library_construction_protocol: Two separate batches of sample collection were performed for this project. In the first batch, 16 organoids were collected on day 22 of differentiation, including one isogenic control organoids, one severe XLAS organoid, and one moderate XLAS organoid, resulting in three libraries. An additional isogenic control was harvested on day 22 (Control_d22_2) and kept in culture for 38 days (Control_d38) and harvested at this later time point (two library, 5 libraries in total). Organoids were dissociated in two rounds of 10 minutes at 37°C with 500 µL of TrypLE. During each round, the tubes were gently flicked every five minutes for homogenization. At the beginning of each round, mechanical dissociation was performed by pipetting up and down five times. After a quick centrifugation, the supernatant was aspirated and inactivated in PBS with 10% SVF on ice. For the second round, 200 µL of TrypLE was added to the remaining organoids. The final suspension was filtered through a 40 µm cell strainer before pelleting the dissociated cells at 300g for 3 minutes. Cells were then resuspended in 1 mL of PBS containing 1% BSA and 1 mM EDTA and kept on ice. Cell viability, as assessed by cell counting, ranged between 70% and 85%. Libraries obtained from these two batches were sequenced together. Libraries were generated using Chromium Next GEM Single Cell 3' and Next GEM Chip G Single Cell Kits (10x Genomics) at the genomics platform at the Institute du Cerveau. The quality of the libraries was assessed using the High Sensitivity D5000 ScreenTape Assay for TapeStation Systems (Agilent), and the libraries were sequenced on NovaSeq X using the NovaSeq X plus 10B Reagent Kit (100 cycles). The minimum requested coverage was 50,000 reads per cell.