Comparing two approaches of miR-34a target identification, biotinylated-miRNA pulldown vs miRNA overexpression. Issue 1 (2nd January 2018)

• Record Type: Journal Article
• Authors: Awan, Hassaan Mehboob; Shah, Abdullah; Rashid, Farooq; Wei, Shuai; Chen, Liang; Shan, Ge
• Is Part Of: RNA biology. Volume 15:Issue 1(2018)
• Page Start: 55
• Page End: 61
• Publication Date: 2018-01-02
• Subjects: Biotinylated miRNA -- Argonaut 2 -- microRNA targets -- RNA pull down -- RNA-seq; RNA -- Periodicals; Molecular biology -- Periodicals; Molecular biology; RNA; Periodicals; 572.8805
• DOI: 10.1080/15476286.2017.1391441
• Languages: English
• ISSNs: 1547-6286

Abstract

ABSTRACT: microRNAs (miRNAs) are critical regulators of gene expression. For elucidating functional roles of miRNAs, it is critical to identify their direct targets. There are debates about whether pulldown of biotinylated miRNA mimics can be used to identify miRNA targets or not. Here we show that biotin-labelled miR-34a can be loaded to AGO2, and AGO2 immunoprecipitation can pulldown biotinylated miR-34a (Bio-miR pulldown). RNA-sequencing (RNA-seq) of the Bio-miR pulldown RNAs efficiently identified miR-34a mRNA targets, which could be verified with luciferase assays. In contrast to the approach of Bio-miR pulldown, RNA-seq of miR-34a overexpression samples had limited value in identifying direct targets of miR-34a. It seems that pulldown of 3′-Biotin-tagged miRNA can identify bona fide microRNA targets at least for miR-34a.