# User:Anthony Lazzaro / SANDBOX

## 30.08.07

Feedback on PR work:

Overal feedback was positive, several points need to be address

1. Motif needs to transcend wiki, t-shirt design, presentation slides and poster and jamboree

2. Motif needs to have a londonesque theme to it eg. Sherlock Homes

3. Opinion of "Cell Free Sensation" is that Sensation part need to change mabye to something like "Cell Free Detectives"

4. Would like a character to help explain what's going on like Imperial Guys did using Sherlock Homes for his invisibility cloack at royal society.

So to move forward i think i need to made a Sherlock homes motif and change name to something like Cell Free Detectives, need to work on motif but recurring pressure from powers at be to make this happen: V is happy with Cell Free sensation.

## Cell By Date Modelling Dump

In the design section we outlined which variants of Cell By Date we would like to implement in different Chassis, as shown in the table below.

 Chassis Level 1 Level 2 Level 3 E.coli X In-Vitro X X X In-Veso X X X

In this section we modify the variants of Cell by Date so that they can be implemented advantageously in each chassis. We also model these modified variants in order to understand the behaviour we expect or to tune the system to realise a particular behaviour. This leads us to write up experiments to determine if our system behaves the way our models predict.

## Overview of Modelling

Overall our modelling for this project will take the form of

$\frac{dFP}{dt}=k_{FP}(t)-d_{FP}[FP]$

1. kFP : Function of Temperature. k is based on the promoter used as promoters take time to turn on.
2. dFP : Function of System. May be considered to be a function of temperature as proteins degrade faster at higher temperatures.

With this in mind we will look at two graphs of k vs. time (special pt is kON.) and [FP] vs. time key point is [FP]ss

• Our major problem at the moment is estimating the errors involved with our fluorometer and experimental procedures, most notably pipetting; we hope to address these through calibration curves.

For each experiment we will do the following

1. Calibration curve to determine error in fluorometer
2. Decay Experiment @ varying temperatures
3. Plug together to find transient response and k
4. Find these parameters as a function of temperature(T)

### Construct Specific Modelling

#### For all constitutive promoters (Ptet promoter used as an example):

Construct employing Ptet

1.Apply the above general equation to this specific construct.

$\frac{d[GFP]}{dt}=k_{Ptet}(t)-d_{GFP}[GFP]$

2.Calibration curve to determine error in fluorometer - we are trying to get this information from BertholdTech
3.Decay Experiment at varying temperatues:

• This is to determine dGFP
• The fluorescence of a pure sample of GFP kept at 'constant temperature' will be measured at time intervals. We expect the fluorescence to decay exponentially with time.
• We plan to determine the decay constant in the following way:
• We cannot measure the decay constant directly; instead we plan to measure/determine the half-life of our reporter. With this done we can calculate the decay constant using the following equation:

$t_{1/2} = \frac{\ln 2}{\lambda}$

DecayPage

4.Having found dGFP we look at transient respone of construct at several constant temperatures to find k at constant temperature
5.Find k & d as functions of temperature:

• This will hopefully have been carried out by interpolation of data from 4. and 3. eg. plots of k vs. temp & d vs. temp
• The problem with this method is that it will not allow us to determine the response time of the promoters eg. the time taken for the promoter to respond to a temperature change.

Otherwise, another plan is as follows:

• This plan has have several temperature slopes as inputs from which we can compare changes in temperature with changes in k to determine the relationship between the two.
• This methods involes comparing rate of temp increase to rate of k change eg. substituting time in k expression for Temp.

6.Following on from this we plan to have several pulse inputs, similar to what we'll have in real life eg. taking meat from the supermarket and bringing it back home to put in your fridge. From this we can work out how well our system behaves in real life scenarios, and how well our model performs.

## 27.08.07 PR Motif

The Competition:

MIT : Wiki looks very professional

Team name : Team DeTox , Project : EcoFilter

Butanerds

1.VesoCops , Veso Powers , an the like

These ideas are very immature, as a PR man i thought to wow audiences with a clever/sexy story - orientated at general public.

2.CFS : Cell Free Sensation ----- Project 1 : Cell by Date , Project 2 : Infector Detector

A more mature approach is to be creative but in a more restrictive framework, taking into account the scientific nature of the competition and also the synthetic biology community as a whole.

