|
| Status |
Public on May 06, 2024 |
| Title |
kidney, TG saline control, day7 [TG_S_119] |
| Sample type |
SRA |
| |
|
| Source name |
kidney
|
| Organism |
Mus musculus |
| Characteristics |
tissue: kidney
genotype: TG
strain: FVB
batch: 3
|
| Extracted molecule |
nuclear RNA |
| Extraction protocol |
Mice were perfused with ice-cold PBS and kidney halves collected for snRNA-seq analysis. Half kidney was cut into small pieces (>25) and transferred into a 2-ml Dounce homogenizer (Sigma, Cat#D8938) loaded with 1 ml of NEZ Lysis Buffer (Sigma, Cat#N3408) with RNase inhibitor (NEB, Cat#M0314) at final concentration of 0.4U/µl) on ice. Samples were then Dounce homogenized on ice with five strokes of the looser pestle every 2 min for 8 min (25 strokes in total). Samples were then slowly Dounce homogenized 25 times with the tighter pestle on ice. The homogenized sample was filtered through a 40-μm Falcon Nylon Cell Strainer, then the filter was washed with 8 ml of 1% BSA PBS, and the nuclear suspension spun in a precooled (4 °C) centrifuge at 650g for 8 min. Supernatant was removed and the pellet resuspended in 1% BSA PBS (plus RNase inhibitor) and moved to a low-bind Eppendorf tube. Nuclear quality and number were assessed with trypan blue staining. Nuclei were sorted as DAPI positive on a BD Aria into 2% BSA solution with RNA inhibitor.
Barcoded single-cell gel beads in emulsion (GEMs) were created by 10x Genomics Chromium TM and then reverse transcribed to generate single-cell RNA-seq libraries using Chromium Single Cell 3’ Library and Gel Bead Kit v2 (10X Genomics) according to manufacturer’s instructions
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
10x Genomics
|
| Data processing |
Sequencing data were processed by CellRanger (version 6.0.0) and reads were aligned to mouse pre-mRNA reference genome (mm10-2020-A) with STAR (v.2.5.1b)
The Cell Ranger cellranger count function output filtered gene–cell expression matrices, removing cell barcodes not represented in cells.
UMI count table utilizing both exonic and intronic reads was generated for downstream analysis.
Assembly: mouse pre-mRNA reference genome (mm10-2020-A)
Supplementary files format and content: Tab-separated values files and matrix files
|
| |
|
| Submission date |
Nov 19, 2022 |
| Last update date |
May 06, 2024 |
| Contact name |
Anna Rinaldi |
| Organization name |
Ente Ospedaliero Cantonale
|
| Department |
Nephrology
|
| Lab |
Cippà
|
| Street address |
Tesserete
|
| City |
Lugano |
| State/province |
Ticino |
| ZIP/Postal code |
6900 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE218376 |
Spatiotemporal landscape of kidney tubular responses to glomerular proteinuria |
|
| Relations |
| BioSample |
SAMN31801972 |
| SRA |
SRX18321262 |
