Nov 13, 2013

Validation of miRNA-mRNA interactions by electrophoretic mobility shift assays

BMC Research Notes
Anna SoléCarlos J Ciudad

Abstract

MicroRNAs are small non-coding RNAs involved in gene expression regulation by targeting specific regions in the 3'-UTR of the mRNA of their target genes. This binding leads to a decrease in the protein levels of such genes either by mRNA degradation or mRNA destabilization and translation inhibition. The interaction between a miRNA and its target mRNAs is usually studied by co-transfection of a reporter expression vector containing the 3'-UTR region of the mRNA and an inhibitory or precursor molecule for the miRNA. This approach, however, does not measure the direct and physical interaction between a miRNA and a specific mRNA. RNA molecules corresponding to miR-224 and to the 3'-UTR of SLC4A4 were incubated together and their interaction studied under different binding conditions using electrophoretic mobility shift assays. A direct and specific interaction between miR-224 and SLC4A4 mRNA was observed. This interaction was abolished in the presence of competitors. In this study, we explored a new application for the electrophoretic mobility shift assay and we demonstrated that it is a useful alternative method to assess, in a direct and specific manner, whether a miRNA binds to a specific predicted target mRNA.

  • References21
  • Citations4

Mentioned in this Paper

Ribonucleotides
SLC4A4 protein, human
MRNA Destabilization
Transfection
expression vector
RNA, Small Temporal
Binding (Molecular Function)
Metabolic Inhibition
Electrophoretic Mobility Shift Assay
MIRN224 microRNA, human

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