Wow factor will come from how professionally and maturily we present our ideas so that the community believes in our ideas and investors want to invest in us. No kiddy diagrams no funny stories. Cell Free Sensation is exciting because it's new, it's lucrative and it makes synthetic biology look good.

## Overview of Modelling

Welcome to our Portal Page for the modelling of Infector Detector.

Infector Dectector (ID) is based on the Quorum Sensing Pathway and our aim in modelling of ID is to determine the concentration of AHL in biofilm we can detect such that we report a visible signal .We are looking at two constructs to emulate the quorum sensing pathway:

### Construct 1

The main feature of this construct is that it constitutively expresses LuxR

Here is what our construct looks like composed of biobricks

Ptet promoting LuxR, Plux promoting GFP

Here is a Block Diagram Picture of how construct 1 will work

Construct 1 - LuxR expressed constitutively

### Construct 2

(image will appear here)

The main feature of this construct is that it does not constitutively expresses LuxR and therefore enables us to determine the initial concentration of LuxR

• Block Diagram Picture of how construct will work
• Chemical Equations

 Construct 1 The main feature of this construct is that it constitutively expresses LuxR Here is what our construct looks like composed of biobricks Ptet promoting LuxR, Plux promoting GFP Here is a Block Diagram Picture of how construct 1 will work Construct 1 - LuxR expressed constitutively Construct 2 (image will appear here) The main feature of this construct is that it does not constitutively expresses LuxR and therefore enables us to determine the initial concentration of LuxR Block Diagram Picture of how construct will work Chemical Equations

## Fooling around with columns to make project definition sexy

Introductory text with mabye a fun motif to introduce to applicaitons: cell by date & infector detector which both have columns below.

### Cell By Date

Descriptive text about cell by date

Cell By Date

### Infector Detector

Infector Detector

 Cell by date descriptive text about cell by date Cell By Date Infector Detector Descriptive text about cell by date Infector Detector

## Modelling Dump

### Previous work

Overall our modelling for this project will take the form of

$\frac{dFP}{dt}=k(t)-\delta_{FP}[FP]$

1. k : Function of Temperature. k is based on the promoter used as promoters take time to turn on.
2. dFP : Function of System. May be considered to be a function of temperature as proteins may degrade at high temperatures.
3. FP : The particular fluorescent protein employed e.g. GFP, DsRed, etc.

Two graphs of k vs. time (special pt is ko.) and [FP] vs. time key point is [FP]ss

• Our major problem at the moment is estimating the errors involved with our fluorometer and experimental procedures, including the use of pipettes; we hope to address these through calibration curves.

For each experiment we will do the following

1. Calibration curve to determine error in fluorometer
2. Decay Experiment @ varying temperatures
3. Plug together to find transient response and k
4. Find these parameters as a function of temperature(T)

### Construct Specific Modelling

Ptet promoting LuxR, Plux promoting GFP

Modelled by 2006 Imperial Team, as part of their prey-sensing module of the molecular predation oscillator:
Test construct modelling & general derivation.
Parts page.
An operating/working version is quoted on the parts registry as T9002 - Link to MIT Parts Registry

The major problem with last year's derivation is that they assumed LuxR concentration to be constant and they didn't look into the co-operativity of the Plux Promoter e.g. the threshold of quorum.

• LuxR concentration:

LuxR will be constituitively expressed in our system, this means that the older our system is the more LuxR present and so the less sensitive our system will be to the amount of AHL around.

We can model this by having a initial condition or history of our system in addition to last year's path way.

$[LuxR]_{0}\; at\; t=t_{0}$

$LuxR+AHL\rightarrow A$

$A+P\leftarrow\rightarrow\;PA\;\rightarrow\;GFP$

$\frac{d[LuxR]}{dt}=k-d_{LuxR}[LuxR]-k_{1}[LuxR][AHL]+k_{2}[A]$

We will first consider the concentration of LuxR to be constant as per last year. Following this we will consider the history of the system and so the initial concentration of LuxR when AHL is detected.

• Cooperativity of Plux Promoter:

We do not know what the cooperativity of the Plux Promoter is and in effect we do not know what exactly the threshold of our system will be.

