GSM9308936: 230852-RN01; Homo sapiens; RNA-Seq
RNA was extracted by Qiagen RNeasy Mini Kit (Cat. 74106) (Qiagen Inc.). The extracted RNA was treated with DNase I to get rid of nay genomic DNA residues. The treated RNA then was cleaned-up using the AMPure XP beads (Beckman Coulter Life Sciences) and eluted into 1x TE buffer. The purified RNA was quantified using Quant-iT™ RiboGreen™ RNA Assay Kit (ThermoFisher SCIENTIFIC) and the RNA quality was accessed using Agilent RNA 6000 Nano Kit and the 2100 Bioanalyzer Instrument (Agilent Technologies). The fragmented cDNA was proceeded to library preparation using SureSelect XT Low Input Reagent Kit with indexes 1-96 (Cat: G9703A, Agilent Technologies) and automated on the Sciclone G3 NGSx Workstation (PerkinElmer, Inc.). This protocol consists of 3 enzymatic reactions for end repair, A-tailing and Adaptor ligation, followed by barcode insertion by PCR using Herculase II Fusion DNA Polymerase (8 to 14 cycles, based on input DNA quality and quantity). PCR primers were removed by using 1x volume of Agencourt AMPure PCR Purification kit (Agencourt Bioscience Corporation). The quality and quantity of the prepared libraries were evaluated using TapeStation 4200 and the DNA High Sensitivity kit (Agilent Technologies) to verify correct fragment size and to ensure complete removal of primer dimers.