characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-06-01T00:15:14Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-06-01T00:15:14Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA110136675
characteristics.characteristic: External Id
characteristics.text: Institute of Immunobiology, Kantonsspital St. Gallen, St. Gallen, Switzerland
characteristics.characteristic: INSDC center name
characteristics.text: 2023-06-01T00:15:14Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-06-01T00:15:14Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS12235495
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11781:Sample 3
characteristics.characteristic: Submitter Id
characteristics.text: 8 to 10
characteristics.unit: week
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: non-hematopoietic cell
characteristics.characteristic: cell_type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Mice were sacrificed at age 8-10 weeks and lungs were harvested for further stromal cell or immune cell isolation and scRNA sequencing analysis. For cell preparation tissues were cut into small pieces. M-CoV immunization protocol: Mice were infected intranasally with 5 × 104 plaque-forming units (PFU) of M-CoV-A59 of M-CoV-GFP . Lungs were collected at 36 hours after infection for further processing for single cell RNA-sequencing Cell isolation from mouse lungs was performed according to standardized protocols. Mice were euthanized at the indicated time points and immediately perfused with PBS. Lungs were transferred into a 6-well plate containing RPMI 1640 medium (Sigma-Aldrich) supplemented with 4 U/mL elastase (Worthington), 1 U/mL Dispase II (Roche) and 10 μl/mL DNAse I (Applichem) and incubated for 45 minutes at 37°C. Lung tissue was then mechanically disrupted and washed in cold RPMI. Tissues were subject to a second enzymatic digestion in RPMI 1640 medium supplemented with 25 μg/mL Liberase TM (Roche) and 0.25 μg/mL DNAse I and incubated for 30 minutes at 37 °C. Any larger tissue fragments were dissociated mechanically followed by washing of the cell strainer with MACS buffer (PBS, 2% FCS, 2mM EDTA (Sigma-Aldrich)). Stromal and hematopoietic cell fractions were enriched using MACS anti-CD45 microbeads (Miltenyi Biotec). Cell sorting was performed on a BD FACSMelody™ cell sorter using BDChorus software. For discrimination between live and dead cells, Seven-aminoactinomycin D (Sigma-Aldrich) was added prior to sorting. Sorted cells were passed through the 10x Chromium (10x Genomics) system and cDNA libraries were generated according to the manufacturer’s recommendations (Chromium Single Cell 3′ Reagent Kit (version 3 chemistry)). cDNA library generation was performed following the established commercial protocol for Chromium Single Cell 3’ Reagent Kit (v3 Chemistry).
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: wild type genotype
characteristics.characteristic: genotype
characteristics.text: CD45-; GFP-
characteristics.characteristic: immunophenotype
characteristics.text: pool of 4 to 8 individuals
characteristics.characteristic: individual
characteristics.text: not applicable
characteristics.characteristic: isolate
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: lung
characteristics.characteristic: organism part
characteristics.text: Mus musculus
characteristics.characteristic: scientific_name
characteristics.text: female
characteristics.characteristic: sex
characteristics.text: C57BL/6N
characteristics.characteristic: strain
characteristics.text: Sample 3
characteristics.characteristic: title