$\frac{d[GFP]}{dt}=\frac{X^a}{1+X^a}$

$X=[LuxR][AHL]\;complex$

a > 1 : initial gradient is zero and we see a step response
a = 1 : response similar to first order eg. no threshold visible we have to define
a < 1 : initial gradient is infinite // to y-axis and mabye we get a step response ???? (is this true)

The threshold for biofilm detection, via AHL quorum sensing molecule is approximately 0.1nM.

#### Feedback from Matty

Overview of modelling this construct:

• Experiment 1 : Get Data - k vs. [AHL]
• Analysis 1 : Determine whether minimum [AHL] neded to get visible level is below target concentration [AHL]t = 0.1nM
• Experiment 1 : Get Data - k vs. [LuxR]
• Analysis 2 : Determine how age of system affects its sensitivity

Breakdown of specific sections:

##### Experiment 1:
1. Determine constant LuxR concentrations to be used
2. Determine Po? (may not be possible)
3. Determine value of rate constants in below derivation
4. Determine Visible GFP level in terms of Fluoresence
5. Convert this level into a concentration
6. Determine error associated with this concentration
7. For constant luxR concentration determine range of AHL concentrations
8. For each AHL concentration record transient resonse esp. Steady State Value (this is the only real value we want) and time taken to reach steady state.
• Protocols said they can Get us either , k vs. [AHL] or [GFP]ss vs. [AHL] for both [LuxR] will be a constant.
##### Analysis 1:
1. We have determine the Menten Kinetics Equations describing our pathway excluding cooperativity to give us insight into what's going on:

$\frac{d[GFP]}{dt}=k_{5}\left\{\frac{[AHL][LuxR][P]_{0}}{K+[AHL][LuxR]}\right\}-\delta_{GFP}[GFP]$

1. plot steady state Value of GFP concentration [GFP] vs. concentration of AHL
2. Draw [GFP] level indicicative of Minimum visible concentration
3. Steady state value is equal to k/delta , using this we can find alpha by comparing to a family of curves
1. $k=\frac{([AHL][LuxR])^\alpha}{1+([AHL][LuxR])^\alpha}$
4. With this plot we can also find the minimum [AHL] needed for a visible [GFP]
5. Compare this with desired value
##### Experiment 2:
• We have not considered this yet
##### Analysis 2:
• We have not considered this yet
##### Considerations
• Modelled deterministically; possibility for additional stochastic modelling.
• Degradation term in model lacks dilution term - look into amending the model to incorporate dilution of specific molecule due to cell growth (dilution)
• HRP system is employed as amplifier of low-level signal (where signal-to-noise ration is high)- amplification here probably amplifies the noise, leading to possible false positives for biofilm presence.
• A potential issue with the experiment is regarding using pTet as the promoter for LuxR expression. It may be necessary to add a gene for TetR expression to ensure control over this promoter. Issue of sensitivity control...

Diffusion coefficient of HSL in water - 4.9*10-6 Daq cm2/s
Diffusion coefficient is approximately half in biofilms [1]

### Case1

Case 1 : Both initial concentrations of LuxR and AHl are controllled

[A] = Kα[AHL]0[LuxR]0

$[AP]=\frac{K_{\alpha}K_{\beta}[AHL]_{0}[LuxR]_{0}[P]_{0}}{1+K_{\alpha}K_{\beta}[AHL]_{0}[LuxR]_{0}}$

$R_{y}(x)=\frac{[AP]}{[P]_{0}}=\frac{K_{\alpha}K_{\beta}xy}{1+K_{\alpha}K_{\beta}xy}$

$R_{y}(x)=1-\frac{1}{1+K_{\alpha}K_{\beta}xy}$

$R_{y}'(x)=\frac{K_{\alpha}K_{\beta}y}{\left(1+K_{\alpha}K_{\beta}xy\right)^2}$

### Case2

Case 3 : Only y is controlled

x = [AHL]0 = [AHL] + [A]

[A] = x − [AHL]

but [A] = Kα[AHL]y

therefore x − [AHL] = Kαy

therefore $[AHL]=\frac{x}{1+K_{\alpha}y}$

$[A]=K_{\alpha}\left(\frac{xy}{1+K_{\alpha}y}\right)$

Now $Ry(x)=\frac{K_{\alpha}K_{\beta}xy}{1+yK_{\alpha}+K_{\alpha}K_{\beta}xy}$

### Construct 1 : Intro

With the chemical pathway below for construct 1 we can make form some initial equations about the system.

