GSM9078299: NK_14days; Mus musculus; RNA-Seq
Live NK1.1+NKp46+ cells from lungs and blood of female 9-weeks-old mice were sorted into PBS containing 2% FCS for the 10X Genomics protocol. Viability of the sorted cells was evaluated with trypan blue and was over 95% for all samples. Sorted cells were loaded into 10x Genomics Chromium in parallel. Libraries were prepared following manufacturer's protocol (Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 protocol) and sequenced on an Illumina NovaSeq sequencer according to 10X Genomics recommendations to a depth of around 50,000 reads per cell. Initial processing was done using Cell Ranger (v7.0.1) mkfastq and count (reads were aligned to GENCODE reference build GRCm38.p6 Release M23). Starting from the filtered gene-cell count matrices produced by CellRranger's built-in cell calling algorithms, we proceeded with Seurat v4 workflow (Hao et al. 2021). Initial processing was done using Cell Ranger (v7.0.1) mkfastq and count (reads were aligned to GENCODE reference build GRCm38.p6 Release M23). Starting from the filtered gene-cell count matrices produced by CellRranger's built-in cell calling algorithms, we proceeded with Seurat v5 workflow. Analysis, including quality control (QC), processing, graph-based clustering, visualizations, and differential gene expression analyses of the scRNA-seq data were performed in R using the Seurat pipeline (version 5.0).