A polymerase chain reaction-based cloning strategy applicable to functional microRNA studies

Abstract MicroRNAs (miRNAs) are key regulatory RNAs that act in concert to coordinately control messenger RNA translation through imperfect recognition of multiple specific binding sites (BSs) located in their 3′ untranslated region. Here, we present a polymerase chain reaction-based cloning strategy that allows the rapid and efficient generation of regulatory elements harboring up to 10 miRNA BSs. Amenable for the study of regulatory elements of any multiplicity, such as those recognized by miRNAs and transcription factors, this methodology will facilitate functional miRNA/miRNA BS studies and accelerate discoveries mainly in the field of gene regulation.