GSM8119085: control/hifu, pooled replicate 2 [B1]; Homo sapiens; RNA-Seq
Organoids were dissociated for single-cell RNA sequencing as described with minor modifications106. In brief, organoids were incubated in Papain (Worthington LK003176) with DNase1 (Worthington LK003170) reconstituted per manufacturer instructions for 60 minutes at 37⁰C with gentle shaking at 27 rpm with regular pipetting. Single-cell suspension was added to Earle's Balanced Salt Solution (EBSS) with Ovomucoid Inhibitor (Worthington LK003182) and DNase1, pelleted, resuspended in ice-cold Neurobasal-A (Invitrogen) with 0.2% bovine serum albumin (BSA), and filtered through 40 µM Flowmi tips (Sigma BAH136800040-50EA). Cell count and viability was determined by Trypan Blue staining using a hemocytometer. Cells were labeled with hashtag-oligonucleotide (HTO) antibodies (Biolegend TotalSeq-B, 405287, 405289, 405291) and libraries were prepared following manufacturer protocols (10X Genomics, CG000206 Rev D). Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (with Feature Barcoding for Cell Surface Protein); PN-1000128, PN-1000079, PN-1000127, PN-1000213