GSM8214875: aNSC_ecMyc_WGS_1; Mus musculus; OTHER
Cells were lysed using cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Immediately after lysis, nuclei were spun at 500g for 10 min using a refrigerated centrifuge. Profiling of chromatin was performed by ATAC-Seq as described by Buenrostro, et al. Briefly, 50,000 fresh aNSCwere washed in cold PBS and lysed. The transposition reaction containing TDE1 Tagment DNA Enzyme (Illumina catalog # 20034198) was incubated at 37°C for 30 minutes. The DNA was cleaned with the MinElute PCR Purification Kit (QIAGEN catalog # 28004) and material was amplified for 5 cycles using NEBNext High-Fidelity 2X PCR Master Mix (New England Biolabs catalog # M0541L). After evaluation by real-time PCR, 4 to 5 additional PCR cycles were done. The final product was cleaned by aMPure XP beads (Beckman Coulter catalog # A63882) at a 1X ratio, and size selection was performed at a 1.5X ratio. Libraries were sequenced on a NovaSeq 6000 in a PE100 run, using NovaSeq 6000 S4 Reagent Kit v1.5 (200 Cycles) (Illumina). An average of 50 million paired reads were generated per sample.