The 80 λdilv phage and its use to detect messenger RNA related to isoleucine-valine metabolism

A 80λ defective phage containing the Escherichia coli K12 genes for the isoleucine-valine biosynthetic enzymes (ilv genes) was used to analyse the level of the specific messenger RNA related to isoleucine-valine metabolism. The mRNA extracted from polysomes of cells derepressed for the ilv genes because of leucine starvation hybridizes specifically to the l strand of 80λdilv. Very little mRNA is found in repressed cells; therefore most of the 80λdilv-specific mRNA is related to isoleucine-valine metabolism. Competition experiments show that, in cells derepressed either because of leucine starvation (five-fold derepressed), or because of the presence of the ilvO603 regulatory mutation (four-fold derepressed), the concentration of mRNA specific for the ilvADGE operon is about 25-fold higher than in repressed cells. We believe that the increase in mRNA concentration in the derepressed cells is due to an increased rate of transcriptional initiation. The mRNA extracted from cells in which the ilvC gene is specifically induced also hybridizes to the l strand of 80λdilv DNA. We conclude from this that the ilvC gene is transcribed in vivo on the same strand as the ilvADGE operon. We also report a detailed genetic map of 80λdilv. The ilv genes clustered at minute 75 on the E. coli map were all present on the phage, as well as the cya gene. The rbs genes appear to be partly present since we get recombination with a specific rbs allele, but no complementation. The bacterial genes substitute for part of the late genes in the left arm of 80λ DNA. Electron microscope analysis of the heteroduplex molecules formed from 80λdilv DNA confirms the genetic data. © 1975 Academic Press Inc. (London) Limited.