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The yeast two-hybrid technology exploits the fact that transcriptional activators are modular in nature. A vector is made containing the transcription factor DBD plus a desired protein called the bait. When placed into yeast with a reporter gene, this fusion protein can bind to the reporter gene promoter, but it cannot activate transcription. A second vector is made where unknown cDNAs are placed adjacent to the activation domain of a transcription factor called the prey. When placed into a yeast strain containing the reporter gene, it cannot activate transcription, since it has no DNA binding domain. When the two vectors are placed into the same yeast, a transcription factor is formed that can activate the reporter gene provided that if the protein made by the second plasmid binds to the X protein (STRATAGENE, 2000). The activation of the reporter gene enables histodine and B-galactosidase. For more details, see the HybriZAP-2.1 Library Construction Kit and HybriZAP �V 2.1 XR cDNA Synthesis Kit by STRATAGENE.FIG 4 |
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