Difference between revisions of "Team:UNOTT/Description"

 
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<p><span style="color: #ffffff;">&nbsp;</span></p>
 
<p><span style="color: #ffffff;">&nbsp;</span></p>
 
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<h1>What?</h1>
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<p><b><i>Key. coli</i> provides a new, more secure, form of key for accessing content.</b> It uses random ligations and large repertoires of possible components to generate unique combinations of expression profiles; this next generation biological key could be the next BIG thing in security; watch this space!
  
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<h3 style="text-align: center;"><span style="color: #D74214;"><strong><em>KEY. COLI </em>- PROTECT YOUR GEMS WITH GERMS!</strong></span></h3>
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<p style="text-align: center;"><span style="color: #ffffff;">________________</span></p>
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<h3 style="text-align: center; color: #339966;"><strong>Why do we need <em>Key.&nbsp;coli?<strong></em></h3>
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<img class="ourkey" src="https://static.igem.org/mediawiki/2017/3/36/T--UNOTT--keyk.png" style="width:40%;height:auto;">
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<h1>Why?</h1>
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<p><b>Major hacking incidents are increasingly common,</b> with accounts being hacked and sensitive information stolen. Many companies are moving away from conventional passwords, which are proving to be unreliable in the hands of the public. Banks are now using physical biometric authentication procedures to correctly identify account owners. This new direction opens a market for biological “passwords”. An ideal system would be as separate from online software programs as possible while maintaining the complexity and uniqueness of a biometric system. Cells are effectively living computers, so we can programme cells to act as a changeable biometric password. </p>
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<h1>How?</h1>
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<p><b>A key must be unique, measurable and unpredictable.</b> In <i>Key. coli</i>, all these requirements are achieved by the random generation of modular vectors that are expressed in Escherichia coli to produce a unique and detectable fluorescent pattern. This pattern is obtained when different fluorescent proteins (GFP, RFP, CFP) and various promoters, subjected to transcription interference by dCas9, are randomly combined during ligation and transformed into the cells to generate the key. </p>
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<p><span style="color: #ffffff;">&nbsp;</span></p>
 
<p><span style="color: #ffffff;">&nbsp;</span></p>
 
<p><span style="color: #ffffff;">&nbsp;</span></p>
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<p>Lately, many critical issues surrounding digital password security have surfaced. Companies are turning to physical security strategies involving biometric and digital keys to secure their clients’ accounts. With the tools available in synthetic biology, there’s significant opportunity to provide secure, synthetically generated biometrics. Consequently, our team developed a randomly assorting fluorescent bacterial key. Three fluorescent proteins are expressed using different promoters, with each capable of being inhibited by dCas9. Null and functioning sgRNAs were given identical restriction sites to compete with one another when ligating, giving an ON/OFF fluorescent state and variance for distinguishable keys. Each combination is modelled from lab data, illustrating discernibility. We’ve designed a safe, portable device for storage of the E. coli, paired to a streamlined authentication procedure tailored to be unsusceptible to current hacking frameworks. This system is scalable to include any type of protein, synonymously expanding the number of combinations and improving security.
 
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<h3 style="text-align: center; color: #339966;"><strong>What is our Project's aim?<strong></em></h3>
 
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<p>Our project aim is to provide a platform for multiple proteins to be expressed simultaneously at different levels. dCas9 will be targeted to repress a given promoter preceding a protein by using a short guide RNA (sgRNA) corresponding to this promoter. Our initial proof of concept system uses fluorescent proteins as the reporter signal and we have taken five promoter-sgRNA pairings from literature and constructed two plasmids which will give many different combinations of protein levels. One plasmid contains three promoters (although this will be expandable) joined up to three different fluorescent proteins. This plasmid will also express dCas9, which we will submit to the iGEM registry. The second plasmid will express 3 different sgRNAs which target each one of these promoters and, by having the option of a non-targeting sgRNA too, we have ON/OFF switches for these fluorescent proteins. In the future, we can expand this to different levels, rather than just two, by creating mismatches in the sgRNA seed region, which has been shown to reduce efficiency of repression by dCas9. Discernible combinations would only be limited by the reproducibility of signals and accuracy of the measurement.
 
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For future expansion, although there is a vast repertoire of fluorescent proteins, any other protein could also be substituted in and measured using other methods. In this way, this system could be used for a vast range of applications, from optimising production of proteins or a metabolic pathway to being used as a biological password. As our assembly method is all interchangeable so any protein can be linked to any promoter, and placed in any position in the plasmid.
 
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We have chosen to focus our project on using this random assortment to create a natural random number generator, with application as a biological key as we felt this is the most exciting to the public and shows the design’s mechanism the best. We envisage a system where these plasmids can be assembled randomly (as this is how we designed our system) to produce an enormous number of combinations, which is a valuable characteristic in security. The potential impact from this application is that we will be providing a new, more secure, form of key for accessing content.
 
