library_construction_protocol: Cells from three organoid batches cultured for 14 weeks were thawed at 37°C and resuspended in HBSS to maintain singularization. The resulting cell suspension for each batch was passed through a dead cell removal kit (Miltenyi Biotec #130-090-10). Details on the specific kit components and protocols could be found on the 10x Genomics website. Following incubation within the GEMs, polyadenylated messenger RNA (mRNA) was reverse transcribed into full-length, barcoded cDNA. This cDNA was then amplified via PCR to achieve sufficient mass for library construction. The quality and quantity of the generated cDNA from each cell suspension were assessed using a fluorometric PicoGreen assay and capillary electrophoresis on an Agilent BioAnalyzer 2100. The cDNA generated from the cell suspensions was then converted into dual index libraries compatible with Illumina sequencers. This conversion utilized 10X Genomics 3' Gene Expression Library Construction kits (PN-1000190). The process involves a series of enzymatic steps: fragmentation of the cDNA, repair of the ends, addition of a poly-A tail, ligation of sequencing adapters, and finally, PCR amplification to generate sufficient library material. The final size distribution of the libraries was validated using capillary electrophoresis, and both the quantity (via fluorimetry with PicoGreen) and quality (via Q-PCR) were assessed.