core Facility |
| Phone |
+41612073541
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| Organization name |
University of Basel
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| Department |
Departement of Biomedicine
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| Street address |
Hebelstrasse 20
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| City |
Basel |
| State/province |
BS |
| ZIP/Postal code |
4053 |
| Country |
Switzerland |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE136808 |
Inactivation of the nuclear interacting SET domain protein 1 (NSD1) impairs GATA1-regulated erythroid differentiation and causes erythroleukemia. NSD1 Add-back experiment. ChIPSeq for Gata1, H3K27ac and H3K36me3. |
| GSE136811 |
Inactivation of the nuclear interacting SET domain protein 1 (NSD1) impairs GATA1-regulated erythroid differentiation and causes erythroleukemia. |
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| Relations |
| BioSample |
SAMN12686896 |
| SRA |
SRX6797017 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4058407_KO3_Input_MM_mm10.bigWig |
439.9 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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