GSM6265809: kidney, wt3; Mus musculus; RNA-Seq
Control and mutant littermate mice (both male and female) were used for single-cell transcriptomic and morphological analysis of the adult kidney. Mice were anesthetized with isoflurane overdose and transcardially perfused with PBS. Kidneys were dissected out. One kidney per mouse was fixed in 4% PFA at 4°C overnight to be used for morphological analysis (see section: Adult kidney preparation for micro-computed tomography). The second kidney was chopped in small pieces and digested with 2 mg/ml collagenase P for 10 min at 37°C, an equal volume of 0,10% trypsin/EDTA was added and tissue was digested for an additional 10 min at 37°C. Following enzymatic digestion, tissue was triturated using a wide-bore pipette tip and clumps were mechanically dissociated using a 100 µm mesh and a syringe plunger. Cell suspensions were washed with PBS/10% FBS twice and resuspended at a concentration of 1,000 cells/µl for processing with a 10X controller. 10x Genomics Chromium Single Cell 3ʹ protocol for Reagent Kits v3 was used for library preparation.