GSM6122992: BM Adult; Mus musculus; RNA-Seq
1) Livers were cut immediately after dissection and kept in ice-cold digestion buffer (1.0 mg/ml Collagenase IV, 2 mg/ml DNase, 5% FBS in HBSS with Ca2++/Mg2++) to prevent loss of hepatic ILCs due to prolonged exposure to accumulating lactate. Embryos were removed from the uterus of pregnant mice and dissected under a microscope. All organs were digested with gentle rocking at 37°C for 20-30 min. After digestion, cells were passed through a 100 μm filter and washed, followed by 3 min incubation in ACK (Ammonium-Chloride-Potassium, Sigma Aldrich) buffer to lyse erythrocytes 2) For harvesting of bone marrow cells, tibiae and femora were carefully separated from surrounding muscle and connective tissue, crushed in a mortar, and passed through a 100 μm filter, followed by lysis of erythrocytes. 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 (paired-end reads, R1=28, i7=8, R2=91) to a depth of around 50,000 reads per cell.