miR-27b directly targets MSTN. (A) Sequence alignment of putative miR-27b-binding site in the MSTN 3′-UTR, showing a high level of complementarity and sequence conservation among mammalian species. The wild type bovine MSTN mRNA sequence is shown with potential binding sites underlined. (B) Luciferase assay was performed to test whether MSTN is a bona fide target of miR-27b. 293 T cells were co-transfected with psi-check2 reporter vectors containing native or mutated MSTN 3′-UTR, and miR-27b mimics or miRNA mimics negative control (miR-142), followed by cell lysis and luciferase assays 48 h later. Mutation of the miR-27b recognition sequence abolished responsiveness to miR-27b co-transfection compared with the wild-type construct. Psi-check2 was used as negative controls. * means values with significant difference (at least p < 0.001).
Miretti et al. BMC Genomics 2013 14:194 doi:10.1186/1471-2164-14-194