Upper limit for DNA packaging by Bacillus subtilis bacteriophage ϕ 105: Isolation of phage deletion mutants by induction of oversized prophages

Summary

We have determined the upper size limit for DNA packaging in Bacillus subtilis bacteriophage ϕ 105 by examining the plaque-forming and transducing capabilities of lysates made from strains containing prophages of various sizes. The upper size limit for efficient packaging of the phage genome appears to be about 40.2 kb, which is about 1 kb larger than the wild-type genome. This places an upper limit of about 5 kb on the size of insertions that can be accommodated in ϕ 105 transfection cloning vectors, such as ϕ 105J27. Induction of prophages that exceed the upper limit, followed by selection for plaque formation or transduction, provides a powerful means of isolating phage deletion mutants. A comparison of the location of each deletion with the resultant phenotype has enabled us to identify non-essential regions of the phage genome, and regions that are required for tail biosynthesis and for host cell lysis.