Chromosomal integration of a binding domain:bait gene into yeast enhances detection in the two-hybrid system

Shuhei Miyashita, Yukio Shirako

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

We have developed a modified yeast two-hybrid system using the GAL4 transcription activator by integrating a BD:bait gene (GAL4 binding domain:bait gene) into the host chromosome. Locus-specific integration by homologous recombination and use of a strong transcription promoter enabled uniform expression of the integrated BD:bait gene in all host cells. Moreover, after introduction of an AD:prey (GAL4 activation domain:prey) plasmid, false positives were nearly undetectable and false negatives were decreased significantly. Using this improved method a cDNA library could be efficiently screened using roughly 1/10 of the materials required for conventional sequential two-hybrid screen.

Original languageEnglish
Pages (from-to)179-184
Number of pages6
JournalJournal of Microbiological Methods
Volume73
Issue number2
DOIs
Publication statusPublished - 2008 May 1
Externally publishedYes

Keywords

  • Aureobasidine A
  • Chromosomal integration
  • GAL4 transcription activator
  • Protein-protein interactions
  • Two-hybrid system
  • cDNA library screening

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Microbiology (medical)

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