GSM9819884: DASMC11 Control siRNA; Homo sapiens; RNA-Seq
Cells were collected 48-hours after treatment and lysed in TRI Reagent (Cat# T9424-100ML, Lot# MKCK9023, Sigma-Aldrich, Oakville, ON, CA). RNA was extracted using the Direct-zol RNA Miniprep kit (Cat# R2050, Zymo Research, California, USA) following the manufacturer's protocol. Samples were prepared for sequencing using the QuantSeq 3' FWD mRNA-Seq Library Prep Kit for Illumina (Lexogen, Austria), according to the manufacturers' protocol, and validated using Qubit 4 (Thermofisher Scientific) with 130ng RNA as the input. Successful libraries were pooled and diluted to an equimolar concentration resulting in a total pool of 4nM. Dilution and denaturation were performed according to Illumina protocols to result in a 2.2pM input (optimized for the Lexogen 3' kit). Single-end sequencing of 75 cycles was performed on an Illumina Nextseq550 using a High Output v2 reagent cartridge with each sample having a minimum depth of 5 million reads.