Validation of miRNA-mRNA interactions by electrophoretic mobility shift assays

BMC Research Notes

Carlos J Ciudad¹  Véronique Noé¹  Xenia Villalobos¹  Núria Mencia¹  Anna Solé¹ 

[1] Department of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona, Avenue Diagonal 643, Barcelona E-08028, Spain

关键词: EMSA;    Target validation;    miRNA;    Binding assay;    3′-UTR;   

Others  :  1140872
DOI  :  10.1186/1756-0500-6-454

 received in 2013-06-20, accepted in 2013-11-08,  发布年份 2013

【 摘 要 】

Background

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.

Findings

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.

Conclusions

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.

【 授权许可】

   
2013 Solé et al.; licensee BioMed Central Ltd.

【 图 表 】

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