Abstract
Because of its adaptability to high-throughput approaches and a low operating cost, the yeast two-hybrid (Y2H) assay remains the most widely used one for high-throughput protein–protein interactions (PPI) mapping experiments. Here we provide a detailed protocol for a liquid culture-based high-throughput binary protein–protein Y2H screen pipeline of a pool of 50 proteins used as baits against a collection of ~12,000 Arabidopsis proteins encoded by sequence-verified open reading frames (ORFs).
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
The Arabidopsis Genome Initiative (2000) Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408:796–815
Zaag R, Tamby JP, Guichard C et al (2015) GEM2Net: from gene expression modeling to -omics networks, a new CATdb module to investigate Arabidopsis thaliana genes involved in stress response. Nucleic Acids Res 43:D1010–D1017
Braun P, Tasan M, Dreze M et al (2009) An experimentally derived confidence score for binary protein–protein interactions. Nat Methods 6(1):91–97
Cusick ME, Yu H et al (2009) Literature-curated protein interaction datasets. Nat Methods 6(1):39–46
Simonis N, Rual JF, Carvunis AR et al (2009) Empirically controlled mapping of the Caenorhabditis elegans protein–protein interactome network. Nat Methods 6(1):47–54
Venkatesan K, Rual JF, Vazquez A, Stelzl U, Lemmens I et al (2009) An empirical framework for binary interactome mapping. Nat Methods 6(1):83–90
Dreze M et al (2010) High-quality binary interactome mapping. Methods Enzymol 470:281–315
Weßling R, Epple P, Altmann S, He Y, Yang L et al (2014) Convergent targeting of a common host protein-network by pathogen effectors from three kingdoms of life. Cell Host Microbe 16(3):364–375
Arabidopsis Interactome Mapping Consortium (2011) Evidence for network evolution in an Arabidopsis Interactome map. Science 333(6042):601–607
Mukhtar MS, Carvunis AR, Dreze M, Epple P et al (2011) Independently evolved virulence effectors converge onto hubs in a plant immune system network. Science 333(6042):596–601
Walhout AJ et al (2000) GATEWAY recombinational cloning: application to the cloning of large numbers of open reading frames or ORFeomes. Methods Enzymol 328:575–592
Acknowledgments
The IPS2 Interactomic platform benefited from the support of Saclay Plant Sciences-SPS (ANR-17-EUR-0007).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Castandet, B., Lurin, C., Delannoy, É., Monachello, D. (2023). High-Throughput Protein–Protein Interactions Screening Using Pool-Based Liquid Yeast Two-Hybrid Pipeline. In: Mukhtar, S. (eds) Protein-Protein Interactions. Methods in Molecular Biology, vol 2690. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3327-4_16
Download citation
DOI: https://doi.org/10.1007/978-1-0716-3327-4_16
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-3326-7
Online ISBN: 978-1-0716-3327-4
eBook Packages: Springer Protocols