Strategies to identify microRNA targets: new advances.

MicroRNAs (miRNAs) are small regulatory RNA molecules functioning to modulate gene expression at the post-transcriptional level, and playing an important role in many developmental and physiological processes. Ten thousand miRNAs have been discovered in various organisms. Although considerable progress has been made in computational methodology to identify miRNA targets, most predicted miRNA targets may be false positive. Due to the lack of effective tools to identify miRNA targets, the study of miRNAs is seriously retarded. In recent years, some molecular cloning strategies of miRNA targets have been developed, including RT-PCR using miRNAs as endogenous primers, labeled miRNA pull-down assay (LAMP) and RNA ligase-mediated amplification of cDNA end (RLM-RACE). The identified miRNA targets should be further validated via effects of miRNA alteration on the target protein levels and bioactivity. This review summarizes advances in strategies to identify miRNA targets and methods by which miRNA targets are validated.