title: A phage display vector optimized for the generation of human antibody combinatorial libraries and the molecular cloning of monoclonal antibody fragments.
abstract: A novel phagemid vector, named pCM, was optimized for the cloning and display of antibody fragment (Fab) libraries on the surface of filamentous phage. This vector contains two long DNA "stuffer" fragments for easier differentiation of the correctly cut forms of the vector. Moreover, in pCM the fragment at the heavy-chain cloning site contains an acid phosphatase-encoding gene allowing an easy distinction of the Escherichia coli cells containing the unmodified form of the phagemid versus the heavy-chain fragment coding cDNA. In pCM transcription of heavy-chain Fd/gene III and light chain is driven by a single lacZ promoter. The light chain is directed to the periplasm by the ompA signal peptide, whereas the heavy-chain Fd/coat protein III is trafficked by the pelB signal peptide. The phagemid pCM was used to generate a human combinatorial phage display antibody library that allowed the selection of a monoclonal Fab fragment antibody directed against the nucleoprotein (NP) of Influenza A virus.
authors: Laboratory of Microbiology and Virology, Vita-Salute San Raffaele University, Milan, Italy.
authors: Laura
authors: L
authors: Solforosi
authors: Nicasio
authors: N
authors: Mancini
authors: Filippo
authors: F
authors: Canducci
authors: Nicola
authors: N
authors: Clementi
authors: Giuseppe Andrea
authors: GA
authors: Sautto
authors: Roberta Antonia
authors: RA
authors: Diotti
authors: Massimo
authors: M
authors: Clementi
authors: Roberto
authors: R
authors: Burioni
mesh_terms: D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000276:immunology; D053118:Influenza A Virus, H1N1 Subtype / Q000276:immunology; D007763:Lac Operon; D008297:Male; D008875:Middle Aged; D019590:Nucleocapsid Proteins; D019151:Peptide Library; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals; D016601:RNA-Binding Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D014758:Viral Core Proteins / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D007140:Immunoglobulin Fab Fragments; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D019151:Peptide Library; D021382:Protein Sorting Signals; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014758:Viral Core Proteins