PMID: 16526412Mar 11, 2006Paper

Direct production and purification of T7 phage display cloned proteins selected and analyzed on microarrays

BioTechniques
James E NowakMichael A Tainsky

Abstract

Phage display technology has emerged into a powerful tool for identifying proteins with specific binding properties. This technology adds amino acid sequences to the carboxy terminus of a phage capsid protein, thus generating a fusion protein displayed on the surface of the phage. Here, we have developed a high-throughput strategy to synthesize purified protein that solves many of the problems associated with crude phage lysates. Phage DNA was used as a template for a nested PCR that added the T7 promoter, ribosome binding site, and a His6-tag. The PCR product was then used as a template for in vitro transcription/translation. The resulting His6-tagged recombinant protein was then purified by nickel affinity chromatography. The functionality of the purified protein was verified using protein microarray analysis.

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Citations

Aug 2, 2006·Molecular BioSystems·Gordon R Whiteley
May 13, 2008·Protein Engineering, Design & Selection : PEDS·M DaiA R M Bradbury
Apr 8, 2010·Current Protocols in Neuroscience·Andrew R M Bradbury

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