Effect of DNA copy number on genetic stability of phage-displayed peptides.
A small model peptide, the FLAG epitope, was cloned into two filamentous phage display vectors, f88-4 and fd88-4, creating phages f88-FLAG and fd88-FLAG, respectively. Both vectors have a gene VIII display cassette (in addition to their normal phage gene VIII) and display the cloned peptide on a few percent of the virion's 3000-4000 pVIII (major coat protein) subunits. Vector f88-4 has a replication defect and attains low DNA copy number in infected cells, while vector fd88-4 has no replication defect and attains the normal, high DNA copy number characteristic of wild-type filamentous phage. Almost no loss of displayed peptide was observed during six rounds of propagation of low copy number f88-FLAG phage. In contrast, when high copy number fd88-FLAG phage was similarly propagated, variant clones that did not display the FLAG epitope accumulated gradually. The loss of displayed peptide from the high copy number vector is undoubtedly slow enough to be overcome by even weak affinity selection, and high copy number vectors have important advantages that make their use worth considering, at least when the displayed peptides are small.
Smith
George P
GP
Division of Biological Sciences, Tucker Hall, University of Missouri, Columbia, MO 65211, USA. smithgp@missouri.edu
Fernández
Ana-María
AM
D000595:Amino Acid Sequence; D001435:Bacteriophages / Q000235:genetics*; D001483:Base Sequence; D004247:DNA / Q000235:genetics*; D000939:Epitopes / Q000235:genetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics*
D000939:Epitopes; D010455:Peptides; D004247:DNA