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| Status |
Public on Sep 05, 2023 |
| Title |
Organoids from proximal part of small intestine of male old mice (105 weeks old) |
| Sample type |
SRA |
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| Source name |
Organoids from proximal part of small intestine of male old mice (105 weeks old)
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| Organism |
Mus musculus |
| Characteristics |
cell type: Epithelial cells
tissue: Organoids
strain: C57BL6/J
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| Treatment protocol |
Organoids devried from old mice were treated with IFNγ 2ng/ml for 24 hours.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Small intestinal crypts were isolated using the established protocol (Sato and Clevers, 2013) with some modifications. Briefly, mouse small intestine was dissected, washed in cold PBS. The villi-free intestinal pieces (2 cm) were washed with cold PBS and transferred to 5 mM EDTA/PBS, followed by two 30-min incubations at 4°C on a rotator. The tissue was transferred to fresh cold PBS and manually shaken for 30 sec. The crypt solution was filtered using a 70 µm cell strainer and centrifuged at 450g for 5 min at 4°C. Isolated crypts were immediately used or snap frozen in liquid nitrogen and stored at -80°C for further experiments. For RNA isolation, the crypts were immediately resuspended in QIAzol Lysis Reagent (Qiagen) and stored at -80°C. Crypts were resuspended in ice-cold Matrigel (Corning, Cat. 356231) and allowed to solidify at 37°C, then were supplied with culture medium (750ng/ml recombinant human Rspondin-1 (lab-made), 100ng/ml recombinant murine Noggin (PeproTech, Cat. 250-38), 50ng/ml recombinant murine EGF (PeproTech, Cat. 315-09), 10 μM CHIR99021 (Tocris, Cat. 4423/10), 100ng/ml recombinant murine Wnt-3a (PeproTech, Cat. 315-20), N2 (Thermo Fisher Scientific, Cat. 15410294), B27 (Thermo Fisher Scientific, Cat. 11500446), 1X Penicillin-Streptomycin (Thermo Fisher Scientific, Cat. 15140122) in Advanced DMEM/F12 (ThermoFisherScientific, Cat. 11550446)). Organoids were cultured for at least 2 weeks, passaged, then IFNgamma (2ng/ml) were added at day 3 and 24 hours later being collected for scRNA sequencing.
For single cell RNA sequencing: The 10X Single Cell 3’ v2 reagent kit (10X Genomics) was used for cell partitioning, cDNA amplification and library construction.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
control group and treated with IFNγ.
Single cell RNA sequencing
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| Data processing |
For single cell RNA sequencing: The fastq files are mapped against mm10.e87 genome following by counting the UMI and removing duplicates for single cells using 'cellranger count' (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/using/count ) with default parameters.
Genome_build: mm10.e87
Supplementary_files_format_and_content: Organoids_scRNAseq_combined2.rds: The processed data is a seurat object saved in RDS file with assigned meta data in trascriptome matrix of intesine
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| Submission date |
May 12, 2021 |
| Last update date |
Sep 05, 2023 |
| Contact name |
Seyed Mohammd Mahdi Rasa |
| E-mail(s) |
SeyedMohammadMahdi.Rasa@uksh.de
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| Phone |
015258164616
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| Organization name |
Universitätsklinikum Schleswig-Holstein
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| Department |
Institut for Immunology
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| Lab |
Immunology (Prof. Sheffold)
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| Street address |
Michaelisstrasse 5
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| City |
Kiel |
| State/province |
Schleswig-Holstein |
| ZIP/Postal code |
24105 |
| Country |
Germany |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE174297 |
In vitro grown organoids treated with IFNγ mimicking in vivo intestinal crypts cells aging |
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| Relations |
| BioSample |
SAMN19114815 |
| SRA |
SRX10852940 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5291229_Organoids_scRNAseq_combined2.rds.gz |
2.2 Gb |
(ftp)(http) |
RDS |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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