GSM9371783: RNA-seq of human iPSC-derived NPCs, EGFP control, biol rep 4; Homo sapiens; RNA-Seq
For RNA-seq: Total RNA was isolated from primary murine NPCs, Ns, and human-derived Ns using Trizol (Sigma-Aldrich) according to the manufacturer's protocol. For CUT&Tag: Nuclei from 100,000 cells were isolated and immobilized on Concanavalin A-coated beads. They were then incubated overnight at 4°C with primary antibodies (anti-MeCP2, anti-V5, or anti-Smarcb1). Following incubation with a secondary antibody, pA-Tn5 transposase was applied to fragment and tag antibody-bound DNA sites. For 4f-SAMMY-seq: 200,000 cells were subjected to DNase I treatment to fragment DNA based on chromatin accessibility. DNA was then fractionated to separate the DNase-sensitive euchromatin fraction (S2S) from the DNase-resistant heterochromatin fraction (S3). For RNA-seq: Library preparation and paired-end sequencing were performed by a commercial service (Genewiz). For CUT&Tag: Following tagmentation, amplicon libraries were generated using the Nextera® DNA Library Prep Kit (Illumina). Barcoded libraries were pooled and sequenced on the NovaSeq 6000 platform. For 4f-SAMMY-seq: Libraries from S2S and S3 fractions were generated using the NEBNext Ultra II DNA Library Prep Kit for Illumina and Unique Dual Index NEBNextMultiplex Oligos. Libraries were pooled and sequenced on an Illumina NextSeq 2000 instrument.