GSM4083094: PKC activator; Homo sapiens; RNA-Seq
Total RNA was isolated with AllPrep DNA/RNA Mini Kit (Qiagen, Mississauga, ON, Canada) according to the manufacturer's instructions. Total RNA integrity and concentration were evaluated on a 2100-Bioanalyzer (Agilent Technologies, Palo Alto, CA). For all samples RNA Integrity Number was 10. The total RNA input used for sequencing was 270 ng for each treatment, a pool of 90 ng for each biological replicates. mRNA was isolated from total RNA using NEBNext Poly (A) mRNA Magnetic Isolation Module (E7490S; NEB). PolyA-selected mRNA was fragmented to a mean size of 200 nt, reverse transcribed to generate double-stranded cDNA, and converted to a paired-end library using NEBNext Ultra RNA Library Prep kit for Illumina (E7530S; NEB) according to the manufacturer's instructions, using Agencourt AMPure XP beads and NEBNext Mutliplex Oligos for Illumina (set1, E7335S; NEB).