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Sample GSM5112198 Query DataSets for GSM5112198
Status Public on Dec 16, 2021
Title KR02
Sample type SRA
 
Source name iPSC-derived kidney organoids without SARS-CoV-2 infection
Organism Homo sapiens
Characteristics tissue: Kidney organoid
condition: CTR
Treatment protocol To study direct effects of the virus on kidney cells independent of systemic effects we chose human induced pluripotent stem cell (iPSC)-derived kidney organoids to model SARS-CoV-2 infection in vitro, using the Bavarian strain of the virus (MT270101.1). iPSC-derived kidney organoids were infected with SARS-CoV-2 at an MOI of 1.0 at d7+18. Virus incubation lastet 4h until changing the medium back to E6 without virus and incubating the organoids for five more days.
Growth protocol To generate human induced pluripotent stem cell (iPSC) derived kidney organoids, iPSCs were seeded using a density of 18,000 cells/cm2 on Geltrex (Thermo Fisher, Breda, the Netherlands) coated 6-well plates (Greiner Bio-one, Alphen aan de Rijn, the Netherlands). The differentiation protocol was modified from Takasato et al. (Takasato et al. 2016). On day 0 (d 0), differentiation was initiated using CHIR-99021 (6 µM, R&D systems, Abingdon, United Kingdom) in Essential 6 medium (E6, Thermo Fisher). CHIR treatment was maintained for 3 or 5 days and medium was replaced sequentially for E6 supplemented with fibroblast growth factor 9 (FGF9), 200 ng/ml, R&D systems) and heparin (1 µg/ml, Sigma-Aldrich, Zwijndrecht, Netherlands) up to d 7. Cell layers were trypsinized and suspensions were counted on d 7. One part 3 days CHIR-differentiated cells was mixed with two parts 5 days CHIR-differentiated cells. To generate cell aggregates, cells were aliquoted using 300,000 cells per 1.5 ml tube and centrifuged. Cell aggregates were plated on Costar Transwell filters (type 3450, Corning, Sigma-Aldrich). One hour CHIR pulse (5 µM) in E6 was used to stimulate self-organizing nephrogenesis and the medium was replaced for E6 supplemented with FGF9 and heparin for additional 5 days. As of d 7+5, organoids were cultured using E6 media supplemented with human epidermal growth factor (hEGF, 10 ng/ml, Sigma-Aldrich), bone morphogenetic protein-7 (BMP7, 50 ng/ml, R&D systems), stromal derived factor 1 beta (SDF1β, 10 ng/ml, R&D systems), vasopressin (10 nM, Sigma-Aldrich) and aldosterone (10 nM, Sigma-Aldrich) until viral infection at d 7+18.
Extracted molecule total RNA
Extraction protocol All protocols to generate scRNA-seq data on the 10x Chromium platform, including sample prep, library prep, instrument and sequencing settings, are available here: https://support.10xgenomics.com/single-cell-gene-expression
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Data processing The Cell Ranger Software Suite (version 3.1.0) was used for processing sequencing information and single cell barcodes.
Seurat (version 3.2.2) was used to integrate the scRNA-seq data and predict cell labels.
Genome_build: GRCh38
Supplementary_files_format_and_content: barcodes: barcodes present in input data
Supplementary_files_format_and_content: matrix: a count matrix with Market Exchange Format (MEX). The first line contains the type code and the second line includes format and cell-ranger versions. Number of peaks, barcodes, and non-zero entries can be found in the third line.  The rest of the lines represent  nonzero entries with the first two numbers corresponding to peak and barcode indices and the last one corresponding to  the number of cut sites.
Supplementary_files_format_and_content: genes: a bed file contains genes corresponding to row indices. For each gene, its gene ID and name are stored in the first and second column of the gene.tsv file, respectively
 
Submission date Feb 26, 2021
Last update date Dec 16, 2021
Contact name Ivan Gesteira Costa
E-mail(s) ivan.costa@rwth-aachen.de
Phone +49 241 80 89337
Organization name UKA RWTH Aachen
Department Joint Research Center for Computational Biomedicine
Lab Institute for Computational Genomics
Street address Pauwelsstr. 19
City Aachen
State/province Northrhine-Westphalia
ZIP/Postal code 52074
Country Germany
 
Platform ID GPL24676
Series (1)
GSE167747 SARS-CoV-2 infects the human kidney and drives fibrosis in kidney organoids
Relations
BioSample SAMN18075507

Supplementary file Size Download File type/resource
GSM5112198_KR02_barcodes.tsv.gz 56.3 Kb (ftp)(http) TSV
GSM5112198_KR02_features.tsv.gz 325.6 Kb (ftp)(http) TSV
GSM5112198_KR02_matrix.mtx.gz 74.7 Mb (ftp)(http) MTX
Raw data not provided for this record
Processed data provided as supplementary file

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