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· (Finley Lab)
This is one of the most comprehensive and detailed guide to yeast two-hybrid system technique with introduction to its background. The following procedures are described: mating assay - small scale for tens of different bait or prey proteins, collecting bait (and prey) strains, large scale for hundreds of different bait or prey strains, interaction mating assay with other yeast two-hybrid systems, recording results, interpreting interaction data, qualitative interpretation and more...
· Yeast Two-Hybrid System (Finley Lab)
The updated version for scaling up the these techniques to facilitate characterization of large networks of interacting proteins. It is also useful for individual hunts.
· Performing a Hunt by Interaction Mating (Finley Lab)
In this protocol one strain is transformed with library DNA and the transformants are collected and frozen in aliquots. For each interactor hunt, an aliquot of this frozen "pretransformed library strain" is thawed and mixed with an aliquot of a bait strain transformed with the bait expression plasmid. Overnight incubation of the mixture on a YPD plate results in mating - i.e., cells of one strain fuse with cells of the other strain to form diploids. The diploids are then exposed to galactose to induce expression of the library encoded proteins, and interactors are selected in the same manner as in the standard hunt protocols.
· GST Fusion Protein Purification from Yeast (Dohlman Lab)
· Two Hybrid System Transformation (Gietz Lab)
This Protocol is for 2 Hybrid system screens
· Purification of MBP (maLTose-Binding Proteins) Fused Proteins (PMCI)
· Purification of GST Fused Proteins (PMCI)
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