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Two-hybrid methods detect interactions between two proteins fused at the C-termini of, respectively, a DNA-binding domain and the activation domain of a transcriptional activator. Thus the N-terminus of none of these proteins is available for interaction. We have tested whether a bait protein with a reverted polarity (i.e. N-bait-LexA-C) is suitable for two-hybrid interaction. We show that such constructs give a specific interaction signal, and document two cases where the sensitivity is dramatically increased. Such constructs might lead to the identification of partners missed during classical two-hybrid screens.

Getting more from the two-hybrid system: N-terminal fusions to LexA are efficient and sensitive baits for two-hybrid studies