Artificial riboswitch selection: A FACS-based approach

Abstract

Riboswitches have a number of characteristics that make them ideal regulatory elements for a wide range of synthetic biology applications. To maximize their utility, methods are required to create custom riboswitches de novo or to modify existing riboswitches to suit specific experimental needs. This chapter describes such a method, which exploits fluorescence-activated cell sorting (FACS) to quickly and efficiently sort through large libraries of riboswitch-like sequences to identify those with the desired activity. Suggestions for the experimental setup are provided, along with detailed protocols for testing and optimizing FACS conditions FACS selection steps, and follow-up assays to identify and characterize individual riboswitches. © Springer Science+Business Media New York 2014.