Antisense RNA does not significantly affect expression of the galK gene of Escherichia coli or the N gene of coliphage lambda.

The effect of antisense RNA on the expression of genes galK and N was studied in vivo. These two genes were either present in the Escherichia coli chromosome, as single copies, or were cloned on plasmid vectors. Antisense RNA was supplied from multicopy vectors where the entire galK or N gene, or only their N-proximal portions, were cloned in the antisense orientation downstream from the strong PL, PR or lacZp promoters. In all of the experiments there was no significant inhibition of the galK or N expression by up to a 50-fold excess of the specific antisense RNAs, for both the in cis and in trans experimental designs. The excess of the antisense RNA was calculated as based on respective copy numbers, but was not experimentally measured. The apparent five-fold regulatory effect observed in one of the experiments was found to be artifactually caused by unexpected creation of a terminator in one of our constructs. To avoid such artifacts, all our constructs were equipped with the nut-N antitermination system. We conclude that the reported antimessenger-mediated inhibition of gene expression is not a general phenomenon, but must require some special features which are not present in the galK and N systems.