GSM7989587: hdSpO1; Homo sapiens; RNA-Seq
For the organoid dissociation, 4–5 randomly selected organoids were pooled to obtain single cell suspension. For assembloids, 1–2 assembloids were pooled for dissociation. To match the relative cell numbers across different regions, 1–2 organoids from each of the four regions, totaling 4–8 organoids, were pooled for dissociation only when comparing organoids to assembloids. For human primary tissue, 1–2 DRG were pooled for dissociation. Samples were incubated in 30 U/mL papain enzyme solution (Worthington Biochemical, LS003126) and 0.4% DNase (12,500 U/mL, Worthington Biochemical, LS2007) at 37°C for 45 minutes. After enzymatic dissociation, organoids were washed with a solution including protease inhibitor and gently triturated to achieve a single cell suspension. Cells were resuspended in 0.04% BSA/PBS (Millipore-Sigma, B6917-25MG) and filtered through a 70 μm Flowmi Cell Strainer (Bel-Art, H13680-0070). To target 7,000 cells after recovery, approximately 11,600 cells were loaded onto a Chromium Single Cell 3′chip (Chromium Next GEM Chip G Single Cell Kit, 10x Genomics, PN-1000127) with Chromium Single Cell 3' Library & Gel Bead kit v3.1 (10x Genomics, PN-1000128 and PN-1000269) and proceeded according to manufacturer's instructions.