title: GSM8442970: RKO-dox-Cas9, sgAAVS1, nucleus, CHX treated, biol rep 2; Homo sapiens; RNA-Seq
library_construction_protocol: For subcellular fractionation, ~2x10^6 cells were gently lysed in REAP buffer (0.1% NP-40 in 1x PBS) at 4°C. Nuclei and cytoplasmic fractions were separated through centrifugation at 3k g for 60 sec at 4°C twice. RNA isolation was done with High Performance RNA Isolation kit (Vienna, VBC Molecular Tools Shop, RNAisomag) and a KingFisher Flex robot (Thermo Fisher Scientific). Poly-A enriched RNA-seq libraries using NEBNext Ultra II RNA Library Prep Kit for Illumina (New England Biolabs) were generated according to manufacturer's recommendations.