• Picture of pathway for construct from Jerry

Our aim in modelling Infector Detector is to determine the concentration of biofilm we can detect such that we report a visible signal. Therefore we want to relate the GFP concentration to the concentration of AHL. Starting that the rate of change in GFP concentration:

$\frac{d[GFP]}{dt}=k_{6}[AP]-\delta_{GFP}[GFP]$

Aussume : [AP] reaches a constant level
Reason : Treat 1st formation of AP complex as a black box - it just reaches steady state.

$\therefore\frac{d[AP]}{dt}=0=k_{4}[A][P]-k_{5}[AP]$

we can infer that $[AP]=\frac{k_{4}}{k_{5}}[A][P]$

Assume : Conservation of Plux Promoters Reason : no damage to Promoters will occour eg. no DNA damage due to old age or cell defence mechanisms attacking the DNA

[P]0 = [P] + [AP] Insertformulahere

### Construct 2 : Case1

Case 1 : Both initial concentrations of LuxR and AHL are controllled

$\displaystyle[A]=K_{\alpha}[AHL]_{0}[LuxR]_{0}$

$[AP]=\frac{K_{\alpha}K_{\beta}[AHL]_{0}[LuxR]_{0}[P]_{0}}{1+K_{\alpha}K_{\beta}[AHL]_{0}[LuxR]_{0}}$

$R_{y}(x)=\frac{[AP]}{[P]_{0}}=\frac{K_{\alpha}K_{\beta}xy}{1+K_{\alpha}K_{\beta}xy}$

$R_{y}(x)=1-\frac{1}{1+K_{\alpha}K_{\beta}xy}$

$R_{y}'(x)=\frac{K_{\alpha}K_{\beta}y}{\left(1+K_{\alpha}K_{\beta}xy\right)^2}$

### Construct 2 : Case 2

Jerry Has this in his sandbox

### Construct 2 : Case3

Case 3 : Only y is controlled

x = [AHL]0 = [AHL] + [A]

[A] = x − [AHL]

but [A] = Kα[AHL]y

therefore x − [AHL] = Kαy

therefore $[AHL]=\frac{x}{1+K_{\alpha}y}$

$[A]=K_{\alpha}\left(\frac{xy}{1+K_{\alpha}y}\right)$

Now $Ry(x)=\frac{K_{\alpha}K_{\beta}xy}{1+yK_{\alpha}+K_{\alpha}K_{\beta}xy}$

## Milestones for PR on week basis

1. Motif
2. Tshirt / Logo / Photo : Finalise Defn of Wiki / Russell Peters
3. Slide Layout : Finalise Tshirt / Logo / Photo
4. Contact Tom Millar
5. Drinks
6. Champange reception

## Imperial's 2007 iGEM Project Definition

### VesoCops - Imperial College iGEM 2007 Team ** Work in Progress We're making VesoCop!!! **

Infector Detector

The Imperial College iGEM 2007 team consists of ten undergraduate bioengineering and bioscience students. This year, we are engineering VesoCops, biological systems that report the presence of nasty bacteria. Under the Cell-Free Intelligence (CFI), we have two divisions. First, a surveillance team called Cell By Date that determines when food is spoilt more accurately than printed sell by dates. It exploits the thermal dependence of the rate of expression of a simple reporter system. The second division consists of an undercover team - Infector Detector, which detects biofilms that are antibiotic-resistant and a major source of infection in hospitals. This system makes use of Lux quorum sensing to eavesdrop on the communication between biofilm-forming bacteria.

Cell By Date

Our contributions to the synthetic biology community will be the characterization of Cell-Free Chassis, the common platform on which Cell By Date and Infector Detector will be built. The cell-free approach is particularly useful for VesoCops to operate in the food and medical industries. We believe this new chassis will unlock fresh potential in simple constructs. Our project strategy is based on the Engineering Cycle, of which we have completed specification and design of the systems. We are starting on modelling and implementation and we aim to test our final constructs in the new chassis. By the end of the summer, the VesoCops will be combat-ready.

