Analysis of Bacillus subtilis Ribonuclease Activity In Vivo.

Ribonucleases remodel RNAs to render them functional or to initiate their degradation. Our laboratory studies these pathways in detail using a variety of techniques, ranging from the isolation of RNAs in various RNase mutants to determine their implication in maturation or decay pathways by Northern blot, to proving their direct roles in RNA cleavage reactions using purified enzymes and in vitro-transcribed RNA. In this chapter, we provide in-depth protocols for the techniques we regularly use in the laboratory to assay RNase activity in vivo, with detailed notes on optimization of these methods. This chapter complements Chap. 25 on assays of ribonuclease action in vitro.