Team:DTU Denmark/USERprogramoutputformat

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   The redoxilator<br><br>
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   The redoxilator<br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/theory" CLASS=leftbar>- Introduction</a><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/genetic_design" CLASS=leftbar>- Genetic design</a><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/yeast" CLASS=leftbar>- Results</a><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/applications" CLASS=leftbar>- Applications and perspectives</a><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/practicalapproach" CLASS=leftbar>- Applications and perspectives</a><br><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/safety" CLASS=leftbar>- Safety considerations</a><br>
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     <br>The USER<sup>TM</sup> assembly standard<br><br>
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     <br>The USER assembly standard<br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprinciple" CLASS=leftbar>- Principle</a><br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprinciple" CLASS=leftbar>- USER fusion of biobricks</a><br>
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    <a href="https://2009.igem.org/Team:DTU_Denmark/USERconcept" CLASS=leftbar>- Proof of concept</a><br>
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    <a href="https://2009.igem.org/Team:DTU_Denmark/USERmanual" CLASS=leftbar>- Manual</a><br><br>
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     <br>USER<sup>TM</sup> fusion primer design software<br><br>
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     <br>USER fusion primer design software<br>
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     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprogram" CLASS=leftbar>- Abstract</a><br>
     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprogram" CLASS=leftbar>- Abstract</a><br>
     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprograminstructions" CLASS=leftbar>- Instructions</a><br>
     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprograminstructions" CLASS=leftbar>- Instructions</a><br>
     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprogramoutputformat" CLASS=leftbar>- Output format</a><br>
     <a href="https://2009.igem.org/Team:DTU_Denmark/USERprogramoutputformat" CLASS=leftbar>- Output format</a><br>
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    <br>Experimental results<br>
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    <a href="https://2009.igem.org/Team:DTU_Denmark/results" CLASS=leftbar>- Results and discussion</a><br>
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   <b>The project</b><br><br><br>
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   <b>The project</b><br><br>
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<font size="4"><b>USER fusion primer design software</b></font><br><br>
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<font size="4"><b>The USER fusion primer design software: PHUSER<br>(<u>P</u>rimer <u>H</u>elp for <u>USER</u>)</b></font><br><br>
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<font size="3"><b>Abstract</b></font><br>
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<font size="3"><b>PHUSER output format</b></font><br>
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<p align="justify">When designing constructs with more than two biobricks using USER(TM) fusion, it is essential to avoid identical fusion tails to ensure correct order of the biobricks. Furthermore, the DNA denaturation temperature (T<sub>M</sub>) of the primer fragments must be pairwise within 2<sup>o</sup>C degrees, for PCR amplication of the biobricks. Selection of optimal fusion tails is achieved by employing a simple, but powerful sorting algorithm utilizing the fact that the relative penalty for increasing length of, and shifting center of fusion regions, is the same, i.e. one base added/removed from final primers. Adjusting the T<sub>M</sub> of primer pairs is done by randomly sampling the various allowed lengths of the primers (18-24 bases), thus changing the CG ration and affecting T<sub>M</sub>, until an acceptable solution is achieved. The suggested primers are presented in a clear and intuitive fashion, diplaying both list view and a graphical overview of fusion regions and related primers. The program is tested to handle primer design for constructs with between 2 and 9 biobricks at the time, but in theory, if enough unique fusion tails exists, many more biobricks can be fused in the same reaction.</p><br>
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<p align="justify">Like the input options, the output from PHUSER is kept simple. First an overview of all needed primers is shown. The output is fairly self-explanatory, although it should be noted that primer consist of a fusion tail (ft) and a binding region (br), easily distinguished from one another by the occurence of the essential uracil. For example:</p><br>
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<a href="https://2009.igem.org/Team:DTU_Denmark/USERprogram" CLASS=leftbar>Design your primers here</a><br><br>
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|--ft--| |-------br-------|<br>
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GGGTTAAU CAAATTATAGCCATACAG
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</font><br><br>
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<p align="justify"><i>Article submitted October 2009 - publication pending</i></p>
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<p align="justify"><b>IMPORTANT: Note that the entire sequence constitutes the primer. The space is simply added for illustrative purposes, and should be omitted when synthesizing the primers</b><br><br>
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Just below is the grafic overview of biobricks and primers. The first block illustrates the forward primer for biobrick 1, i.e. binding region to biobrick 1, and fusion tail to the selected USER cassette. The following blocks (the number depends on how many biobricks were entered), illustrate the region at which biobrick 1 is fused to biobrick 2 and so forth. The last 40 bases of biobrick 1 and the first 40 bases of biobrick 2 are displayed, along with the positioning of the fusion tails and binding regions for both reverse and forward primers. The last block of the graphic overview illustrated the last biobrick and its binding region to the selected cassette, diplayed in a similar fashion as described for the first block.<br><br>
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Lastly, the primers are displayed again, only this time with more detailed information, such as T<sub>M</sub> and GC ratio. More parameters are to be added to the output.<br><br>
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<b>Design your primers with PHUSER <a href="http://igem.grafiki.org/" CLASS=leftbar target="blank">here</a></b><br><br>
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  <b>Synthetic Biology</b><br><br>
 
