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| Status |
Public on Aug 27, 2021 |
| Title |
RNA-seq_E EOMES |
| Sample type |
SRA |
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| Source name |
Hematopoietic stem cell
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| Organism |
Homo sapiens |
| Characteristics |
tissue: umbilical cord blood
cell type: Hematopoietic stem cell
donor: Donor E
treatment: Retroviral transduction
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| Treatment protocol |
Pre-cultured CD34+ HPC were harvested, transferred to RetroNectin (Takara Bio, Saint-Germain-en-Laye, France)-coated plates and viral supernatant was added. Additional cytokines were added to keep the concentrations constant after virus addition. The plates were centrifuged at 950 g and 32°C during 90 min. Another 48 hours after transduction, lineage-(CD3/CD14/CD19/CD56) CD34+eGFP+ HPC were sorted using a BD FACSAria™ Fusion cell sorter (BD Biosciences).
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| Growth protocol |
HPC were obtained by isolating peripheral blood monocytes form umbilical cord blood by density gradient centrifugation and subsequently CD34+ HPC were enriched by magnetic activated cell sorting (MACS). Isolated cord blood-derived CD34+ HPC were cultured in complete IMDM containing 10% FCS and supplemented with thrombopoietin (TPO) (20 ng/ml), stem cell factor (SCF) (100 ng/ml) (all from Peprotech, London, UK.) and FMS-like tyrosine kinase 3 ligand (FLT3-L) (100 ng/ml, R&D Systems, Minneapolis, MN, U.S.A.) for 48 hours.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total mRNA was extracted using the RNeasy micro kit (Qiagen). The concentration and quality of the total extracted RNA was checked by using the Quant-it ribogreen RNA assay (Life Technologies) and the RNA 6000 nano chip (Agilent Technologies, Santa Clara, CA, U.S.A), respectively.
Illumina sequencing library preparation was performed using the QuantSeq 3' mRNA-Seq Library Prep Kits (Lexogen, Vienna, Austria) according to manufacturer's protocol. Libraries were quantified by qPCR.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
HSCsample_15
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| Data processing |
Fastq files were aligned to human reference genome GRCh38 using STARv2.42 and gencode v25 as guide gtf
Counts were generated on the fly by STAR
QC of fastq files was done with FastQC
Genome_build: GRCh38
Supplementary_files_format_and_content: a tab separated raw count matrix file
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| Submission date |
Feb 09, 2021 |
| Last update date |
Aug 27, 2021 |
| Contact name |
Wouter Van Loocke |
| E-mail(s) |
[email protected]
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| Organization name |
Ghent University
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| Department |
Biomolecular Medicine
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| Street address |
C. Heymanslaan 10
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| City |
Ghent |
| State/province |
East Flanders |
| ZIP/Postal code |
9000 |
| Country |
Belgium |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE166437 |
Transcriptome profiling of hematopoietic progenitor cells (HPC) transduced with transcription factors T-BET or EOMES |
| GSE166439 |
ATAC-seq and RNA-seq of hematopoietic progenitor cells (HPC) and Natural Killer (NK) cells transduced with transcription factors T-BET or EOMES |
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| Relations |
| BioSample |
SAMN17841260 |
| SRA |
SRX10058259 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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