The 'prey' in yeast two-hybrid is fused to:
In yeast two-hybrid, functionality is split between two fusion proteins. Bait carrying the DNA-binding domain occupies promoter elements but lacks activation capacity. Prey consists of library proteins fused to a transcriptional activation domain that can recruit RNA polymerase II machinery and coactivators. Only upon physical binding between bait and prey does the activation domain localize to the reporter promoter, restoring a complete transcription factor. This design allows nuclear interaction to be converted into selectable reporter expression, forming basis for high-throughput binary interactome screening.
Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.