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Yeast One-Hybrid Screens for Detection of Transcription Factor DNA Interactions

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Plant Reverse Genetics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 678))

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.

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Acknowledgements

This work was supported by grants CerealGene Tags (European Commission FP5 project QLG2-CT-2001-01453) and NWO 901-07-206 (Netherlands Organization of Scientific Research) to P.B.F.O. and TF-STRESS (European Commission FP5 project QLK3-CT-2000-00328) and ZF-Models (European Commission FP6 project LSHG-CT-2003-503496) to A.H.M.

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Authors and Affiliations

  1. Sylvius Laboratory, Institute of Biology (IBL), Leiden University, Leiden, The Netherlands

    Pieter B. F. Ouwerkerk

  2. Clusius Laboratory, Institute of Biology (IBL), Leiden University, Leiden, The Netherlands

    Annemarie H. Meijer

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  1. Pieter B. F. Ouwerkerk
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  2. Annemarie H. Meijer
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Editors and Affiliations

  1. , Virginia Bioinformatics Institute, Virginia Tech, Washington Street, Blacksburg, 24061, Maryland, USA

    Andy Pereira

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Ouwerkerk, P.B.F., Meijer, A.H. (2011). Yeast One-Hybrid Screens for Detection of Transcription Factor DNA Interactions. In: Pereira, A. (eds) Plant Reverse Genetics. Methods in Molecular Biology, vol 678. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-682-5_16

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  • DOI: https://doi.org/10.1007/978-1-60761-682-5_16

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60761-681-8

  • Online ISBN: 978-1-60761-682-5

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