GSM7950267: NGN2 pre; Homo sapiens; RNA-Seq
For ParseBio, to obtain single cell suspension, induced neurons cultured in 12-well plates were washed with PBS and incubated with 300 μL accutase at 37°C for 5 min. Dissociation reaction were stopped with 700 μL PBS containing 0.04% BSA, and cells were gently trituated into single cell suspension. Cells were washed twice with 500 μL PBS containing 0.04% BSA before nuclei isolation. To isolate the nuclei from single cell suspension, cells were trituated ten times in 100 μL lysis buffer (10 mM Tris-Cl pH7.4, 3 mM NaCl, 10 mM MgCl2, 1% BSA, 0.1% Tween-20, 1 mM DTT, 0.2U/μL RNase inhibitor (Takara), 0.1% NP-40, 0.01% Digitonin (Invitrogen)) and incubate 2 min on ice. The isolated nuclei underwent two subsequent washes in 500 μL wash buffer (10 mM Tris-Cl pH7.4, 3 mM NaCl, 10 mM MgCl2, 1% BSA, 0.1% Tween-20, 1 mM DTT, 0.2U/μL RNase inhibitor). Isolated nuclei were fixed and cryopreserved for snRNA-seq according to the instructions provided by the manufacturer (Parse Biosciences).   For 10x Multiome, single-cell suspensions obtained from each NGN2-iN pre-patterning condition were divided into two distinct pools (Supplementary Table 1). Subsequently, each pool, along with the control NGN2-inducible iPSCs, underwent nuclei isolation individually following provider's instruction (10x Genomics user guide CG000365). Libraries were constructed according to the manufacter's instructions. For 10x Genomics user guide CG000338 was used.