Method for detecting protein-protein interaction.

摘要

A method for detecting an interaction between a first test protein and a second test protein, comprising: (a) providing a yeast host cell containing a reporter gene, wherein the reporter gene expresses a detectable protein when the gene indicator is activated by an amino acid sequence including a transcriptional activation domain when the transcriptional activation domain is sufficiently close to the reporter gene; (B) providing a first hybrid gene that is expressed in the yeast host cell, comprising the first hybrid gene a DNA sequence encoding a first hybrid protein, the first hybrid protein comprising: (i) a DNA-binding domain that recognizes a binding site on the reporter gene in the host cell; and (ii) a first test protein or fragment thereof, which is to test the interaction with at least one second test protein or fragment thereof; (C) proporcionarun second hybrid gene is expressed in the yeast host cell, the second chimeric gene a DNA sequence encoding a second hybrid protein, the second hybrid protein comprising comprising: (i) the transcriptional activation domain; and (ii) a second test protein or fragment thereof, which is to test the interaction with the first test protein or fragment thereof; wherein the interaction between the first test protein and the second test protein in the yeast host cell causes the transcriptional activation domain activates transcription of the reporter gene; (D) introducing the second chimeric gene into a host cell strain of yeast of a first type of binding and subsequently introducing said cells into an arraying means, thereby creating a master library plate; (E) introducing cells master library plate into a second arraying means which is an arraying means for liquid media thereby creating a joint assembly; (F) introducing the first chimeric gene into a host cell strain of yeast type opposite to (d) binding and subsequently introducing said cell into the mating set, thereby allowing the binding to occur in liquid medium; and (g) selecting the product of the interaction of the first and second genes.