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Screening Arrayed Libraries with DNA and Protein Baits to Identify Interacting Proteins

Part of the Methods in Molecular Biology book series (MIMB,volume 1794)

Abstract

Molecular interactions are an integral part of the regulatory mechanisms controlling gene expression. The yeast one- and two-hybrid systems (Y1H/Y2H) have been widely used by many laboratories to detect DNA–protein (Y1H) and protein–protein interactions (Y2H). The development of efficient cloning systems have promoted the generation of large open reading frame (ORF) clone collections (libraries) for several organisms. Functional analyses of such large collections require the establishment of adequate protocols. Here, we describe a simple straightforward procedure for high-throughput screenings of arrayed libraries with DNA or protein baits that can be carried out by one person with minimal labor and not requiring robotics. The protocol can also be scaled up or down and is compatible with several library formats. Procedures to make yeast stocks for long-term storage (tube and microplate formats) are also provided.

Key words

  • Arrayed libraries
  • DNA–protein interaction
  • High-throughput
  • One-hybrid system
  • Open reading frame
  • Protein–protein interaction
  • Transcription factors
  • Two-hybrid system
  • Yeast

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  • DOI: 10.1007/978-1-4939-7871-7_9
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Acknowledgments

The work in L.O.-S. lab is supported by MINECO grants BIO2013-46076-R and BIO2016-77840-R. We thank to all people that contributed to the development of the Arabidopsis TF library [22] as well as all the labs that have used this resource, thus helping to ascertain its potential. We apologize to those authors whose publications are not cited here due to space limitations.

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Correspondence to Luis Oñate-Sánchez .

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Sánchez-Montesino, R., Oñate-Sánchez, L. (2018). Screening Arrayed Libraries with DNA and Protein Baits to Identify Interacting Proteins. In: Oñate-Sánchez, L. (eds) Two-Hybrid Systems. Methods in Molecular Biology, vol 1794. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7871-7_9

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  • DOI: https://doi.org/10.1007/978-1-4939-7871-7_9

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