Team:DTU-Denmark/Project

From 2010.igem.org

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<li><a href="https://2010.igem.org/Team:DTU-Denmark/Regulatory_sytems">Regulatory Systems</a></li><br>
<li><a href="https://2010.igem.org/Team:DTU-Denmark/Regulatory_sytems">Regulatory Systems</a></li><br>
<li><a href="https://2010.igem.org/Team:DTU-Denmark/Switch">The Switch</a></li><br>
<li><a href="https://2010.igem.org/Team:DTU-Denmark/Switch">The Switch</a></li><br>
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<li><a href="https://2010.igem.org/Team:DTU-Denmark/SPL">Synthetic Promoter Library</a></li><br>
<li><a href="https://2010.igem.org/Team:DTU-Denmark/Team1">Anti-Repressor group</a></li><br>
<li><a href="https://2010.igem.org/Team:DTU-Denmark/Team1">Anti-Repressor group</a></li><br>
<li ><a href="https://2010.igem.org/Team:DTU-Denmark/Team2">Anti-Terminator group</a></li><br>
<li ><a href="https://2010.igem.org/Team:DTU-Denmark/Team2">Anti-Terminator group</a></li><br>
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== Synthetic promoter library (SPL) ==
 
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How does it work, examples, what have it been used to characterize?
 
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how do you construct it? Figures and illustrations to explain.
 
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Figures to explain our use? And example on our specific design primer sequences illustration on the double stranded DNA, with BB - prefix suffix.
 
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= Characterizing BBricks as parts of the switch =
 
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'''(Materials and Methods)''' section<br>
 
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Mainly the hard control of the switch is due to a double regulation system build on a both terminator-anti-terminator and repressor anti-repressor regulation. It was out of the scope of this project to construct the entire theoretical developed switch, and characterize the fully constructed switch. Have focused on characterizing the two regulatory systems individually. This was done in order to investigate if the responses were satisfactory to use in a future complete switch construction. (????  By getting the regulatory mechanism of the subparts we further, by modeling, could conclude constraints for successful function of the system and other subparts.
 
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In this section we describe the design of and the experimental setup used to characterize the subparts of the system and our bio-bricks.
 
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== Anti-terminator function ==
 
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(experimental work)
 
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==== selecting subparts ====
 
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Why were these subparts chosen ?<br>
 
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AIM
 
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==== Design and experimental setup ====
 
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'''presentation - Figure of setup and explanation'''
 
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==== Materials and methods ====
 
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HOW ? what plasmids and why, what measurering method and why?
 
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refer to the notebook page with protocols - and actual info from lab.
 
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'''mRNA-stability'''
 
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when introducing non-coding sequences problems will acour  with rna-degredation of RNAP is not attracted to the area, to fast degredation, unwanted steam loops. (Reference to the terminator screening plasmids for BB)
 
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'''Synthetic promoter library (SPL)'''<br>
 
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How does it work, examples, what have it been used to characterize?
 
-
how do you construct it? Figures and illustrations to explain.
 
-
Figures to explain our use? And example on our specific design primer sequences illustration on the double stranded DNA, with BB - prefix suffix.
 
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==== Results ====
 
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comments to the results and reference to the BB pages with info and results.
 
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== Repressor function ==
 
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(experimental work)
 
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==== selecting supparts ====
 
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Why were these supparts chosen ?<br>
 
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AIM
 
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==== Design and experimental setup ====
 
-
'''presentation - Figure of setup and explanation'''
 
-
 
-
==== Materials and methods ====
 
-
HOW ? what plasmids and why, what measurering method and why?
 
-
refer to the notebook page with protocols - and actual info from lab.
 
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==== Results ====
 
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comments to the results and reference to the BB pages with info and results.
 
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== References ==
== References ==

Revision as of 12:41, 6 October 2010

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Welcome to the DTU iGEM wiki!

Background

What is Synthetic Biology?

Not many people have heard of Synthetic Biology. Synthetic biology essentially aims to utilize natures tricks to design and build artificial biological systems for engineering purposes as well as a way to get a better understanding of why biological systems are set up as they are. The term “synthetic biology” was first used on genetically engineered bacteria that were created with recombinant DNA technology. Parts from natural biological systems are taken, characterized and simplified and used as a component of a highly unnatural, engineered, biological system. The term was then used when referring to when organic synthesis is used to generate artificial molecules that mimic natural molecules such as enzymes.

There are two types of synthetic biologists:
  • those who deal with re-designing and fabricating existing biological systems.
  • those who deal with designing and fabricating biological components that do not already exist in the real world.

Synthetic biology provides us with a new perspective from which we can understand and ultimately utilize life for our own benefits. [1,2,3]



Introduction

Aim

The goal of our project is to enable colonies of ''E. coli'' bacteria to transition between production of two different reporter proteins. In our system, switching between states will be induced by exposing the bacteria to light. Each of the states will have a specific frequency associated with it. There are multiple potential applications for biologicals "switches" such as these, this includes the improved control of production of additives in industrial biotechnological processes.

Project Concept

As previously stated, the main goal of our project is to design a bistable switch. The switching between the two states will be controlled by the introduction of two different wavelengths of light, each wavelength responsible for the induction of a different state. As a proof of concept, we’re using fluorescent proteins as reporter genes which makes it easy to observe and characterise the system. In principle, however, any reporter gene can be used.

Our original project concept revolved around using light-receptors to instigate the switch between the two stable state. It was thought that the production of the first reporter protein would be induced by red light (660 nm). At the same time, production of the other reporter will be suppressed by a coexpressed repressor. Conversely, production of the second reporter would be induced by blue light (470 nm). Bistability of the system is achieved by using two repressors which negatively regulate each other’s expression. This enables the system to sustain state without continuous input, i. e. once production of a reporter protein is initiated, it will persist until the system is forced into the other state.

Our project concept has since changed to using two different carbohydrate sources as a means of switching between the two stable states. This means that the state in which the bacteria will be found depends on which one of two carbohydrate sources it was last exposed.

References

  • (Gottesman et.al. 2002) Gottesman. Max E, Nudler. Evgeny, 2002 ”Transcription termination and anti-termination in E.coli” Genes to cells. (a good introduction review to termination function)
  • (Franklin et.al. 1989) NC Franklin, JH Doelling - Am Soc Microbiol "Overexpression of N antitermination proteins of bacteriophages lambda, 21, and P22: loss of N protein specificity." - Journal of bacteriology, 1989
  • (Jensen 2004) Ole Nørregaard Jensen, “Modification-specific proteomics: characterization of post-translational modifications by mass spectrometry,” Current Opinion in Chemical Biology 8, no. 1 (February 2004): 33-41.
  • [1] http://syntheticbiology.org/FAQ.html
  • [2]http://www.nature.com.globalproxy.cvt.dk/nrg/journal/v6/n7/execsumm/nrg1637.html
  • [3]http://www.nature.com.globalproxy.cvt.dk/msb/journal/v2/n1/full/msb4100073.html