Abstract
Stable RNA, mainly comprised of rRNA and tRNA, accounts for the majority of cellular RNA. Although normally stable under favorable growth conditions in the laboratory, these RNA species undergo extensive degradation responding to many environmental changes and stress conditions. Multiple ribonucleases and other enzymes may be involved in the decay of stable RNA. The onset and rate of degradation are probably determined by the status of the RNA as well as the availability of the degrading activities. The elucidation of pathways for stable RNA decay has been benefited by many biochemical and genetic approaches. These include purification of the enzymes and characterization of their substrate specificity in vitro, and studies of stable RNA decay by inactivating and overexpressing the degradation activities in vivo. Furthermore, RNA degradation intermediates have been characterized in detail, such as determining the sizes, the sequences, the 5'- and 3'-termini, etc. In this work, we describe the methods that are most commonly used in the study of the degradation and processing of stable RNA in E. coli. Most of them should be also useful in studies of other RNA species or RNA from other organisms.