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<p align="left"><i>“Synthetic Biology is an art of engineering new biological systems that don’t exist in nature.”</i><br></p>
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<strong>Achievements</strong><br>
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<p align="right"><i>-Paras Chopra & Akhil Kamma</i><br><br></p>
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<br>
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<strong>Redox sensing device</strong><br>
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Two novel genes have been designed and synthesized each comprised of 5 genetic elements. Together they function as a device termed the Redoxilator that can sense the internal redox state of a yeast cell, and output a reporter signal. Extensive mathematical modelling was performed to simulate how the construct would operate <i>in vivo</i>.<br>
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<br>
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<strong>Biobricks</strong><br>
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DNA of several new biobricks have been designed and submitted including a yeast optimized GFP reporter protein, a protein degradation sequence and a fast degradable yeast GFP. (Bronze medal)<br>
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<br>
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We have demonstrated that our USER fusion biobrick works as expected and documented it (silver medal)<br>
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<br>
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<strong>USER fusion Assembly standard</strong><br>
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A new biobrick assembly standard that allows the rapid construction of multi-part devices have been developed and documented. The assembly standard offers many benefits: All restriction sites are allowed, multiple biobricks can be joined in one step, the result is scar-free making it ideal for protein fusions and more. (Gold medal)<br>
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<br>
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<strong>USER-fusion primer design software</strong><br>
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A novel and very useful software tool have been developed that can automatically design the optimal primers for USER fusion assembly of 2-9 biobricks, taking several parameters into account.
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</p>
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<p>In nature, biological molecules work together in complex systems to serve purposes of the cell. In synthetic biology these molecules are used as individual functional units that are combined to form tailored systems exhibiting complex dynamical behaviour. From ‘design specifications’ generated from computational modelling, engineering-based approaches enables the construction of such new specified gene-regulatory networks. The ultimate goal of synthetic biology is to construct systems that gain new functions, and the perspectives of the technology are enormous. It has already been used in several medical projects2 and is predicted to play a major role in biotech-production and environmental aspects.</p>
 
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Latest revision as of 03:17, 22 October 2009

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The project

Home The Team The Project Parts submitted Modelling Notebook


The redoxilator

- Genetic design
- Applications and perspectives
- Safety considerations


The USER assembly standard

- USER fusion of biobricks


USER fusion primer design software

- Abstract
- Instructions
- Output format


Experimental results

- Results and discussion
The project

 The USER fusion primer design software: PHUSER
(Primer Help for USER)

PHUSER output format

Like the input options, the output from PHUSER is kept simple. First an overview of all needed primers is shown. The output is fairly self-explanatory, although it should be noted that primer consist of a fusion tail (ft) and a binding region (br), easily distinguished from one another by the occurence of the essential uracil. For example:


|--ft--| |-------br-------|
GGGTTAAU CAAATTATAGCCATACAG

IMPORTANT: Note that the entire sequence constitutes the primer. The space is simply added for illustrative purposes, and should be omitted when synthesizing the primers

Just below is the grafic overview of biobricks and primers. The first block illustrates the forward primer for biobrick 1, i.e. binding region to biobrick 1, and fusion tail to the selected USER cassette. The following blocks (the number depends on how many biobricks were entered), illustrate the region at which biobrick 1 is fused to biobrick 2 and so forth. The last 40 bases of biobrick 1 and the first 40 bases of biobrick 2 are displayed, along with the positioning of the fusion tails and binding regions for both reverse and forward primers. The last block of the graphic overview illustrated the last biobrick and its binding region to the selected cassette, diplayed in a similar fashion as described for the first block.

Lastly, the primers are displayed again, only this time with more detailed information, such as TM and GC ratio. More parameters are to be added to the output.

Design your primers with PHUSER here

Achievements

Redox sensing device
Two novel genes have been designed and synthesized each comprised of 5 genetic elements. Together they function as a device termed the Redoxilator that can sense the internal redox state of a yeast cell, and output a reporter signal. Extensive mathematical modelling was performed to simulate how the construct would operate in vivo.

Biobricks
DNA of several new biobricks have been designed and submitted including a yeast optimized GFP reporter protein, a protein degradation sequence and a fast degradable yeast GFP. (Bronze medal)

We have demonstrated that our USER fusion biobrick works as expected and documented it (silver medal)

USER fusion Assembly standard
A new biobrick assembly standard that allows the rapid construction of multi-part devices have been developed and documented. The assembly standard offers many benefits: All restriction sites are allowed, multiple biobricks can be joined in one step, the result is scar-free making it ideal for protein fusions and more. (Gold medal)

USER-fusion primer design software
A novel and very useful software tool have been developed that can automatically design the optimal primers for USER fusion assembly of 2-9 biobricks, taking several parameters into account.

Comments or questions to the team? Please -- Comments of questions to webmaster? Please

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