Mu-specific properties of lambda phages containing both ends of Mu depend on the relative orientation of Mu end DNA fragments.

Abstract

Abstract To facilitate the study of bacteriophage Mu transposition we have constructed plaque-forming λ phages containing all the Mu genes and sites known to be required for normal levels of Mu transposition (i.e., Mu genes A and B and the two Mu DNA ends). These phages were made by in vitro recombination of Eco R1 DNA restriction fragments from two donor λ phages, each containing an Eco R1 fragment carrying one end of Mu. Theoretically the resulting phages with both ends of Mu could possess the two Mu-containing fragments in different orientations with respect to each other and to the lambda DNA, producing eight classes of hybrid phages. The results of agarose gel electrophoresis of DNA fragments produced by Hind III digestion of phage DNA allowed most phages to be assigned to one of the eight expected classes. Electron microscopic analysis of DNA heteroduplexes was used to confirm these classifications. Intact representatives were recovered for only six of the eight expected classes. Those phages containing the two Mu ends in closest proximity to one another were not recovered intact; however, deleted derivatives of this type of phage were found. The relative orientation of Mu-containing DNA fragments in phages from the six recovered classes was critical in determining the Mu-specific reactions which they carried out. All intact λ-Mu clones complemented Mu phages defective in genes A and B , but only those phages from the two classes with the Mu-containing fragments in their normal relative orientation complemented A -defective phage at a high level. Phages in the same two classes were also unique in their ability to carry out a second Mu-specific reaction which resulted in inhibition of their own growth.