title: GSM9307537: Induced cortical neurons, TDP-43 CRISPRi, replicate 1; Homo sapiens; RNA-Seq
library_construction_protocol: RNA was extracted and purified from cultured cells using the Direct-zol™ RNA Miniprep Plus (R2071, Zymo Research, Irvine, CA, USA). RNA concentration was determined with the Qubit RNA BR Assay Kit (Q10210, ThermoFisher Scientific, Waltham, MA, USA) and Qubit 4 Fluorometer (Q33226, ThermoFisher Scientific, Waltham, MA, USA). RNA quality was assessed using the RNA 6000 Nano (5067-1511, Agilent, Santa Clara, CA, USA) on the Agilent 2100 Bioanalyzer (G2939BA, Agilent, Santa Clara, CA, USA). CrypTE-seq read libraries were prepared from ~300ng total RNA per sample following a modified PacBio Iso-Seq v2 library protocol (103-071-500, PacBio, Menlo Park, CA, USA) and pooled using the SMRTbell Prep Kit 3.0 (102-182-700, PacBio, Menlo Park, CA, USA). Modifications include the use of post-primer annealing with first strand cDNA synthesis performed with MarathonRT (200035, RNAConnect, Branford, CT, USA) followed by Iso-seq RT and then addition of the template switch oligo. All other steps followed the Iso-seq v2 library protocol (103-071-500, PacBio, Menlo Park, CA, USA).