GSM9450983: Stimulated Control Total RNA 3; Mus musculus; RNA-Seq
Cells were washed with PBS containing cycloheximide and lysed in polysome extraction buffer with triton. Incubated for 30 minutes on ice, vortexing every 5 to 10 minute and centrifuged for 15 minutes. Lysate was frozen as input or loaded onto a sucrose gradient to collect polysomal information. mRNA was isolated using oligo(dT)-attached magnetic beads. First-strand cDNA was synthesized with random hexamer primers, second-strand by incorporating dUTP instead of DTTP. Directional libraries prepared through end repair, A-tailing, adapter ligation, size selection, PCR amplification, and purification.