GSM7433534: AngII+Si-NAT10_RNA_1; Mus musculus; RNA-Seq
For acRIP-seq: Briefly, the concentration of total RNA was measured by Qubit RNA HS assay kit (Invitrogen, Q32852).   Total RNA was fragmented into 100-200 nt RNA fragments using ZnCl2.   Subsequently, the purified RNA was incubated with anti-ac4c polyclonal antibody  for 3 h at 4℃ and then with protein A/G magnetic beads (Thermo Fisher Scientific, 88802) at 4 ℃ for an additional 2 h to obtain immunoprecipitated RNA fragments.   The ac4c-enriched RNA was eluted from the beads in RLT buffer supplied in RNeasy Mini Kit (QIAGEN, 74106).   The RNA-seq library was prepared by NEBNext® Ultra™ II RNA Library Prep Kit for Illumina® (NEB, E7775).  For Ribo-seq:tissue were pulverized manually under liquid nitrogen and treated by ice-cold PBS containing 100 μg/mL cycloheximide.     Ribosome profiling was performed using Epi™ Ribosome Profiling Kit (Epibiotek, R1814).     Subsquently, RPFs (ribosome-protected RNA fragments) was extracted using RNA clean&ConcentratorTM-5 kit (ZYMO, R1016).     EpiTM RiboRNA Depletion Kit  (Epibiotek, R1805) was used for rRNA depletion.     Sequencing libraries were constructed using QIAseq miRNA Library kit (QIAGEN, 1103679). RNA libraries were prepared for sequencing using standard Illumina protocols acRIP-seq; Ribo-seq; RNA-seq