Phage display of cDNA repertoires: the pVI display system and its applications for the selection of immunogenic ligands.
The selection of phage displayed cDNA repertoires on an immobilised target has been reported to be an efficient way to rapidly identify interacting partners. To date, however, only a few successful applications have been reported. Here, we present a review of the current status of the display and selection of cDNA libraries using phage. As an example, we report the construction of a set of phage display vectors suitable for cDNA display based on fusion to the minor bacteriophage coat protein 6 (pVI) of filamentous phage. We have evaluated these vectors through the display of the C(H)3 domain of human IgG and of the Escherichia coli alkaline phosphatase (PhoA) gene. Both the C(H)3 domain of IgG and PhoA are shown to be displayed on pVI, and PhoA is also shown to be enzymatically active. We have constructed primary colorectal tumor cDNA repertoires in these vectors and performed selections on both a monoclonal antibody to beta2 microglobulin (beta2M) and polyclonal antibody sera to human IgG. In both cases, relevant ligands were recovered from the phage displayed cDNA repertoire. These vectors may be used for selection of phage displayed cDNA libraries with polyclonal sera from patients. This will allow the identifying antigenic cDNA products in such diseases as cancer, viral/bacterial infections or autoimmune disease. Furthermore, by selections with other specific biomolecules, this display system may aid the identification of interacting partners in functional genomics.
Hufton
S E
SE
Research Institute Growth and Development, Department of Pathology, Maastricht University, PO Box 5800, 6202 AZ, Maastricht, Netherlands.
Moerkerk
P T
PT
Meulemans
E V
EV
de Bruïne
A
A
Arends
J W
JW
Hoogenboom
H R
HR
D000469:Alkaline Phosphatase; D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000235:genetics* / Q000378:metabolism; D036022:Capsid Proteins; D018844:Cyclin-Dependent Kinases / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary; D029968:Escherichia coli Proteins; D005822:Genetic Vectors; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology; D008024:Ligands; D008969:Molecular Sequence Data; D019151:Peptide Library
D036022:Capsid Proteins; D018076:DNA, Complementary; D029968:Escherichia coli Proteins; D007074:Immunoglobulin G; D008024:Ligands; D019151:Peptide Library; C109203:gene VI protein, bacteriophage phi-Lf; D018844:Cyclin-Dependent Kinases; D000469:Alkaline Phosphatase; C114089:phoA protein, E coli