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This is a hot topic at present; numerous major hacking incidents fixate online security at the forefront of many business’s concerns, as accounts are being hacked and sensitive information stolen. Many large companies are deviating away from conventional online character passwords, which are proving to be unreliable in the hands of the public. For instance, many banks are now developing physical biometric authentication procedures to correctly identify the true owner of an account. This new direction opens a market for biological “passwords”. An ideal system would be as decoupled from online software programs as possible, while maintaining the complexity and uniqueness of a biometric system. Cells are effectively living computers so we can programme cells to act as a changeable biometric password. Our product would have no human influence over the outcome of combinations due to our design, making it less prone to issues commonly associated with human carelessness such as using simple, easy to guess passwords.
 
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This system combines aspects comparable to the traditional mechanical key, a digital key, and biometric keys - a fourth alternative to the paradigm. Also, this track of research will hopefully be of interest to many and stimulate further research into developing tools for advanced biological computation. The parallels between computation and genetic regulation are astounding, unlocking the potential for fully realised programmable genetic systems would be invaluable to society in numerous ways.
 
  
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For the physical product, we plan on freeze-drying our “programmed” bacteria for storage in a device. We will optimise the conditions for this to optimise reproducibility and allow accurate authentication. The two signals need to have as low a variance as possible, otherwise the accuracy and completeness of emission signals is invalidated and fewer discernible combinations are possible. Therefore, we are using the data collected to form models to predict the outcomes of fluorescence under all possible conditions, and allow each key to be categorised separately by their output.</p>
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<img src="https://static.igem.org/mediawiki/2017/8/84/UNOTT2017-How1.png" alt="" width="100%" height="100%">
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<h5><b> Figure 1:</b> Two-plasmid modular process used to generate random<i>Key. coli</i> construct(s)<p>
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<p><span style="color: #ffffff;">&nbsp;</span></p>
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<p><span style="color: #ffffff;">&nbsp;</span></p><p><span style="color: #ffffff;">&nbsp;</span></p>
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<p><span style="color: #ffffff;">&nbsp;</span></p>
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<img src="https://static.igem.org/mediawiki/2017/b/bc/UNOTT2017-How2.png" alt="" width="100%" height="100%"></h5>
  
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<h5><b>Figure 2:</b> Random ligation process and colony picking allows large numbers of plasmid variants to be created for use in keys. </p> <p><span style="color: #ffffff;">&nbsp;</span></p></h5>
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<p><span style="color: #ffffff;">&nbsp;</span></p>
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<br>
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<p>A key transport device, based on freeze-dried <i>Key. coli</i>, allows the bacteria to survive and be transported anywhere with ease. Once entry to a lock is desired, the <i>Key. coli</i> device can be activated, and the output read in a suitable detection device. </p>
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<h1><i>Key. coli</i> Summary</h1>
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<p><img src="https://static.igem.org/mediawiki/2017/8/83/UNOTT2017-summary.png" alt="" width="100%" height="auto" /></p>
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Latest revision as of 03:45, 2 November 2017

 

 

 

PROJECT DESCRIPTION

 




What?

Key. coli provides a new, more secure, form of key for accessing content. It uses random ligations and large repertoires of possible components to generate unique combinations of expression profiles; this next generation biological key could be the next BIG thing in security; watch this space!

Why?

Major hacking incidents are increasingly common, with accounts being hacked and sensitive information stolen. Many companies are moving away from conventional passwords, which are proving to be unreliable in the hands of the public. Banks are now using physical biometric authentication procedures to correctly identify account owners. This new direction opens a market for biological “passwords”. An ideal system would be as separate from online software programs as possible while maintaining the complexity and uniqueness of a biometric system. Cells are effectively living computers, so we can programme cells to act as a changeable biometric password.

How?

A key must be unique, measurable and unpredictable. In Key. coli, all these requirements are achieved by the random generation of modular vectors that are expressed in Escherichia coli to produce a unique and detectable fluorescent pattern. This pattern is obtained when different fluorescent proteins (GFP, RFP, CFP) and various promoters, subjected to transcription interference by dCas9, are randomly combined during ligation and transformed into the cells to generate the key.

 

 

Figure 1: Two-plasmid modular process used to generate randomKey. coli construct(s)

 

 

 

 

Figure 2: Random ligation process and colony picking allows large numbers of plasmid variants to be created for use in keys.

 

 


A key transport device, based on freeze-dried Key. coli, allows the bacteria to survive and be transported anywhere with ease. Once entry to a lock is desired, the Key. coli device can be activated, and the output read in a suitable detection device.

Key. coli Summary