This chapter provides detailed methodologies for isolating total RNA and polyadenylated RNA from E. coli and other prokaryotes, along with the procedures necessary to analyze the processing and decay of specific transcripts and determine their 3'- and 5'-ends. The RNA isolation methods described here facilitate isolating good-quality RNA in a very cost-effective way compared to the commercially available RNA isolation kits, without employing phenol and/or alcohol precipitation. We also discuss the limits associated with polyacrylamide and agarose gels for the separation of small and large RNAs. Methods useful for the analysis of post-transcriptionally modified transcripts and the processing of very large polycistronic transcripts are also presented.