LambdaZLG6: a phage lambda vector for high-efficiency cloning and surface expression of cDNA libraries on filamentous phage.

We describe a vector, lambdaZLG6, combining the high efficiency of cDNA library cloning in bacteriophage lambda with filamentous phage display of cDNA-encoded products. The cDNAs are expressed as fusions to the 3' end of M13 gene VI. The lambdaZLG library is converted to a pZLG6-cDNA phagemid library by in vivo mass excision. Helper phage infection generates a library of phagemid particles displaying the cDNA-encoded products and containing the corresponding nucleotide sequences within.

Surface expression and ligand-based selection of cDNAs fused to filamentous phage gene VI.

We describe a novel phage display system that affords the surface expression and hence affinity selection of cDNAs. The strategy is based on a new approach to functionally display proteins on filamentous phage through the attachment to the C-terminus of the minor coat protein VI. The utility of the method was evaluated using a cDNA library derived from the parasite Ancylostoma caninum. cDNA sequences were fused in each of the three reading frames to the 3'-end of the M13 gene VI expressed by a phagemid vector. Phages rescued from this cDNA expression library were subjected to biopanning against two serine proteases, trypsin and the human coagulation factor Xa. This led to the identification of cDNAs encoding novel members of two different families of serine protease inhibitors. The authenticity of the cDNA selected with trypsin as the target was demonstrated by purifying the encoded potent Kunitz-type inhibitor from an Ancylostoma caninum extract. The rapid isolation of specific cDNAs with the protein VI monovalent display system should facilitate the search for novel biologically important ligands.