GSM4982800: abdominal aortic aneurysm tissue, IgG control_RIP-seq; Homo sapiens; RIP-Seq
RIP-seq was first performed using the Magna RIP™ RNA-binding Protein Immunoprecipitation Kit (Millipore, Massachusetts, USA), according to the manufacturer's protocol. Briefly, magnetic beads coated with 10 μg of specific antibodies against Aly/REF (Abcam, Cambridge, UK) or normal IgG (Millipore, Massachusetts, USA) were incubated with prepared indicated cell lysates overnight at 4 °C. Washed RNA-protein complexes were treated with proteinase K digestion buffer. The coprecipitated RNAs were purified with phenol: chloroform: isoamyl alcohol and subsequently subjected to purity and concentration analysis using NanoDrop 2000c. rRNAs were removed from the immunoprecipitated RNA and input RNA samples by using (New England Biolabs, Inc., Massachusetts, USA). RNA libraries were constructed by using rRNA-depleted RNAs with NEBNext® Ultra™ II Directional RNA Library Prep Kit (New England Biolabs, Inc., Massachusetts, USA) according to the manufacturer's instructions. Libraries were controlled for quality and quantified using the BioAnalyzer 2100 system (Agilent Technologies, Inc., USA).