Structural analysis of microRNA-target interaction by sequential seed mutagenesis and stem-loop 3' RACE.

ABSTRACT:

Background

As a consequence of recent RNAseq efforts, miRNAomes of diverse tissues and species are available. However, most interactions between microRNAs and regulated mRNAs are still to be deciphered. While in silico analysis of microRNAs results in prediction of hundreds of potential targets, bona-fide interactions have to be verified e.g. by luciferase reporter assays using fused target sites as well as controls incorporating mutated seed sequences. The aim of this study was the development of a straightforward approach for sequential mutation of multiple target sites within a given 3' UTR.

Methodology/principal findings

The established protocol is based on Seed Mutagenesis Assembly PCR (SMAP) allowing for rapid identification of microRNA target sites. Based on the prese

SUBMITTER: Bohmer M 

PROVIDER: S-EPMC3839922 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature