Filamentous phage infection-mediated gene expression: construction and propagation of the gIII deletion mutant helper phage R408d3

Abstract

We describe the use of transcriptional fusions to the phage shock protein ( psp) promoter. These fusions are expressed only when cells are infected by filamentous phage. In an application, the psp promoter was fused to the protein coding part of filamentous phage gene I I I ( gIII). Protein III (pIII) is needed to complement mutant f1 phage containing a deletion of gIII, but its synthesis also renders cells resistant to infection. By inducing pIII production from psp– gIII only in the cells that are already infected with phage, it was possible to obtain plaques from phage in which gIII had been completely deleted. gIII was deleted from two helper phages: R408 and VCSM13, which were then propagated on cells containing the psp– gIII fusion. These two phages were tested for use in a phage display method that requires generation of noninfectious, phagemid-containing virion-like particles. Both helpers worked, but R408d3 was superior to VCSM13d3, because it generated about 1800-times fewer background infectious particles.