GSM8513080: B16F10_S127A3; Mus musculus; RNA-Seq
Total RNA was extracted from normal or mutated B16F10 cells using TRIzol reagent (15596018CN, Invitrogen). Total RNA was quantified using a NanoDrop spectrophotometer and an Agilent 2100 bioanalyzer (Thermo Fisher Scientific). RNA library construction and subsequent RNA sequencing were performed using BGI-Shenzhen (Shenzhen, China) software. First-strand cDNA was generated by random hexamer-primed reverse transcription followed by second-strand cDNA synthesis. Afterwards, A-Tailing Mix and RNA Index Adapters were added by incubation for end repair. The cDNA fragments obtained from the previous step were amplified by PCR and the products were purified using Ampure XP Beads and dissolved in an EB solution. The product was validated using an Agilent Technologies 2100 Bioanalyzer for quality control. Double-stranded PCR products from the previous step were denatured and circularized using splint oligo sequences to obtain the final library. Single-stranded circular DNA used as the final library.