GSM7383216: Renal organoids, d24, scRNA-seq, batch 1; Homo sapiens; RNA-Seq
For each sample, ~5000 organoids were collected in 15ml tubes, washed once with PBS then dissociated to single cells in 500µL Bacillus Licheniformis protease (Sigma, P5380) on ice for 20 min (Adam et al., 2017), with gentle agitation by pipetting every 2 min, followed by resuspension in PBS with 10% FBS to neutralise the protease. Cells were centrifuged at 300g for 5 min, then resuspended in PBS with 10% FBS and filtered through a 30µm strainer prior to centrifugation and final resuspension in PBS with 0.5% BSA. Cells were counted with Countess II (Life Technologies) to ensure cell viability > 90% and to achieve a targeted cell recovery of 1,600 cells per sample for batches 1 and 2, and 5,000 cells per sample for batch 3. Cells were processed for library preparation using a 10X Chromium sc 3'mRNA kit (v3, 10X Genomics) at the Cancer Research UK Genomics Facility, Cambridge, UK.