Rneasy 74106

To isolate RNA from MEFs and pancreatic organoids, we used TRIzol (Thermo Fisher Scientific, #15596018) according to the manufacturer’s instructions, and contaminating DNA was removed by DNase treatment for 10 minutes and column purification (Qiagen RNeasy #74106). Pancreas tissue portion for RNA purification was consistently collected from the tail of the organ and immediately cut into smaller pieces and immersed in RNAlater stabilization solution (Thermo Fisher) and incubate at 4°C overnight before storing the sample at −80 °C until RNA extraction was performed. Samples were homogenized using a Tissue Master 125 (Omni) and RNA purified using the RNAeasy Kit (Qiagen).

For probe synthesis, a total of 1 μg of linearized plasmid DNA was transcribed in vitro using either SP6 or T7 RNA polymerase (Promega) using RNA DIG labeling mix (Roche). The probe was cleaned subsequently through a column (Qiagen RNEasy 74106) and dissolved in diethylpyrocarbonate (DEPC)-treated water containing 50% v/v formamide.^{40 (link)}In situ hybridisation was performed as described previously.⁴¹