GSM6265659: sample1_Kras LA2; Mus; RNA-Seq
LLC1 or KRasLA2 tumor bearing mice CD45 cell isolation. Briefly, lungs were minced into small pieces and treated with Collagenase 5% and DNAase 1% containing buffer for 30 min at 37°C. Afterwards cells were passed through a 100µM cell strainer followed by 40µm nylon mesh. After a 10 min centrifugation step at 500 x g, cells were incubated in 2ml Erylysis buffer for red blood cell removal. After 4 min incubation time, lysis was stopped by adding 23 ml PBS. Cells were resuspended in PBS and FACS sorted for CD45 cells using an AriaIII-sorter and 7-AAD as indicator for dead cells. After sorting the cell suspensions were counted with MOX Z cell counter and diluted according to manufacturer's protocol to obtain 10.000 single cell data points per sample. Each sample was run separately on a lane in Chromium controller with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (10xGenomics). Single cell RNAseq library preparation was done using standard protocol.