title: Illumina NovaSeq 6000 paired end sequencing; RNA-Seq of WT HeLa Flp-In T-REx and UPF3A overexpression cell lines
design: RNA-Seq of WT HeLa Flp-In T-REx and UPF3A overexpression cell lines
library_construction_protocol: HeLa Flp-In-T-REx (established by Elena Dobrikova and Matthias Gromeier, Duke University Medical Center) WT and UPF3A overexpressing cells were maintained at 37 C, 5% CO2 and 90% humidity in Dubleccos Modified Eagle Medium (DMEM, Thermo Fisher Scientific) supplemented with 9% fetal bovine serum (FBS) and Penicillin-Streptomycin (both Thermo Fisher Scientific). To induce expression of the stably integrated UPF3A construct, 30 g/ml cumate was added and the cells were harvested after 48h. Total RNA was extracted using the Direct-zol RNA MiniPrep kit including the recommended DNase I treatment (Zymo Research) according to the manufacturer's instructions. Ribosomal depletion and strand specific library preparation was carried out with TruSeq Stranded Total RNA (with Ribo-Zero Gold) according to the manufacturer's instructions.