 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jul 01, 2019 |
| Title |
in 1x3hr in vivo +TNFα rep3 |
| Sample type |
SRA |
| |
|
| Source name |
Bone marrow HSCs (Lin-/cKit+/Sca1+/Flk2-/CD48-/CD150+)
|
| Organism |
Mus musculus |
| Characteristics |
cell type: HSCs
treatment: in vivo 1x3hr in vivo +TNFα
strain: C57BL/6
tissue: Bone marrow
age: 6-12 weeks old
|
| Treatment protocol |
For ex vivo TNFα treatment, FACS-purified HSCs and GMPs were cultured in Iscove’s modified Dulbecco’s media (IMDM) containing 5% FBS (StemCell Technology), 50 U/ml penicillin, 50 μg/ml streptomycin, 2 mM L-glutamine, 0.1 mM non-essential amino acids, 1 mM sodium pyruvate and 50 μM 2-mercaptoethanol, supplemented with the following cytokines unless otherwise indicated (all from PeproTech except for TNFα): SCF (25 ng/ml), TPO (25 ng/ml), Flt3-L (25 ng/ml), IL-11 (25 ng/ml), IL-3 (10 ng/ml), GM-CSF (10 ng/ml) and EPO (4 U/ml) ± TNFα (1 μg/ml; Genentech).
For in vivo TNFα treatment, C57BL/6 mice were injected retro-orbitally with a single dose of 2 μg TNFα (Genentech) in 100 μl PBS or 100 μl PBS alone (untreated animals) 3 hours prior to BM harvest.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using RNeasy Plus Micro Kit (Qiagen). RNA integrity number (RIN) was determined by Bioanalyzer (Agilent Technologies) and RNA samples with RIN > 8.0 were subjected to further processing.
The double-stranded cDNA was generated using Ovation RNA-Seq System V2 (Nugen), and the sequencing libraries were prepared using LTP Library Preparation Kit (Kapa Biosystems). Different adaptors were used for multiplexing samples in one lane.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 3000 |
| |
|
| Data processing |
Data quality was verified on Sequencing Analysis Viewer (Illumina) and demultiplexing was performed with CASAVA 1.8.2 (Illumina).
Adapter trimming, alignment, and gene-level expression quantification was performed using STAR (Dobin et al., 2013).
Normalization was performed in R using the DESeq2 package (Love et al., 2014)
Genome_build: GRCm38/mm10
Supplementary_files_format_and_content: Tab-delimited text files include normalized read count values
|
| |
|
| Submission date |
Jun 06, 2018 |
| Last update date |
Jul 04, 2019 |
| Contact name |
Masayuki Yamashita |
| Organization name |
Institute of Medical Science, University of Tokyo
|
| Department |
Division of Stem Cell and Molecular Medicine, Center for Stem Cell Biology and Regenerative Medicine
|
| Street address |
4-6-1 Shirokanedai
|
| City |
Minato-ku |
| State/province |
Tokyo |
| ZIP/Postal code |
108-8639 |
| Country |
Japan |
| |
|
| Platform ID |
GPL21493 |
| Series (1) |
| GSE115403 |
Inflammation-produced TNFα protects hematopoietic stem cells from necroptosis and promotes hematopoietic regeneration |
|
| Relations |
| BioSample |
SAMN09374690 |
| SRA |
SRX4172836 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3177658_in_1x3hr_HSC_+TNF_rep3.txt.gz |
326.4 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
