Yeast one-hybrid screens for detection of transcription factor DNA interactions

Methods Mol Biol. 2011;678:211-27. doi: 10.1007/978-1-60761-682-5_16.

Abstract

The yeast one-hybrid system is widely recognized as a valuable and straightforward technique to study interactions between transcription factors and DNA. By means of one-hybrid screens, transcription factors or other DNA-binding proteins, expressed from cDNA expression libraries, can be identified due to the interactions with a DNA sequence-of-interest that is linked to a reporter gene, such as the yeast HIS3 gene. Usually, the library is constructed in an E. coli-yeast shuttle vector designed for production of hybrid proteins consisting of a library protein and the trans-activating domain (AD) from the yeast GAL4 transcription factor. Here, we describe an optimized system of vectors for one-hybrid screenings together with detailed step-wise protocols, an elaborate trouble-shooting guide and many technical tips to conduct successful screenings. This system and other yeast genetic selection procedures derived from one-hybrid methodology proved highly useful to help understanding the regulatory networks controlling expression of the genome.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA / metabolism*
  • Genotype
  • Polymerase Chain Reaction
  • Protein Binding
  • Transcription Factors / metabolism*
  • Two-Hybrid System Techniques*
  • Yeasts / genetics
  • Yeasts / metabolism*

Substances

  • Transcription Factors
  • DNA