title: RNA-Seq of mus musculus: Nf1/Mll3 mutant glioblastoma cells w/ TMZ
design: For RNAseq samples preparation, cell lines harboring specific gene knockouts were cultured for 7-9 days under the selection pressure of puromycin, and then plated into 6-well plates at a density of 2 x 105 cells per well in triplicates. 24 h after plating, 1 mM TMZ or DMSO in fresh DEME + 10% FBS was added and cultured for another 48 h. Then, cellular RNA of control or treated cells were extracted by adding 350 킠 TRIzol Reagent (Invitrogen) directly into 6-well plates to lyse the cells, followed by gently shaking the plates and incubation for 5-10 min to complete and lysis and homogenization. Then, 70 킠 chloroform was added, vigorously mixed, and centrifuged at 16,000 g for 15 min. Transfer the RNA containing aqueous phase to a new tube and further purified using RNeasy mini kit (Qiagen). After eluting RNA from column using Nuclease free water, the concentrations of sample RNA were normalized into 150-300 ng/uL for RNAseq.
library_name: Nf1+Mll3_TMZ_3