|
| Status |
Public on Sep 10, 2019 |
| Title |
Control Sample 2 (snRNAseq) |
| Sample type |
SRA |
| |
|
| Source name |
Human Kidney
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: kidney
disease state: control
age: 62
a1c: NA
gfr: 60.7
global glomerulosclerosis: None <10%
ifta: 1-10%
gender: M
|
| Growth protocol |
ES cell–derived NS cells were routinely generated by re-plating d 7 adherent neural differentiation cultures (typically 2–3 × 10e6 cells into a T75 flask) on uncoated plastic in NS-A medium (Euroclone, Milan, Italy) supplemented with modified N2 and 10 ng/ml of both EGF and FGF-2 (NS expansion medium).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Nuclei were isolated with Nuclei EZ Lysis buffer (NUC-101; Sigma-Aldrich) supplemented with protease inhibitor (5892791001; Roche) and RNase inhibitor (N2615; Promega and AM2696; Life Technologies). Samples were cut into <2-mm pieces and homogenized using a Dounce homogenizer (885302–0002; Kimble Chase) in 2 ml of ice-cold Nuclei EZ Lysis buffer, and they were incubated on ice for 5 minutes with an additional 2 ml of lysis buffer. The homogenate was filtered through a 40-μm cell strainer (43–50040–51; pluriSelect) and then centrifuged at 500×g for 5 minutes at 4°C. The pellet was resuspended and washed with 4 ml of the buffer, and then, it was incubated on ice for 5 minutes. After another centrifugation, the pellet was resuspended in Nuclei Suspension Buffer (1× PBS, 0.07% BSA, and 0.1% RNase inhibitor), filtered through a 20-μm cell strainer (43–50020–50; pluriSelect), and counted.
Libraries were prepared according to Illumina's instructions and accompanying 10X Genomics Chromium Single Cell 5' Library and Gel Bead Kit
Single Nucleus RNA Sequencing (snRNAseq)
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
snRNAseq
|
| Data processing |
Count matrices were generated with zUMIs v2.0 and saved as .rds files
Seurat objects were created from the count matrices and filtered using min.cells=3 and low.thresholds=500 (Seurat v2.3.4)
Seurat objects from control and diabetic samples were combined using the RunMultiCCA function to generate combined.rds
Genome_build: GRCh38
Supplementary_files_format_and_content: individual rds files were generated from count matrices using zUMIs v2.0 and Seurat v2.3.4
|
| |
|
| Submission date |
May 29, 2019 |
| Last update date |
May 27, 2020 |
| Contact name |
Parker C Wilson |
| Organization name |
Washington University in St. Louis
|
| Street address |
660. S Euclid Ave., CB 8118
|
| City |
SAINT LOUIS |
| State/province |
MO |
| ZIP/Postal code |
63110 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE131882 |
The Single Cell Transcriptomic Landscape of Early Human Diabetic Nephropathy |
|
| Relations |
| Reanalyzed by |
GSM4572193 |
| BioSample |
SAMN11878527 |
| SRA |
SRX5914366 |
