|
| 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 |
|
| Relations |
| BioSample |
SAMN17001946 |
| SRA |
SRX9632458 |
