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Nucleic Acids Research, 2003, Vol. 31, No. 11 e59
© 2003 Oxford University Press

A novel helper phage that improves phage display selection efficiency by preventing the amplification of phages without recombinant protein

R. Arjen Kramer, Freek Cox, Marieke van der Horst, Sonja van den Oudenrijn, Pieter C. M. Res, Judith Bia, Ton Logtenberg and John de Kruif

Crucell Holland B.V., PO Box 2048, 2301CA, Leiden, The Netherlands

*To whom correspondence should be addressed. Tel: +31 71 5248779; Fax: +31 71 5248702; Email: j.dekruif{at}crucell.com

Phage display is a widely used technology for the isolation of peptides and proteins with specific binding properties from large libraries of these molecules. A drawback of the common phagemid/helper phage systems is the high infective background of phages that do not display the protein of interest, but are propagated due to non-specific binding to selection targets. This and the enhanced growth rates of bacteria harboring aberrant phagemids not expressing recombinant proteins leads to a serious decrease in selection efficiency. Here we describe a VCSM13-derived helper phage that circumvents this problem, because it lacks the genetic information for the infectivity domains of phage coat protein pIII. Rescue of a library with this novel CT helper phage yields phages that are only infectious when they contain a phagemid-encoded pIII-fusion protein, since phages without a displayed protein carry truncated pIII only and are lost upon re-infection. Importantly, the CT helper phage can be produced in quantities similar to the VCSM13 helper phage. The superiority of CT over VCSM13 during selection was demonstrated by a higher percentage of positive clones isolated from an antibody library after two selection rounds on a complex cellular target. We conclude that the CT helper phage considerably improves the efficiency of selections using phagemid-based protein libraries.


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