...
TEV cleavage site high or low affinity?
Experiment
- Western blot HEK293 cells for Syk
- anti-Syk antibody
- Check BCR membrane localization
- add CD79a/b + mIgM and verify with fluorescent anti-IgM antibodies
- Check IgM binding
- biotinylated beta-amyloid with fluorescent secondary antibody
- Check Lyn and Syk membrane localization using fluorescent fusions
- Lyn-YFP
- Syk-mKate
- Make Syk+TEVp fusions
- Fusions ->
...
- PrioritizeFusions
- N terminus: 0 linking aa
- N terminus: 15 linking aa
- C terminus: 0 linking aa
- C terminus: 15 linking aa
- Inducible expression under TetR -> find upper limit for constitutive behavior
Syk+TEVp -> find lower limit of beta amyloid (threshold)
High Priority Fusions
- Syk-BFP
...