title: GSM9231186: BPNA Colon Tumour scRNA-Seq Rep4; Mus musculus; RNA-Seq
library_construction_protocol: Colonic tumour samples were collected in PBS and processed immediately for scRNAseq. Briefly, samples were chopped to achieve a paste-like consistency using a Mcllwain Tissue Chopper. Samples were transferred to GentleMACS C tubes (Miltenyi Biotec, 130-093-237) for tissue homogenisation using the GentleMACS Octo Dissociator with heaters (Miltenyi Biotec, 130-096-427) in combination with the Mouse tumour dissociation kit (Miltenyi Biotec, 130-096-730). Following tissue digestion, samples were resuspended in RPMI + 10% FBS + 2mM EDTA and filtered through 70μm and 40μm strainers respectively. The cell suspension was centrifuged at 400g for 3 mins and washed with RPMI + 0.4% BSA. The sample suspensions were centrifuged at 400g for 3 mins and the resultant pellets resuspended in 2% FCS + 25 mM HEPES + 2 mM EDTA + PBS for cell sorting. Cells were sorted using a BD FACSAria sorter (BD Biosciences) and DAPI (Invitrogen, D1306) to remove dead cells. Live sorted single cells were loaded onto a Chromium Chip G using reagents from the 10x Chromium Single-Cell 3' v3 Gel Bead Kit and Library (10x Genomics) according to the manufacturer's protocol.