Membrane-Based Yeast Two-Hybrid

Principle

Screening for transmembrane protein interactions using a split-ubiquitin system. The split-ubiquitin system consists of the N-terminal (Nub) and C-terminal (Cub), which typically have a strong affinity and can function as wild-type ubiquitin. Here, the isoleucine at position 3 of Nub is mutated to glycine (NubI is mutated to NubG), reducing its affinity with Cub. Meanwhile, the Cub portion is fused with the artificially synthesized LexA transcription activation factor to form a fusion protein Cub-LexA. Then, the proteins to be tested are separately fused with NubG and Cub-LexA, forming bait fusion protein (bait-Cub-LexA) and prey fusion protein (prey-NubG). If the bait and prey interact, NubG and Cub come into close proximity, recognized by UBPs, which results in the cleavage of LexA, allowing it to enter the nucleus and activate the transcription of the reporter gene.

Delivery Time

Our Service Advantages:

A. Library Selection:

Customized yeast libraries with a unique "one library, four uses" feature (single hybrid, double hybrid, triple hybrid, and overexpression vector libraries): Utilizing an optimized version of Clontech's pGADT7-Rec2, GATEWAY secondary recombination, and "one-step recombination" methods.

Offering commercialized yeast cDNA libraries.

B. Screening Service:

The yeast membrane-based two-hybrid system can be used to study the interaction modes between membrane proteins or cytoplasmic proteins and protein binaries. The screening steps are optimized, with a well-established experimental workflow, providing accurate and reliable experimental data and original images.

C. Post-screening can also proceed with high-throughput screening Y2H-seq simultaneously.