Selective enrichment and high-throughput screening of phage surface-displayed cDNA libraries from complex allergenic systems.
Phage surface display technology, first described in 1985, enables the construction of large combinatorial peptide and antibody libraries. The basic concept of linking the phenotype, expressed as gene product displayed on the phage surface, to its genetic information integrated into the phage genome, allows the survey of large libraries for the presence of specific clones using the discriminative power of affinity purification. The selection procedure involves the enrichment of phage by binding to an immobilized target molecule. As a consequence of the physical linkage between genotype and phenotype, sequencing the DNA of the integrated section of the phage genome can readily elucidate the amino acid sequence of a displayed gene product. Phage surface display technology has revolutionized our ability to select agonist and antagonist peptides, antibodies with desired specificities, and DNA-binding molecules. We have extended phage surface display to access cDNA libraries with this powerful screening technology based on affinity selection of desired clones. Here we discuss construction of cDNA libraries displayed on the phage surface, selective enrichment of clones and robotics-based high-throughput screening of enriched libraries.
Crameri
R
R
Swiss Institute of Allergy and Asthma Research (SIAF), Davos, Switzerland. crameri@siaf.unizh.ch
Walter
G
G
D000485:Allergens / Q000235:genetics*; D001435:Bacteriophages / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary; D006801:Humans; D012371:Robotics
D000485:Allergens; D018076:DNA, Complementary