Phage-based expression cloning to identify interacting proteins.
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.
Stone
J M
JM
University of Missouri, Columbia, Columbia, Missouri, USA.
D001435:Bacteriophages / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary; D007202:Indicators and Reagents; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D012996:Solutions
D018076:DNA, Complementary; D007202:Indicators and Reagents; D011993:Recombinant Fusion Proteins; D012996:Solutions