GSM6454745: KHRNABCSK_0007 [24013470]; Homo sapiens; RNA-Seq
RNA was extracted from archival FFPE tissue using a modified QIAGEN Rneasy® FFPE kit protocol. Briefly, prior to deparaffinization of the FFPE tissue, the samples are incubated with 1X acidic antigen retrieval solution at 90C for 5 minutes. Following incubation, samples are cooled to room temperature, and any excess paraffin is removed from the tube. We then proceeded through the standard kit protocol. RNA yield was quantified using the Qubit® RNA Broad Range kit and Qubit® 4.0 fluorometer. For RNA sequencing, 100 ng of total RNA molecules were used to construct libraries using Illumina TruSeq RNA Exome Library Prep Kit, following manufacturer's protocols. The final libraries were then quantified using Agilent D1000 Screen Tape as well as sequenced on Illumina MiSeq V2 Micro Kit to assess insert sizes and integrity before sequencing on a high-throughput sequencer. Each library was normalized to 4 nM and pooled and sequenced on Illumina NextSeq500 High Output Kit (Illumina, San Diego, CA).