RNA visualization in bacteria by fluorescence in situ hybridization

Abstract

Detecting localized RNA in bacteria is difficult due to the properties of RNA and the small size of the cell. Fluorescence in situ hybridization (FISH) has been an invaluable method for detecting and imaging RNA. In FISH, RNA is fixed in its native subcellular position through chemical cross-linking. An oligonucleotide probe conjugated to a fluorophore is annealed to the target RNA, and the target RNA/probe hybrid is visualized using fluorescence microscopy. This chapter describes the use of FISH to visualize tmRNA, a regulatory RNA required for trans-translation. The method can be adapted to visualize the localization of other regulatory and messenger RNAs as well. © 2012 Springer Science+Business Media, LLC.