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Updated: Feb 27, 2026

Author Spotlight: Enhanced Isolation of Interaction-Null Mutants in Yeast
Published on: December 29, 2023
Protein-Protein Interactions: Yeast Two-Hybrid System
1Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan.
Abstract:
The yeast two-hybrid system is a powerful and commonly used genetic tool to investigate interactions between artificial fusion proteins inside the nucleus of yeast. Here we describe how to use the Matchmaker GAL4-based yeast two-hybrid system to detect the interaction of the Agrobacterium type VI secretion system (T6SS) sheath components TssB and TssC41. The bait and prey gene are expressed as a fusion to the GAL4 DNA-binding domain (DNA-BD) and GAL4 activation domain (AD, prey/library fusion protein) respectively. When bait and prey fusion proteins interact in yeast nucleus, the DNA-BD and AD are brought into proximity, thereby activating the transcription of reporter genes. This technology can be widely used to identify interacting partners, confirm suspected interactions, and define interacting domains.
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