GSM8606709: U87-Osimertinib, replicate2, bulk-RNA-seq; Homo sapiens; RNA-Seq
Cells were harvested and lysed in 1 mL Trizol (Gibco, 15596018), followed by adding 0.2 mL CHCl3. Then solution was vortex for 15~30 s and set at room temperature for 2~3 minutes, followed by centrifugation for 15 minutes at 12,000g under 4°C. After centrifugation, the water layer was transferred into a new 1.5ml tube with 0.5 mL isopropanol (IPA) and set at -80°C overnight for RNA precipitation. The RNA pellet was obtained by centrifugation for 10 minutes at 12,000g under 4°C followed by removing the liquid. RNA pellet was washed using cold 75% ethanol, and was centrifuged for 5 minutes at 12,000g under 4°C. After removing liquid, RNA pellet was resuspended in 20ul DEPC-water for further experiments. The RNA-seq libraries were constructed using the using VAHTS Universal V6 RNA-seq Library Prep Kit for Illumina® according to manufacturer's instructions. Briefly, total RNA samples were enriched for eukaryotic mRNA by Oligo(dT) magnetic beads followed by fragmentation and cDNA synthesis and purification. Purified double-stranded cDNA was subjected to end repair, adaptor ligation, and PCR amplification to obtain cDNA libraries.