GSM6421790: ASC48_WT_nuc_RNA_rep2; Mus musculus; RNA-Seq
For total RNA-seq, 1ml Trizol(Invitogen) were added to cells and RNA were extracted according to manufacturer's protocol. For subcelluar RNA-seq, cells were treated with buffer A (HEPES-KOH 50 mM pH 7.5, 10 mM KCl, 350 mM sucrose, 1 mM EDTA, 1 mM DTT, 0.1% Triton X-100) for 5 min on ice and then homogenized with a T 10 basic ULTRA-TURRAX® homogenizer at 4th gear for 1min. The nuclei were harvested by brief centrifugation (2000g, 5 min), while the supernatant was collected as the cytoplasmic fraction. Nuclei pelltet were resuspended with same volume buffer A. Then 1ml Trizol were added to both nuclear and cytoplasmic fraction and subjected for RNA extracion. All RNA were subjected to polyA selection (Ambion, 61006) before library preparation. RNA libraries were prepared with about 1μg RNA according to the manufacturer's protocol (NEB UltraII RNA library kit for illumina). The libraries were sequenced in 2 × 150 nt manner on HiSeq X ten (Illumina).