Phage-based expression cloning to identify interacting proteins.

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Abstract

Phage-based expression cloning is a simple, rapid, and powerful technique to identify interacting proteins. A protein of interest is expressed as a recombinant fusion protein and labeled with (32)P at an engineered recognition site to facilitate detection. b-gal proteins that are fused in-frame to cDNA inserts in a phage-derived expression library are produced by the phage and adsorbed onto nitrocellulose filters. The filters are then screened with the radioactive protein probe to identify phage clones that express the interacting protein. This technique leads directly to the isolation of a cDNA encoding the interacting protein, bypassing the need for labor-intensive protein purification, microsequencing, or antibody production.

Original languageEnglish (US)
Pages (from-to)Unit 20.3
JournalCurrent protocols in molecular biology / edited by Frederick M. Ausubel ... [et al.]
VolumeChapter 20
StatePublished - May 2001

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Bacteriophages
Organism Cloning
Proteins
Complementary DNA
Recombinant Fusion Proteins
Collodion
Antibody Formation
Clone Cells

ASJC Scopus subject areas

  • Molecular Biology

Cite this

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