|
| Status |
Public on May 22, 2024 |
| Title |
scRNAseq_CITEseq_CD34+cells_DONaK_CTRLaK_HTO |
| Sample type |
SRA |
| |
|
| Source name |
Umbilical cord blood
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Umbilical cord blood
disease state: Healthy
cell type: CD34+ cells
treatment: Mixed samples (DON 2.5uM + aK 5mM, Untreated + aK 5mM)
library type: HTO
hto: TotalSeq A0251 anti-human hashtag1, TotalSeq A0252 anti-human hashtag2
|
| Treatment protocol |
Cells were stimulated with cytokines after thawing and were grown in the same medium with or without metabolic inhibitor (2-DG, DON or AOA) until lysis was performed. Cells were collected and lysed after 96h of cytokine stimulation.
|
| Growth protocol |
CD34+ cells were purified from human healthy cord blood with a two-step immunomagnetic beads protocol and were then frozen in a cryopreservation medium. After thawing, CD34+ cells were cultured in a 96-well plate in a humidified 5% CO2 incubator at 37°C. Cells were grown in prestimulation medium made of X-Vivo (Lonza) supplemented with penicillin/streptomycin (respectively 100U/mL and 100µg/mL - Gibco, Thermo Scientific), 50 ng/ml h-FLT3, 25 ng/ml h-SCF, 25 ng/ml h-TPO, 10 ng/ml h-IL3 (Miltenyi) final concentration.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
High viability percentage was obtained using a Dead Cell Removal kit (Miltenyi). scRNA-seq with feature barcoding technology for cell surface protein was performed as described in the Chromium Next GEM Single Cell 3’ protocol for v3.1 from 10X Genomics (CG000206 Rev D). 20 000 cells were loaded on the chip to obtain around 5000 cells encapsidated in GEM.
Library was prepared following the 10X Genomics protocol and sequenced on a NextSeq 500/550.
scRNA-seq combined with CITE-seq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Demultiplexing + Matrix generation with Cellranger (pipeline version cellranger-6.0.1)
R (version 4.2.1) & Seurat (version 4.3.0) & sctransform (version 0.3.5) & ggplot2 (version 3.4.2) & org.Hs.eg.db (version 3.15.0) & clusterProfiler (version 4.4.4) & ReactomeGSA (v1.10.0) & ReactomeContentService4R (v1.2.0)
Filters : 1000<number of genes detected in a cell<6500, <25%mRNA, >5%rRNA, mitochondrial genes removed, genes detected in more than 3 cells
Assembly: hg38 (GRCh38)
Supplementary files format and content: cell ranger outputs
|
| |
|
| Submission date |
Sep 12, 2023 |
| Last update date |
May 22, 2024 |
| Contact name |
Laëtitia RACINE |
| E-mail(s) |
laetitia.racine@hotmail.fr
|
| Organization name |
EPHE
|
| Lab |
CRSA
|
| Street address |
27 rue de Chaligny
|
| City |
Paris |
| ZIP/Postal code |
75012 |
| Country |
France |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
| GSE243005 |
Metabolic adaptation pilots the differentiation of human hematopoietic cells (scRNA-Seq and CITE-Seq) |
| GSE243006 |
Metabolic adaptation pilots the differentiation of human hematopoietic cells |
|
| Relations |
| BioSample |
SAMN37367217 |
| SRA |
SRX21762899 |
