GSM2875751: Cor4U_5Gy_2; Homo sapiens; RNA-Seq
Cells were washed two times with phosphate buffered saline and centrifuged at 6,000 rpm for 2 min after incubation for additional 48 h prior to RNA isolation. Isolation was performed with the RNeasy Mini kit (Qiagen, RNeasy Mini Handbook 06/2012) following the manufacturer's protocol. RNA concentration was determined by means of a Qubit 3.0 Fluorometer (Thermo Scientific, Waltham, MA, USA) and quality was assessed using Bioanalyzer (Agilent 2100 Bioanalyzer, Agilent, Santa Clara, CA, USA). Library preparation of RNA samples was performed with the NEBNext® Ultra II Directional RNA Library Kit for Illumina (rRNA Depletion Kit, New England Biolabs Inc., Ipswich, MA, USA, protocol version 1.0 4/17). 500 ng RNA was used as input. In brief, rRNA was depleted and cDNA synthesis was executed before adapter ligation using appropriate primer design (NEBNext® Multiplex Oligos for Illumina® (Index Primers Set 1), Ipswich, MA, USA, protocol 2016). DNA fragment length was determined with Agilent Bioanalyzer (Agilent 2100 Bioanalyzer, Agilent, Santa Clara, CA, USA) and concentration was measured with a Qubit 3.0 Fluorometer (Thermo Scientific, Waltham, MA, USA).