|
| Status |
Public on Aug 25, 2022 |
| Title |
implanted organoid +PBMC 2, scRNAseq |
| Sample type |
SRA |
| |
|
| Source name |
human induced pluripotent stem cell-derived kidney organoid
|
| Organisms |
Homo sapiens; Mus musculus |
| Characteristics |
mouse strain: BALB/c IL2Ry-/- RAG2-/-DKO
tissue: human induced pluripotent stem cell-derived kidney organoid
protocol: implanted organoid +PBMC
|
| Treatment protocol |
none
|
| Growth protocol |
A human iPSC line was pre-treated with CHIR99201 for four days and fibroblast growth factor 9 (FGF9) for three days after which cell pellets were formed, which were placed on a Transwell membrane at the air – medium interphase. The organoids were harvested on day 18. Whole kidney organoids were then subcutaneously implanted into 4 dorsal pockets (2 per pocket) in immune deficient mice. PBMCs from healthy human donors were adoptively transferred 1 month after implantation. PBMC were thawed and 10 million PBMC were administered in 200 μL of phosphate buffered saline (PBS) via intraperitoneal injection. Two months after kidney organoid implantations and four weeks after human PBMC administration, mice were sacrificed by cervical dislocation. The organoids were harvested.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Single cell suspension of organoids were generated using TrypLE Select for 15 minutes at 37°C and stored on ice in APEL
Single cell libraries were prepared using the Chromium Single Cell 3’ Reagent Kit v3 (10x Genomics, Pleasanton, CA, USA). Next generation sequencing (28-8-0-91 cycles) was carried out on an Illumina NovaSeq6000 platform (Illumina, San Diego, USA).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
Sample 5 (PBMC 2)
10x Genomics
|
| Data processing |
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v6.0.0
Reads containing sequence information were mapped to both the GRCh38 human genome and mm10 mouse genome (barnyard) reference
Generation of BAM files and filtered gene-barcode matrices was accomplished using the Cell Ranger Software (version 6.0.0)
Using Seurat (version 4.0.5) standard pre-processing was done. Selection and filtration of cells was done (genes that were expressed in fewer than three cells, and cells with fewer than 200 distinct transcripts were removed). Data was normalized and scaled, followed by the detection of highly variable genes.
From the generated Seurat-Object, unnormalized data (raw counts) were exported as a comma separated table in CSV files.
Assembly: GRCh38 mm10 (barnyard)
Supplementary files format and content: Comma separated values files and matrix files
|
| |
|
| Submission date |
Aug 22, 2022 |
| Last update date |
Sep 19, 2022 |
| Contact name |
Hector Tejeda Mora |
| E-mail(s) |
h.tejedamora@erasmusmc.nl
|
| Organization name |
Erasmus MC
|
| Department |
Internal Medicine
|
| Lab |
Nephrology and Transplantation
|
| Street address |
Doctor Molewaterplein 40
|
| City |
Rotterdam |
| ZIP/Postal code |
3015 GD |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL25526 |
| Series (1) |
| GSE211803 |
The immunogenicity of human kidney organoids |
|
| Relations |
| BioSample |
SAMN30443517 |
| SRA |
SRX17177747 |
