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Sample GSM4956335 Query DataSets for GSM4956335
Status Public on Dec 05, 2020
Title WT_HSC
Sample type SRA
Source name WT hematopoietic stem cell
Organism Mus musculus
Characteristics genotyping: Ezh2 +/+ CreERT+/-
surface markers: Lin- Sca-1+ c-kit+ CD34- CD150+ CD48- Flt3-
strain: C57BL/6
Treatment protocol To generate a gene set upregulated in Ezh2-deficient HSCs, BM cells (1-5x106) from Cre-ERT;Ezh2+/+ and Cre-ERT;Ezh2fl/fl CD45.2 mice were transplanted into lethally irradiated CD45.1 recipient mice. Ezh2 was deleted by intraperitoneal injection of 100 μl of tamoxifen (10 mg/ml) a day for 5 consecutive days as previously described (Mochizuki et al., Blood 2015). HSCs (CD150+CD48–CD34–Flt3–LSK) were purified from recipient mice 1 months after tamoxifen injection, and were subjected to RNA-seq analysis.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using RNeasy Plus Micro Kit (Qiagen)
Total RNA were subjected to reverse transcription and amplification with a SMARTer Ultra Low Input RNA Kit for Sequencing (Clontech). After sonication with an ultrasonicator (Covaris), the libraries for RNA-seq were generated from fragmented DNA with 8 cycles of amplification using a NEB Next Ultra DNA Library Prep Kit (New England BioLabs). After the libraries were quantified using the Bioanalyzer (Agilent), samples were subjected to sequencing with Hiseq1500 (Illumina) and 61 cycles of the sequencing reactions were performed.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 1500
Data processing alignment:TopHat2 (version 2.0.13; with default parameters) and Bowtie2 (version 2.1.0) were used for alignment to the reference mouse genome (mm10 from the University of California, Santa Cruz Genome Browser;) with annotation data from iGenomes (Illumina)
Row counts for each gene symbol are generated by StringTie
Normalization and statistical analysis were performed using DEGES-edgeR of TCC package, and 448 genes were selected using cutoff of q<0.05
For making tdf file, bedtools genomecov was used to make bedgraph file and
Genome_build: mm10
Submission date Dec 04, 2020
Last update date Dec 05, 2020
Contact name Atsushi Iwama
Organization name The Institute of Medical Science, The University of Tokyo
Department Center for Stem Cell Biology and Regenerative Medicine
Lab Division of Stem Cell and Molecular Medicine
Street address 4-6-1, Shirokanedai
City Minato-ku
State/province Tokyo
ZIP/Postal code 108-8639
Country Japan
Platform ID GPL18480
Series (2)
GSE162659 Epigenetic traits inscribed in chromatin accessibility in aged hematopoietic stem cells (EZH2_KO_HSC)
GSE162662 Epigenetic traits inscribed in chromatin accessibility in aged hematopoietic stem cells
BioSample SAMN17001946
SRA SRX9632458

Supplementary file Size Download File type/resource
GSM4956335_EZH2_WT_HSC_RNA_raw_count_gene.txt.gz 93.6 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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