title: Dual luciferase gene reporter assays to study miRNA function.
abstract: This chapter describes one of the most reliable quantitative assays to test the silencing of a possible target gene by a specific miRNA using a luciferase reporter gene. The experimental procedure first consists in cloning both the wild-type and mutated forms of the 3'UTR of the miRNA predicted mRNA target downstream of a firefly luciferase reporter. Next, each construct is co-transfected together with the miRNA into HeLa cells, and the reporter expression is monitored. Changes in luciferase levels will indicate whether or not a miRNA can bind to the UTR and regulate its expression.
authors: CNRS UMR 7365, IMoPA, Université de Lorraine, 9 avenue de la Forêt de Haye, Vandoeuvre-lès-Nancy, 54506, France.
authors: Thomas
authors: T
authors: Clément
authors: Véronique
authors: V
authors: Salone
authors: Mathieu
authors: M
authors: Rederstorff
mesh_terms: D003001:Cloning, Molecular; D020868:Gene Silencing / Q000502:physiology*; D017930:Genes, Reporter / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D008163:Luminescent Measurements / Q000379:methods; D035683:MicroRNAs / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D020506:Untranslated Regions / Q000235:genetics*
chemical_list: D035683:MicroRNAs; D012333:RNA, Messenger; D020506:Untranslated Regions; D008156:Luciferases