Guidelines on Designing MicroRNA Sponges: From Construction to Stable Cell Line.
Single microRNA (miRNA) can be inhibited using antagomiR which efficiently knockdown a specific miRNA. However, the effect is transient and often results in subtle phenotype. Here we report a guideline on designing miRNA sponge inhibiting a miRNA family. As a model system, we targeted miR-30 family, known as tumor suppressor miRNAs in multiple tumors. To achieve an efficient knockdown, we generated perfect and bulged-matched miRNA binding sites (MBS) and introduced multiple copies of MBS. The protocol here demonstrates the miRNA sponge as a useful tool to examine the functional impact of inhibition miRNAs.
Post Graduate Program in Health Science, São Francisco University, Av São Francisco de Assis, 218 - Jardim São José, Bragança Paulista, 12916-900, São Paulo, Brazil. manoela.ortega@usf.edu.br.
Manoela Marques
MM
Ortega
Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX, USA.
Hakim
H
Bouamar
D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D003062:Codon; D055785:Gene Knockdown Techniques; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*
D003062:Codon; D035683:MicroRNAs
Vectors construction; miR-30a inhibition; microRNAs (miRNA) sponges