Oscillator based
From 2006.igem.org
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Contents |
Scope
This group is focussing on Ideas related to Counters and Oscillators.
Motto
ASCII is an art form!
Organisation
People
People currently working on these Ideas:
Please feel free to add your name.
Meetings
Friday, 5. August, 09:00 @ Christophe's bureau (CAB F61.2) Summary of the meeting Monday, 8. August, 15:00 @ Christophe's bureau (CAB F61.2) Tuesday, 9. August, 09:00 @ Christophe's bureau (CAB F61.2)Summary of the meeting Wednesday, 10. August, 09:15 @ Christophe's bureau (CAB F61.2)
Modules
We try to divide all the Ideas related to our scope in some 5 or 6 Modules. Our Goal on the one hand is to define the modules in a way that allows one to design, implement and test the individual modules independently. On the other hand, we also want to ensure that it is possible to connect them to build bigger components with more functionality.
+---------------+
| OSCILLATOR |
| ============ |
+---------------+
| |
| cell division | +-------------------+ +-------------------+
| | | | | |
+---------------+ | DIVISOR/COUNTER | | |
| oscillator |----> | =============== |----> | reporter gene |
| from previous | | | | |
| work | | | | |
+---------------+ +-------------------+ +-------------------+
| triggering |
| by hand |
| |
+---------------+
TRIGGERING
cell division
Uwe Sauer suggests to look into ftsZ as a protein that is expressed only once per cycle. We could try to get their promoter and use it to build a peak as input for the counter.
to read
- FtsZ Dynamics during the Division Cycle of Live Escherichia coli Cells
- Regulation of FtsZ during the Caulobacter cell cycle
- Course Summary
- Anatomy of a bacterial cell cycle
- How Do Prokaryotic Cells Cycle
oscillator
attenuated oscillator from cell articel
Bad news! I read the paper about oscillatory behavior in E coli (atkinson03) and if I did get it right then they didn't completely succed to develop a stable oscillator. Their design enabled them to achieve 4 different behaviors of the circuit by changing the promoters, repressors, copy numbers and cooperativity. Due to theire simulations the circuit should have been able to act as a toggle switch, a damped oscillator and a stable oscillator (with different frequencies and peak concentrations). They succeded in establishing reproducibly a damped oscillation. But by shifting the system parameters into the region of stable oscillation the damping didn't disappear. Pushing it further could possibly help but I don't know if it's worth it and if we have enough time for this.
Furthermore they didn't include all the genes into peptides but a part also in the DNA of the E coli itself. I don't know if this would be a problem but I wanted to mention it anyway.
repressilator
The repressilator is working but as we know it has its drawbacks. On one hand it isn't oscillation isn't really very regullary and on the other hand it's amplitude is increasing with time. With this oscillator you could perhaps count to 2 or 3. Because if the minimum amplitude is to high thecounter wouldn't recognice the difference anymore. But instead of using the simple hand triggering this could be a alternative to test the counter.
another part from the registry of standard parts
triggering by hand
Let's brainstorm about stuff that could be used to generate an oscillation by hand.
- IPTG? What about degradation time?
- ...
DIVISOR/COUNTER
This module will get an input concentration from the interface module and will count, due to the changing input, to 4. Also a reporter (for example different colors) will be produced which could serve as an indicater for the state the circuit is currently occupiing.
Idea and Implementation
During our discussion about the divisor implementation we noticed that the previous one wasn't quite stable. Talking about toggle switches we suddenly had the idea of combining the base structur of the toggle switch we learned to know during the crash course with the allready existing design. The follwing drawings show this new design which is basically a combination of two such toggle switches.
All substances which are involved should be produced and decomposed quickly
S = concentration signal from the interface
R1 is produced by gene (1) and represses gene (0) and (3)
R2 is produced by gene (2) and represses gene (0) and (1)
R3 is produced by gene (3) and represses gene (1) and (2)
R0 is produced by gene (0) and represses gene (2) and (3)
---S---
R1 / \ R3 R3 R0 R1 R2
\ | | / | | | |
\/|____ | | ____|\/ /|___ | | | | ____|\
___/ (1) |_____v__ __v_____| (3) \___ ___/ (2) |_=_=____ ____=_=_|(0=4) \___
\ ____| = = = = |____ / \ ____| = = |____ /
\| | | | | |/ /\| | | |/\
| | | | / | | \
R2 R3 R0 R1 R2 \ / R0
---S---
The following time lines should be the result of this gene circuit:
S:
----- ----- ----- ----- Let's assume that the counter is in state 0
| | | | | | | | (expressing R0). As soon as the concentration
| | | | | | | | of S goes up (0) and (2) are repressed
--- ----- ----- ----- ---- (no further expressing of R0). Now R1 can be
R1: expressed while (3) is still repressed
---- ---- by R0. The production of R1 ensures that (3)
/ \ / \ won'tbe active when R0 is totally decomposed.
/ \ / \ -> We are in state (1).
--- ---------------- --------------- When the S goes down again R1 can't be
R2: expressed anymore. Now R2 can be
---- ---- expressed while (0) is still repressed by R1.
/ \ / \ The production of R2 ensures that (0) won't
/ \ / \ be active when R1 is totally decomposed.
--------- ---------------- --------- -> We are in state (2)
R3: And so on...
---- ----
/ \ / \
/ \ / \
--------------- ---------------- ---
R0:
--- ---- ---
\ / \ /
\ / \ /
---------------- ----------------
Challenges
- The combination of promoters, repressing and activating substances will be quite a challenge. They should change the state rather quick. Therefore most of the substances have to be expressed and decomposed quickly. But on the other hand the repression at certain points should stay long enough to ensure that there is only one state the system can fall in.
- I think the repression by the input signal S will affect the expression of R2 and R0 quickly. But will the decreasing concentration of S have the same affect on R1 and R3 in comparable time? Or is a second input needed with the reverse effect of S?
REPORTER
This will probably be one or more colors to indicate the current state of the counter.
Discussions
We invite anyone to make comments and participate in discussions here
- Coolness
- Modularity
- Usefulness
- Feasibility
Attic
Have a look at Oscillator_based_Attic for old toggle stuff.
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