GSM9376790: Sham.Number-2; Mus musculus; RNA-Seq
Total RNA was extracted from cerebral ischemic penumbra tissue of the ipsilateral cortex using TRIzol reagent. RNA integrity and quantity were precisely assessed using an Agilent 2100 Bioanalyzer. For eukaryotic samples, poly(A) mRNA was enriched using Oligo(dT) magnetic beads. The enriched mRNA was fragmented and used as a template for first-strand cDNA synthesis with random hexamer primers. Second-strand cDNA was then synthesized using buffer, dNTPs, DNA polymerase I, and RNase H. Double-stranded cDNA was purified using AMPure XP beads, followed by end repair, A-tailing, and sequencing adapter ligation. Size selection was performed with AMPure XP beads, and final libraries were amplified by PCR. The resulting libraries were validated for insert size using an AATI analyzer and quantified by Q-PCR for accurate concentration measurement prior to sequencing.