Oscillator based
From 2006.igem.org
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)
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Contents |
Scope
This group is focussing on Ideas related to Counters and Oscillators.
Motto
ASCII is an art form!
People
People currently working on these Ideas:
Please feel free to add your name.
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.
Module Tree
---------------- -----------
| | | | |
|cell division |-----> | | |
| | | | |
---------------- | | |
|----| | ---------------------
---------------- | | | | | ------------
| | | | | | | | |
| oscillator |-----> | | |---->| divisor/counter |----->| reporter |
| | | | | | | | |
---------------- | | | | | ------------
|----| | ---------------------
---------------- | | |
| triggering | | | |
| by hand |-----> | | |
| | | | |
---------------- -----------
Interface
cell division
oscillator
Bad news! I raed 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.
triggering by hand
interface
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.
'4 genes' counter
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:
--- ---- ---
\ / \ /
\ / \ /
---------------- ----------------
Challanges
- The combination of promoters, repressing and activating substances will be quite a challange. It should change the state rather quick. Therefore most of the substances have to be decomposed quickly. But on the other side 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?
Toggle switch
Idea
During our discussion, it became apparent that another possibiliy to build a counter / divisor would be to use a toggle switch. It seems that the term "toggle switch" is not well-defined, i.e. it can stand for a few different functionalities of electronic circuits.
When I use the term "toggle switch" in this section, it shall stand for a genetic device with one input and one output, where both of them have two possible states (high and low). When the input is low, the output doesn't change (it keeps its current state). When there is a peak in the input signal (i.e. it jumps to the high state but doesn't stays there to long, where the length of "to long" depends on the parameters of the circuit), the output changes to the state opposite to its current state (from low to high or from high to low). It "toggles".
Concept for Implementation
A possible implementation of this concept could look as following:
The first device needed is just a bistable switch, as described in the lecture by Jörg (if someones remembers the paper where this came from, please add a reference to that paper here)
R1 R2
\ /
\/|____ ____|\/
___/ (1) |_______ _______| (2) \___
\ ____| = = = = |____ /
\| | | | | |/
| | | |
R2 R3 R4 R1
The bistable switch produces either R1 or R2. When R1 is present, it represses the production of R2. If R2 is present, it represses the production of R1. Thus, this device has two stable states. I will refer to the state where R1 is produced as State 1 and to the state where R2 is produced as State 2. The repressors R3 and R4 should be ignored in this stage, they get their relevance when the second component of the toggle switch is introduced below.
Now, we need another device that allows us to switch the bistable switch from one state to the other. I call it switcher. This could be achieved in the following way:
S R1 R3 S R2 R4
| | / | | /
| | ___|\/ | | ___|\/
_|__|__| (3) \___ _|__|__| (4) \___
|___ / |___ /
|/ |/
The idea of the switcher is to repress the production of the repressor (R1 or R2) that is currently produced by the bistable switch. It does so only when the input signal S is present. The concept of the dynamics of a toggling is as folloeing:
Given the bistable switch is currently in state 1 with gene (1) activated and gene (2) repressed. When the concentration of S rises, S in combination with R1 leads to the production of R3, which itself cuts of the production of R1, allowing R2 to be produced. The bistable switch has been toggled.
Challenges
To make this device work, one has to adjust the time constants. There are some challenges to face:
- Assume the bistable switch has just been toggled from state 1 to state 2. When S is still present, R2 produced in state 2 can lead to a retoggling to state 1. This behavior is clearly undesired. To avoid this, S should be a substance that is degraded quickly.
- Even when S has sharp peaks, there is still a problem during transition. Given the toggle switch is in state 1. S rises, R3 is produced, and the production of R1 in reality is rather first lowered than directly cut off. This leads to a lower production of R3, thus slowing down the toggle process. To avoid this, R3 should be stable on a longer timescale than the timescale of the processes in the toggle switch. If this is ensured, a short peak of S is sufficient to produce some persistent R3, that is able to stop the production of R1. This allows one to make the peak short enough to guarantee that the above described "double toggling" does not happen.
Using the toggle switch as a counter
If you have such a device working, it is possible to convert an input signal with frequency to a signal with frequency 2f. This is a divisor. If you combine n of these divisors, you are able to count to 2^n. That's a counter!
Evaluation of the idea
I think that this concept can be implemented. The problems described above should be possible to deal with. I expect the toggle switch to be quite stable and noise resistant, and yet sufficiently simple regarding the different interactions. The catenation of the toggle switches to a counter may prove to be not so easy. A toggle switch is a device, that can be used in a wide area of applications, given the importance of the concept in electrical engineering and computer science.
A test-tube that changes its color from red to green when some substance is added, is not so spectacular, since we all saw this in our high school chemical class. A tube changes back fron green to red when another chemical is added, is slightly better. But what a cool demo, when the tube switches it's color again and again, every time ONE SINGLE SUBSTANCE is added! (for comments and opinions to the coolness of such a demo, please use the talk page...))
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
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