|
| Status |
Public on Jan 17, 2023 |
| Title |
lung,PCW7, rep1 [10x299w3] |
| Sample type |
SRA |
| |
|
| Source name |
lung
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: lung
age: PCW7
technology: Chromium
library technology: 10Xv3
gender: female
|
| Treatment protocol |
N/A
|
| Growth protocol |
N/A
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The tissues were triturated with glass Pasteur pipettes every 15-20 min to enhance dissociation. After digestion, the cell suspension was filtered in a 15ml Falcon tube using a 30μm cell strainer (CellTrics, Sysmex), to remove clumps and debris. The cell suspension was kept ice cold and was diluted (roughly 1:2) with ice cold HBSS. The filtered cells were pelleted at 200g for 5 min at 4 °C and the pellet resuspended in a small volume of calcium- and magnesium-free HBSS (Gibco, cat no. 14170) and transferred to 1.5ml Eppendorf tubes precoated with 30% BSA (A9576, Sigma-Aldrich). A Bürker chamber was used for cell counting.
Single-cell RNAseq was carried out with the Chromium Single Cell 3´ Reagent Kit v2 and v3. Cell suspensions were counted and diluted to concentrations of 800 – 1200 cells/μl for a target recovery of 5000 cells on the Chromium platform. Downstream procedures including cDNA synthesis, library preparation and sequencing were performed according to the manufacturer’s instructions (10X Genomics Inc.). Libraries were sequenced on an Illumina NovaSeq 6000 (Illumina). We aimed to obtain 75.000 and 200.000 sequencing reads/cell for the v2 and v3 libraries respectively to match the different performances of the Chromium Single Cell 3´ Reagent v2 and v3 Kits and to achieve sufficient sequencing saturation. Across all 39 libraries we obtained an average of 187242 reads/cell.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
NextSeq 2000 |
| |
|
| Data processing |
Reads were aligned to the human reference genome and libraries were demultiplexed and aligned with the 10X Genomics pipeline CellRanger (version 3.0.2).
Loom files were generated for each sample by running Velocyto (0.17.17) 76 in order to map molecules to unspliced and spliced transcripts.
Assembly: GRCh38-3.0.0
Supplementary files format and content: Matrix tables with raw gene counts for every gene and every sample
|
| |
|
| Submission date |
Oct 17, 2022 |
| Last update date |
Jan 17, 2023 |
| Contact name |
Paulo Czarnewski |
| E-mail(s) |
czarnewski@gmail.com
|
| Organization name |
KTH
|
| Street address |
Tomtebodavägen 23b
|
| City |
Solna |
| ZIP/Postal code |
171 65 |
| Country |
Sweden |
| |
|
| Platform ID |
GPL30173 |
| Series (2) |
| GSE215895 |
A topographic atlas defines developmental origins of cell heterogeneity in the human embryonic lung [SC] |
| GSE215898 |
A topographic atlas defines developmental origins of cell heterogeneity in the human embryonic lung |
|
