A new phage display system to construct multicombinatorial libraries of very large antibody repertoires.
We present an easy and efficient technique for the construction of large phage-displayed antibody (Ab) repertoires through the recombination of two separate heavy (VH) and light (VL) chain gene libraries. Here, the system has been applied to the display of a chimpanzee anti-HIV gp160 Ab. The process, which makes use of lambda phage att recombination sites, leads to the irreversible physical association between plasmid and phagemid vectors carrying, respectively, VL and VH sequences. The heat-inducible expression of the Int recombinase allows perfect control of recombination. Selection of the recombinant phagemid is made possible by the assembly, in vivo, of a genetic marker (chloramphenicol resistance) created only after the correct recombination event. Theoretically, all possible associations between the VL and VH sequences should be obtained, and it should be possible to generate multicombinatorial libraries of close to 10(12) clones.
Geoffroy
F
F
Department of Molecular Immunology, Pasteur Mérieux Sérums et Vaccins, Marcy l'Etoile, France.
Sodoyer
R
R
Aujame
L
L
D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D015686:Gene Products, env / Q000032:analysis / Q000276:immunology*; D005803:Genes, Immunoglobulin; D006678:HIV / Q000276:immunology / Q000302:isolation & purification; D015483:HIV Antibodies / Q000235:genetics*; D019302:HIV Envelope Protein gp160; D007140:Immunoglobulin Fab Fragments / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007147:Immunoglobulin Light Chains / Q000235:genetics*; D002679:Pan troglodytes; D010957:Plasmids; D011498:Protein Precursors / Q000032:analysis / Q000276:immunology*; D011995:Recombination, Genetic; D015183:Restriction Mapping
D015686:Gene Products, env; D015483:HIV Antibodies; D019302:HIV Envelope Protein gp160; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D011498:Protein Precursors