The Story:

We approach synthetic biology with a view of making bacteria help us. The media is rife with stories of Synthetic Biology being a threat as it could make the next super weapon and how bacteria are bad and make us sick. We want bacteria that serve us, to inspire confidence in Synthetic biology, and that protect us from these daily threats to show that bacteria can be well behaved. The idea of to protect & serve leads us to the idea of a cop , a bactocop of you will.

The Project:

BactoCops, Imperial College’s iGEM 2007 project, delivers a new breed of grime fighting officers. The current state of affairs is that only the run of the mill BactoCops are in operation, the make up the BAPD is you will. These BactoCops can’t do a lot because they use E.Coli as a chassis – they can’t be near open wounds or near our food for example. We are focusing on bringing a new type of agency to the BactoCop world consisting of SuperBactocops – we call it the CIA : the Cell Free Intelligence Agency. This new agency has unbelievable potential and to demonstrate this we have two amazing teams. Firstly, a surveillance team called Cell By Date that determines when food is spoilt more accurately than printed sell by dates. It exploits the thermal dependence of the rate of expression of a simple reporter system. Our second team goes undercover, codename - Infector Detector, it detects biofilms that are antibiotic-resistant and a major source of infection in hospitals. This system makes use of Lux quorum sensing to eavesdrop on the communication between biofilm-forming bacteria.

 Infector Detector Cell By Date

Our contributions to the synthetic biology community will be the characterization of Cell-Free Chassis, the common platform on which Cell By Date and Infector Detector will be built. The cell-free approach is particularly useful for BactoCops to operate in the food and medical industries. This is because living, replicating engineered bacteria pose major health risks. We believe this new chassis will unlock fresh potential in simple constructs.

The Progress:

Our project strategy is based on the Engineering Cycle, of which we have completed specification and design of the systems. We are starting on modelling and implementation and we aim to test our final constructs in the new chassis. By the end of the summer, the BactoCops will be combat-ready.

Re-Revised Project dEscription corrections:

1. Remove BactoCops

2. Make Biofilm Detector Picture look more medical

My Project Description Corrections:

1. Story needs to be re-written it paints syn bioin a very negative light

2. Cell by date doesn't report the presence of nasty bacteria

3. BactoCops is misleading because we are not using bacteria

Possible way forward is to extend the RoboCop / VesoCop Idea

### Potentials for Motifs

1. Mr.Bean
2. James Bond
3. Dr.Who
4. Austin Powers
5. Sherlock Homes
6. Robin Hood
7. Alice in Wunderland
8. The Queen
9. MI6
10. Scotland Yard
11. Inspector Morse
12. The Bill
13. Bobbies
14. Red Phone Box
15. The Avengers
16. Red Bus
17. Royal Guards
19. Repressed Women
20. The Beetles
21. Oasis
22. Spice Girls
23. David Beckham
24. Monty Python
25. Billy Conelly
26. Beer
27. Drunkedness
28. The InVesible Man

#### James Bond

We'll be Q-Branch

1. Bond as human get new gadget which is our project cell by date / infector detector

2. Bond is a cell : he goes through transformation and becomes a super agent

Project Description:

Projects are missions

1. Cell by Date :

vehicle sounds like vesicle

James Bond 007 : Licence to Glow

Vesper Vesicle - Bond's no. 57 on the bang list.

Pussy Galore - Lipids Galore

Octopussy - Spherical Pussy - Pleasure from all angles

#### Revised VesoCop Story

1. Back Story - how VesoCop came to be
2. VesoCop in action - Project Definition

The Story: Current state of affairs is that syn. Bio has made tremendous progress in terms of making Bacteria Do wonderful things for us protecting us in Variety of situations, they behave like the police men around us they are if you will BactoCops. But these bactocops can't do everything, with E.Coli as a Chassis they are ineffective in certain situation. There is a need now for a new breed of BactoCop that can go to these places, further than any BactoCop before him : we can him VesoCop The Project: The Progress:

