Tagged polymerase chain reaction subtractive hybridization for the enrichment of phage display random peptide libraries.
Affinity selection of phage display peptide libraries is routinely used for isolating peptides capable of binding a range of molecules, including antibodies and receptors. This process is most successful when the selecting molecule is relatively pure, for example, a monoclonal antibody. However, isolation of peptides able to bind to target molecules present in a complex mixture is more difficult because the affinity selection process isolates peptides capable of binding to all molecules present in the mixture. Here we describe the development of a tagged polymerase chain reaction (PCR) subtractive hybridization method that is universally applicable for the targeted isolation of peptides able to bind to unique molecules within a complex mixture. We also describe a discriminatory limiting dilution PCR method that can be used to optimize hybridization conditions.
Tarr
Alexander W
AW
Institute of Infections, Immunity, and Inflammation, School of Molecular Medical Sciences, Queen's Medical Centre, University of Nottingham, Nottingham, NG7 2UH, UK.
Boneham
Steven P
SP
Grabowska
Anna M
AM
Ball
Jonathan K
JK
D001435:Bacteriophages / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D006801:Humans; D006824:Hybridization, Genetic; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism; D019151:Peptide Library; D016133:Polymerase Chain Reaction; D013382:Subtraction Technique
D010446:Peptide Fragments; D019151:Peptide Library