Decay of mRNA in Escherichia coli: specific segments of transcripts (bla/ompA/RNA half-life/transcriptional termination/SI nuclease)

An assay was developed to investigate the fate of specific segments of i-lactamase (bla) and ompA gene transcripts in Escherichia coli. DNA probes cloned in bacteriophage M13 were treated with an endonuclease capable of cleaving single- stranded DNA, the fragments produced were annealed with total cellular RNA, and the resulting RNA-DNA hybrids were subjected to SI nuclease treatment and'gel fractionation. By using this assay, direct evidence was obtained for 3'-to-5' directionality in the decay of the long-lived,mRNA encoded by the ompA gene, and no pref- erential stability was observed for translated versus untranslated mRNA segments. In the case of bla mRNA, initial cleavage of the full-length transcript was rate limiting, and no decay intermedi- ates were detected. No difference in.degradation rate was seen for bla transcripts having variant 3' or 5' termini.