Biomedical Engineering Reference
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FIGURE14.6 Schematic representation of theY2Hassay. The genes coding two investigated
proteins are inserted into different plasmids in frame with genes coding for two parts of the
transcription factor. The proteins are expressed and, if they bind, the transcription factor
becomes functional, thus expressing the measurable reporter gene.
These two fragments, able to reconstruct a functional transcription factor when inter-
acting physically, are fused with two proteins (A and B) whose interaction has to be
assessed. Separate plasmids are generated, one containing protein A in a frame with
the BD, and the other containing protein B in a frame with the AD (Figure 14.6). The
expression of both genes leads to the synthesis of two chimeric proteins composed
of the proteins studied associated with the two fragments of the transcription factor.
If the two proteins do interact, the two transcription factor fragments can bind, thus
forming a functional transcription factor. Then, the functional transcription factor
induces a measurable expression of the reporter gene. The classical transcription
factor-reporter system used in the two-hybrid assay encompasses the exploiting
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