Team:LMU-Munich/TEV-System/Functional Principle

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If one has the Cellline which is stable transformed with TetinduciblePromoter+TEVrecognition+Ndegron+SF3B+bak one can transform these cells with the Construct containing the Protein of interest. If the plasmid has been incorporated by the cell, both protein complexes should be synthesized when the Tet inducible Promoter is induzed by doxycyclin. Both protein-complexes are supposed to attach because p14* and SF3B are interacting proteins. The TEV-protease is supposed to cut the recognition sites. Now the protein of interest (eGFP) is split apart and the N-terminus of the N-degron is uncovered. This is a signal for the cell to deplete the protein complex including the deadly bak. So all protein should be destroyed except the protein of interest. If no plasmid has been incorporated, the cell will die because of Bak.

TEV Construct 1: Inserted in celline

TEV Construct 2: Contains gene of interest

@@ Line 7: / Line 7: @@
 [[Image:TEV1.jpg|thumb|TEV Construct 1: Inserted in celline]]
 [[Image:TEV2.jpg|thumb|TEV Construct 2: Contains gene of interest]]
-[[Image:Jump1.jpg|thumb|Jump-and-Stop Construct 1: Inserted in cellline]]
-[[Image:Jump2.jpg|thumb|Jump-and-Stop Construct 2: attP attachment site and gene of interest]]
-[[Image:Jump3.jpg|thumb|Jump-and-Stop Construct 3: Integrase PhiC31o]]
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