GSM7849056: HEK293 cells, PKA Cα, doxycycline, biol rep 1; Homo sapiens; RNA-Seq
After treatment, the cells were washed with PBS, total RNA was isolated with a RNeasy Mini Kit (74104, Qiagen) including an on-column DNase I digestion and quantified using a Nanodrop ND-1000 (Thermo Scientific). Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers followed by the second strand cDNA synthesis. The library was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries were pooled and sequenced on Illumina Novaseq 6000 platform, according to effective library concentration and data amount.