Premature neural differentiation in cerebral organoids derived from early-onset autism individuals in a birth cohort

Diversity clinical phenotypes of autism spectrum disorders (ASD), caused by heterogeneity of genetic and environmental factors, hampered the investigation in uncovering pathological mechanisms. Here we generated ASD patient-cerebral organoids from induced pluripotent stem cells (iPSCs) that reprogramed from the early-onset ASD individuals at around 2 years old with common clinical diagnosis in a prospective birth cohort, which was excluded typical autistic mutations and environmental exposures. Overall design: The organoids were dissociated by trypLE (Life technology) into single cells. The cell debris and other aggregates were removed by an additional low-speed centrifugation. About 12,000 cells were captured and the released RNA were barcoded through reverse transcription in individual GEMs. Reverse transcription was performed on a S1000TM Touch Thermal Cycler (Bio Rad) at 53 degrees Celsius for 45 min, followed by 85 degrees Celsius for 5 min and final hold at 4 degrees Celsius. cDNA was generated and then amplified, and quality was assessed using an Agilent2100. scRNA-seq libraries were prepared with the Chromium Single Cell 3' Library & Gel Bead Kit V3 (10x Genomics, 1000075) and then sequenced on an Illumina NovaSeq6000 with a sequencing depth of at least 50000 reads per cell with a paired-end 150 bp (PE150) reading strategy.