The phage-displayed peptide library vector M13KE was derived from the well-known cloning vector M13mp19. Consequently, it contains a lacZα insert near its origin of replication, which significantly decreases the propagation rate of the phage. We have previously reported thirteen compensatory mutations in the 5’-untranslated region of gene II, which effectively allow the phage to resume a wild-type propagation rate. Recently, we have discovered two additional types of genetic change that increase the propagation rate of M13KE: (i) a point mutation in gene IV that reverts back to the nucleotide found in wild-type M13, and (ii) an ejection of the lacZα insertion. Phage propagation assays confirm that both these changes also compensate for the lacZα insertion. While the compensatory nature of the gene IV mutation remains unknown, we suspect a connection between the location of the lacZα insert and the mutations in the 5’-untranslated region of gene II. We hypothesize that these mutations increase the expression of pII, the protein that initiates the rolling circle replication of the M13 genome, and that such increased levels of pII compensate for the lacZα disruption of the origin. We will present newly discovered phage clones from all three categories, as well as our preliminary data for the quantification of pII levels in wild-type phage, M13KE, and M13KE mutants.

Various genetic changes compensate for a lacZ α insertion in a sensitive area of the M13 bacteriophage genome

251st National Meeting (2016)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.