GSM2528069: TT-seq_Raji-CDK9as_1NAPP1_rep2; Homo sapiens; RNA-Seq
Cells were harvested by centrifugation at 3,000 x g for 2 min. Total RNA was extracted using QIAzol according to the manufacturer's instructions. RNAs were sonicated to generate fragments of <1.5 kb using AFAmicro tubes in a S220 Focused-ultrasonicator (Covaris). 4sU-labeled RNA was purified from 150 µg total fragmented RNA. Separation of labeled RNA was performed using streptavidin beads (Miltenyi Biotec). Prior to library preparation, 4sU-labeled RNA was purified and quantified. Enrichment of 4sU-labeled RNA was analyzed by RT-qPCR as described before in (Schwalb et al., 2016). Input RNA was treated with HL-dsDNase (ArcticZymes) and used for strand-specific library preparation according to the Ovation Universal RNA-Seq System (NuGEN). The size-selected and pre-amplified fragments were analyzed on a Fragment Analyzer before clustering and sequencing on the Illumina HiSeq 1500. NuGEN Ovation Universal RNA-Seq System, starting with 100 ng RNA input
GEO Accession
GSM2528069