Experimental approaches for the discovery and characterization of regulatory small RNA

Abstract

Following the pioneering screens for small regulatory RNAs (sRNAs) in Escherichia coli in 2001, sRNAs are now being identified in almost every branch of the eubacterial kingdom. Experimental strategies have become increasingly important for sRNA discovery, thanks to increased availability of tiling arrays and fast progress in the development of high-throughput cDNA sequencing (RNA-Seq). The new technologies also facilitate genome-wide discovery of potential target mRNAs by sRNA pulse-expression coupled to transcriptomics, and immunoprecipitation with RNA-binding proteins such as Hfq. Moreover, the staggering rate of new sRNAs demands mechanistic analysis of target regulation. We will also review the available toolbox for wet lab-based research, including in vivo and in vitro reporter systems, genetic methods and biochemical co-purification of sRNA interaction partners.