GSM8164512: HCO day 18; Homo sapiens; RNA-Seq
organoids were recovered from the Matrigel using Cell Recovery Solution and transferred in 15mL tubes. Then, organoids were washed using dPBS and dissociated with 500mL of trypsin-EDTA at 37°C. Full dissociation was achieved by manual dissociation using a micropipette. Trypsin-EDTA was inhibited by the addition of 4.5mL of DMEM/F12 supplemented with 10% FBS. Solutions were filtered using a 40µm cell strainer and centrifuged at 300g for 5min. Supernatant was removed and replaced by DMEM/F12 supplemented with Y-27632. Cells were counted and viability assessed using Trypan blue exclusion dye. scRNA-seq libraries were prepared using the 10x Genomics Chromium platform using version 3.1 chemistry. Sequencing was performed using a NovaSeq 6000 (Illumina) machine and Raw scRNA-Seq data was processed using CellRanger (v2.0.0 10x Genomics)