title: GSM8488285: C12_20d_10Dol_1; Homo sapiens; RNA-Seq
library_name: GSM8488285
library_construction_protocol: To minimize organoid-to-organoid variability, 5-10 organoids were pooled into one sample for analysis. Organoids were homogenized in 400 μL TRIzol (Invitrogen) by manual pipetting until complete lysis. RNA was isolated by addition of 80 μL chloroform (Sigma-Aldrich), vortex and centrifugation at 4ºC for 15 minutes at 12,000 g, and careful collection of 200 μL of the upper aqueous layer. RNA clean-up was performed using a silica spin column (Zymo) according to the manufacturer's instructions, followed by elution in 15 μL water and analysis by NanoDrop 2000 (Thermo Scientific). RNA was immediately extracted, minimizing any additional steps that could lead to RNA degradation, which preserves RNA integrity for further processing. RNA-seq library preparation was performed as previously described (Jacob et al 2020, Weng et al 2018).