GSM6625296: scRNA_KO_Spleen_Th1_Toxo_day7; Mus musculus; RNA-Seq
cell isolation protocol: Cells from spleen were obtained by mechanical disruption. Peritoneal cells were obtained by  washing of the peritoneal cavity with cold PBS. Isolated cells were further sorted as based on their surface markers. in vitro culture protocol: All cells were stimulated in RPMI medium with 10% (vol/vol) FCS (Invitrogen), 2 mM glutamine (Invitrogen), 100 IU/ml penicillin (Invitrogen), 0.1 mg/ml streptomycin (Invitrogen), and 20 mM HEPES buffer, pH 7.2–7.5 (Invitrogen), and 2 mM β-mercaptoethanol (Sigma-Aldrich). NK cells were treated with 1000 U/ml IL-2 and 10  ng/ml IL-12 (R&D) for 6 hours; ILC2 cells were treated with 50 ng/ml IL-25 (BioLegend) and 50 ng/ml IL-33 (Biolegend) for 4 hours; NCR+ILC3 were treated with 50ug/ml of IL-23 (R&D Systems) for 6 hours. Single cell suspensions were sorted using a FACS Aria Fusion (BD). ~15K cells at a concentration of 800 cells/μl were processed using the Chromium Next GEM Single Cell 3ʹ GEM, Library & Gel Bead Kit v3.1 (10x Genomics, #1000121), the Chromium Next GEM Chip G Single Cell Kit (10x Genomics, #1000120) and the Chromium Controller (10x Genomics, #1000202). cDNA libraries were constructed using the Chromium Single Cell 3ʹ Library Kit (10x Genomics, #1000079) and Single Index Kit T Set A (10x Genomics, #1000213).