PMID: 9547290Jun 20, 1998Paper

A recombination based method to rapidly assess specificity of two-hybrid clones in yeast

Nucleic Acids Research
R PetermannH Kovar

Abstract

The yeast two-hybrid system is frequently used to identify protein-protein interactions. Confirming the specificity of candidate clones requires separation and isolation of yeast plasmids, propagation in bacteria and testing combinations of DNA-binding and activation domain hybrids in yeast. In order to simplify this procedure, we developed a rapid method based on PCR amplification of library insert DNAs and in vivo cloning into the activation domain hybrid vector. Reporter gene activity is assayed in parallel for combinations with different DNA-binding domain hybrids. Further characterization of inserts does not require plasmid isolation and intermediate hosts.

References

Nov 1, 1991·Proceedings of the National Academy of Sciences of the United States of America·C T ChienS Fields
Jan 1, 1985·Cold Spring Harbor Symposia on Quantitative Biology·L Breeden, K Nasmyth

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Citations

Mar 9, 2004·Methods : a Companion to Methods in Enzymology·Maxim PoustovoitovPeter D Adams
Jul 16, 2005·BioTechniques·Jennifer I SempleDavid Markie
Apr 4, 2009·Current Protocols in Molecular Biology·Elena KotovaIlya Serebriiskii
Feb 12, 2008·Current Protocols in Molecular Biology·Elena KotovaThomas Coleman

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