Protocol

Artificial Riboswitches

Volume 1111 of the series Methods in Molecular Biology pp 259-267

Date:

Construction of Ligand-Responsive MicroRNAs that Operate Through Inhibition of Drosha Processing

  • Chase L. BeiselAffiliated withDepartment of Chemical and Biomolecular Engineering, North Carolina State University
  • , Ryan J. BloomAffiliated withDepartment of Bioengineering, Stanford University
  • , Christina D. SmolkeAffiliated withDepartment of Bioengineering, Stanford University

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Abstract

MicroRNAs (miRNAs) offer powerful tools for targeted gene silencing in almost all eukaryotes. These tools have received considerable attention for their utility in both fundamental genetic studies and as therapeutic agents. Rendering individual microRNAs responsive to endogenous or exogenously applied molecules (or ligands) can improve the stringency of silencing and can mediate autonomous control. This chapter describes the construction of ligand-responsive miRNAs that undergo reduced processing and subsequent gene silencing when bound by the recognized ligand. Following a simple set of rules, the engineered microRNAs can be readily modified to target different sequences and to bind different ligands. Individual miRNAs also can be incorporated into the same transcript for tunable, multi-gene silencing.

Key words

Aptamers Drosha Gene regulation Mammalian cells MicroRNA Riboswitch RNA interference