The double mutant tells you the order
- The idea
- Predict
- See it
- Why
- Check
The idea
“The single mutants tell me the order, so I don't need the strain with both changes.”
Two clean single-mutant results feel like plenty of evidence. When locking either protein on gives the same result, they can't say which comes first.
The short version
The mistake: The single mutants tell me the order, so I don't need the strain with both changes.
What's actually true: Locking each protein on alone only shows it can drive the response. Lock one on and remove the other to see which acts first.
- A protein locked on, or constitutively active, stays on with no signal. If locking either protein on starts the response, that fits either order.
- The double mutant settles it. Lock one protein on and remove the other. If the response still happens, the locked-on protein acts after the missing one.
- If the response fails, the locked-on protein needs the missing one to pass the signal on. So the missing one acts later.
On the exam: If a locked-on protein still works with the other one gone, it acts later in the pathway.