GSM6604804: 56093_sCJD; Homo sapiens; RNA-Seq
Frozen frontal cortex brain slices were left to thaw and were hand-dissected. Approximately 50-100 mg of the superior frontal gyrus was homogenised in Tri Reagent (Zymo Research) using a pellet pestle (Kimble) and incubated at room temperature for 2 hours. Total RNA was purified using the Direct-zol RNA miniprep kit (R2050; Zymo Research) following the manufacturer's instructions. DNAse I treatment was performed in tube followed by column purification (Zymo Research). All RNA samples were subsequently run on a TapeStation 2200 (Agilent, Santa Clara, US) using High Sensitivity RNA ScreenTapes. 100 ng of Total RNA was used as input material to pepare librairies using the Universal Plus Total RNA-Seq with Human AnyDeplete (Tecan) according to the manufacturer's instructions. Resulting libraries were visualized and quantified on the 2200 TapeStation Instrument and diluted at 2nM concentration and pooled in two batches of 7 libraries each. Each pool was sequenced on the Illumina NextSeq 500 after the addition of 5% PhiX sequencing control library (FC-110-3002; Illumina) using the following settings: paired-end, read 1: 75 nt, read 2: 75 nt.