MeRIP-Seq of Blood Vessel
Library selection was performed by immunoprecipitation with anti-mA antibody to enrich mA-methylated RNA fragments.The RNA m6A was sequenced by MeRIP-seq at Novogene (Beijing, China). Briefly, atotal of 2 g RNAs were extracted from the VSMCs. The integrity and concentration of extracted RNAs were detected using an Agilent 2100 bioanalyzer (Agilent) and simpliNano spectrophotometer (GE Healthcare), respectively. Fragmented RNA (~100 nt) was incubated for 2 hr at 4 with anti-m6A polyclonal antibody (Merk Millipore) in the immunoprecipitation experiment. Then, immunoprecipitated RNAs or Input was used for library construction with Ovation SoLo RNA-Seq System Core Kit (NuGEN). The library preparations were sequenced on an Illumina Novaseq or Hiseq platformwith a paired-end read length of 150 bp according to the standard protocols.