bulk RNA-seq NOHA
bulk RNA-seq NOHA
Neural organoids were harvested at DIV 50 into 2ml Eppendorf tubes and washed three times with 1.5 ml PBS to remove medium residues before being snap-frozen and stored at -80 Â°C for a maximum of 1 month before RNA extraction. For each of the replicates, 1 organoid was collected unless size constraints required the pooling of multiple organoids  After thawing the pellets on ice, RNA was extracted using the PureLink RNA Mini kit (Thermofisher, 12183025) and eluted in 30ul of RNAse free water. On-column DNAse I treatment was performed with RNAse-free DNase set (Qiagen, 79265) according to protocol recommendations. Purified RNA was quantified with Nanodrop One (Thermofisher) to estimate the concentration range, while the RNA Integrity Index (RIN) was measured by TapeStation analysis. A RIN >7.5 (scale 1-10) was chosen as a cutoff for further sample processing. Samples with RIN <7.5 were discarded and RNA was re-extracted from backup pellets.  RNA concentration was then normalized, and cDNA libraries were prepared using Illumina Stranded mRNA Prep with polyA-capture technology with an initial RNA input of 100 ng/sample