Tagged polymerase chain reaction subtractive hybridization for the enrichment of phage display random peptide libraries |
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Authors: | Tarr Alexander W Boneham Steven P Grabowska Anna M Ball Jonathan K |
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Affiliation: | Institute of Infections, Immunity, and Inflammation, School of Molecular Medical Sciences, Queen's Medical Centre, University of Nottingham, Nottingham, NG7 2UH, UK. |
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Abstract: | 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. |
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Keywords: | Subtractive hybridization Random peptide libraries Positive enrichment DNA selection Tagged PCR Phage display Phage libraries |
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