Definition:

The λ repressor two-hybrid system is a protein-protein interaction technique mainly used to study the direct interaction between two proteins. This method employs fusion technology to link the protein of interest (bait) with the λ repressor protein, which contains a DNA-binding domain and a dimerization domain. The bait protein specifically binds to the λ operator sequence through the DNA-binding domain of the λ repressor, which is located upstream of the reporter gene promoter.
The corresponding target protein (prey) is fused to the N-terminal of the RNA polymerase α subunit. When the bait and prey proteins meet and interact within the cell, their binding promotes the stable binding of RNA polymerase to the promoter, thus initiating the transcription of the reporter gene (such as HIS3). The protein interaction is detected by measuring the activity of the reporter gene to determine the presence and strength of the interaction.
However, since both the λ repressor protein and the N-terminal of the RNA polymerase α subunit have a tendency to dimerize, this may affect the interpretation of the experimental results, especially when analyzing proteins that have an inherent tendency to aggregate. Therefore, this system is more suitable for studying proteins whose interactions depend on aggregation with other proteins.