1. Potential as a motif - Look forward to slide layout / presentation / Logo / Tshirt Design

#### Austin Veso Story

• Feedback from Group on Austin Veso:
• Jerry likes big chicks
• Teeth need changing - make them whiter less crooked
• Colar needs fixing
• Consider 3/4 view
• Text is too bland - make it in crazy 60s theme
1. Back Story - how Austin Veso came to be
2. Austin Veso in action - Project Definition

The Story:
Synthetic Biology has amde awe inspiring Progress to date : it has helped to produce a malaria vacine and by the the looks of previous iGEM it will improve our lives in many other ways(Need more wow! milestones here). We feel that we can contribute to this amazing movement through the exploration of Cell Free Chasis. In Looking into Cell Free Chassis we think we've come up with something special we call him Veso Powers: International Bag Of Lipids
The Project:
Veso Powers: International Bag of Lipids is the ultimate spy, using his cell free mojo he can do amazing things. Check out two incredible missions that Veso Powers will be able to carry out by the end of summerof after he has completed our rigerous trainging programme in the ways of the Mojo at our super secret HQ in london.
Mission 1: Thermal Steakout
Veso Powers' First mission involves our food. Using his special Cell By Date ability he can determine when food is spoilt more accurately than printed sell by dates. Veso Powers Cell By Date ability exploits the thermal dependence of the rate of expression of a simple reporter system.

Mission 2: A lot of Biofilm's Penthouse
Austin Veso Second Mission is to go undercover, using his Infector Detector ability. With this ability Veso Powers can detect biofilm that are antibiotic-resistant and a major source of infection in hospitals. The Infector Detector ability system makes use of Lux quorum sensing to eavesdrop on the communication between biofilm-forming bacteria.

Through Veso Powers our contributions to the synthetic biology community will be the characterization of Cell-Free Chassis, the common platform on which Cell By Date and Infector Detector will be built. The cell-free approach is particularly useful for Veso Powers to operate in the food and medical industries. We believe this new chassis will unlock fresh potential in simple constructs.

The Progress:
Our project strategy is based on the Engineering Cycle, of which we have completed specification and design of the systems. We are starting on modelling and implementation and we aim to test our final constructs in the new chassis. By the end of the summer, Veso Powers will have his mojo in full swing and will be mission ready - Yeah Baby !.

1. Potential as a motif - Look forward to slide layout / presentation / Logo / Tshirt Design
Picture5

#### Sherlock Veso Story

1. Back Story - how VesoCop came to be
2. VesoCop in action - Project Definition

The Story: Current state of affairs is that syn. Bio has made tremendous progress in terms of making Bacteria Do wonderful things for us protecting us in Variety of situations, they behave like the police men around us they are if you will BactoCops. But these bactocops can't do everything, with E.Coli as a Chassis they are ineffective in certain situation. There is a need now for a new breed of BactoCop that can go to these places, further than any BactoCop before him : we can him VesoCop
The Project:
The Progress:

1. Potential as a motif - Look forward to slide layout / presentation / Logo / Tshirt Design

Pictures:

Picture5

asdfasdfasdf

text below picture

Anchoring

Citations for PubMed

[1, 2]

Internal : internal

External : ign

Time Stamp

Anthony Lazzaro 11:23, 12 July 2007 (EDT)

Search Term site :openwetware.org

Well Characterised Part in Registry:

Recievers:

BBa_F2620 :

• Input: 3OC6HCL
• Output: Pops

Summary of Brainstorming 18th July:

Hrp has promise as a black box feedback loop mechanism

• Application Thermoregulation
• General application to a system that needs feedback loop - duh!

Hrp has promise as a very specific switch

Other Promising Ideas:

Lucas: Bacteria that move away from a source

• Could expand to bacteria that move away or towards a chemical source & incorporate feedback loop seen in signals
• Possible conflict with chemotaxis (?) from last year need to check could reuse there parts if no conflict

Density flow receptor: Not only detect presence of bacteria but also how much of it there is a particular location
eg.how much dust here compared to here

• Need some kind of measurement device or quorum sensing set up (so more stuff more cells more reporter)

Uses for Hrp System: (Blood testing)

Immediate blood-test results can mean the difference between life and death in medical situations such as poisonings and infectious diseases. In many cases, by the time the test results are analyzed and a diagnosis made, the patient is in critical condition or beyond treatment

Blood poisoning is an illness due to an infectious agent or its toxin spreading through the bloodstream. The presence of bacteria in the blood is called bacteremia.

