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Updated: Jul 23, 2025
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A Yeast 2-Hybrid Screen in Batch to Compare Protein Interactions
Published on: June 6, 2018
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High-Throughput Protein-Protein Interactions Screening Using Pool-Based Liquid Yeast Two-Hybrid Pipeline
Benoît Castandet1, Claire Lurin1, Étienne Delannoy1
1Institute of Plant Sciences Paris-Saclay (IPS2), Université Paris-Saclay, Université Paris-Cité, CNRS, INRAE, Université Evry, Gif sur Yvette, France.
Methods in Molecular Biology (Clifton, N.J.)
|July 14, 2023
Summary
This study details a high-throughput yeast two-hybrid (Y2H) screening pipeline for mapping protein-protein interactions (PPI). The method efficiently tests interactions between 50 bait proteins and ~12,000 Arabidopsis proteins.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- The yeast two-hybrid (Y2H) assay is a key method for mapping protein-protein interactions (PPIs).
- Its adaptability to high-throughput screening and low cost make it widely used.
- Efficient pipelines are crucial for large-scale PPI mapping.
Purpose of the Study:
- To provide a detailed protocol for a high-throughput binary protein-protein Y2H screen.
- To establish a liquid culture-based pipeline for PPI mapping.
- To screen a large collection of Arabidopsis proteins against a defined set of bait proteins.
Main Methods:
- Development of a liquid culture-based yeast two-hybrid (Y2H) screening pipeline.
- Utilizing a pool of 50 proteins as baits.
- Screening against a collection of approximately 12,000 Arabidopsis proteins.
- Employing sequence-verified open reading frames (ORFs).
Main Results:
- A robust and detailed protocol for high-throughput Y2H screening was established.
- The pipeline successfully tested binary interactions between bait proteins and a large Arabidopsis proteome subset.
- Demonstrated the feasibility of large-scale PPI mapping using Y2H.
Conclusions:
- The developed Y2H pipeline offers an efficient approach for high-throughput protein-protein interaction mapping.
- This protocol facilitates the systematic exploration of Arabidopsis interactomes.
- The method is adaptable for studying protein interactions in various biological systems.
