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).
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