Short bursts of low levels of bacteria in the blood usually do not cause problems. For example, mild bacteremia typically occurs during a dental cleaning or when brushing your teeth. Your body's immune system fights off these bacteria.

If bacteria persist in the blood, however, they may cause sepsis, a serious, life-threatening condition.

• Slovenia has already covered this what other blood test could we do ?

20th July

### Further Brainstorming for new ideas:

All my ideas have been rejected so far

Avenues to explore for new ideas:

1.Pre-2006 iGEMs

• 2005:

##### 1.Easy Application

-Make Bacteria Bind to something when told to do so

• E. coli MC1061 synthesize type I pili encoded by the fim (fimbria) operon, which bind to

mammalian surface carbohydrates [2]

• We could use this to simply bind bacteria to carbohydrates when the operon is expressed

(eg. simple promoter activation)

• 'The ability of enteropathogenic Escherichia coli (EPEC) to form attaching and effacing intestinal lesions is a major characteristic of EPEC pathogenesis' a group has identified a chromosomal gene (eae, for E. coli attaching and effacing) that is necessary for this activity.[3]
Error fetching PMID 16330045:
1. Error fetching PMID 16330045: [1]
Error fetching PMID 2172966:
1. Error fetching PMID 2172966: [2]
##### 2.Technical Use

-Determine how many bacteria are attached to target

##### 3.Real World Application

-Determine A certain concentraion of a species on a surf

#### Focus on Biofilm:

##### 1.Easy Application

Secrete Biofilm when inducer given:

• Chemical Control of Biofilm Production [taken from MIT 2006 Wiki] :

1. Oxindolyl-L-alanine inhibits, in a dose-dependent manner, indole production and biofilm formation by strain S17-1 grown in Luria-Bertani (LB) medium. Supplementation with indole at physiologically relevant concentrations restores biofilm formation by strain S17-1 in the presence of oxindolyl-L-alanine and by mutant strain E. coli 3714 (S17-1 tnaA::Tn5) in LB medium
2. Here are 2 more articles on the mutant indole-negative e. coli strain 3714
• I hope that we can get mutant strain 3714 -- (or make our own mutant) -- what do you guys think about e. coli biofilm and cell-cell signaling systems/applications?? anyways, take a look...
• Error fetching PMID 14569285:
1. Error fetching PMID 14569285: [Martino]
• Error fetching PMID 11958566:
1. Error fetching PMID 11958566: [Martino]
##### 2.Technical Use
• Apparantly biofilm production is initiated when bacteria adhese to a surface through a CPX signalling pathway the genes encoding for this pathway have been found.[2]
Error fetching PMID 11830644:
1. Error fetching PMID 11830644: [1]

### 22nd July Last Minute Susan Time

Major Focus on Biofilm

##### 1.Easy Application

Secrete Biofilm when inducer given:

• Chemical Control of Biofilm Production [taken from MIT 2006 Wiki] :
• Oxindolyl-L-alanine inhibits, in a dose-dependent manner, indole production and biofilm formation by strain S17-1 grown in Luria-Bertani (LB) medium. Supplementation with indole at physiologically relevant concentrations restores biofilm formation by strain S17-1 in the presence of oxindolyl-L-alanine and by mutant strain E. coli 3714 (S17-1 tnaA::Tn5) in LB mediumlink. Problem with this is that we need to get this mutant e. coli 3714 strain?
• 2-Component System Control:
• Apparantly biofilm production is initiated when bacteria adhese to a surface through a CPX signalling pathway the genes encoding for this pathway have been found.[5]
##### 4.Modular Real World Application

Papers:

Error fetching PMID 14569285:
1. Error fetching PMID 14569285: [Martino]
Error fetching PMID 11958566:
1. Error fetching PMID 11958566: [Martino]
Error fetching PMID 11830644:
1. Error fetching PMID 11830644: [1]

### Quick note on Noise reduction in iGEM

I think one way to reduce noise & cross talk in iGEM projects is to have 'comm channels'

'Channels' used by the chassis should be chara