Difference between revisions of "Team:AshesiGhana/Description"

 
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    <specialh2 style="text-transform: lowercase;">Project Description</specialh2> <br><br><br><br><br><br>
 
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<specialh3>Why are Co-cultures useful?</specialh3>
 
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<p>In nature, microorganisms do not exist in isolation but interact and cooperate in complex ecosystems, a phenomenon which synthetic biological systems have yet to fully harness. Technologies that enable the engineering of synthetic ecosystems, or co-cultures, are crucial not only for the study of these natural systems but also for the advancement of synthetic biology.  Developments that enable this foundational leap  in how we engineer biology will allow the creation of synthetic populations that grow together and work together, unlocking the full potential of multicellular engineering in synthetic biology. From creating antibiotic-free human therapeutics and chemical-free biofertilizer based on microbiome engineering, to reprogrammable and dynamic biomaterials, engineering cooperation into synthetic ecosystems and co-cultures has the potential to change how we use biology forever.</p>
 
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<specialh3> Applications of Co-cultures</specialh3>
 
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<button class="btn-link" data-balloon-length="large" data-balloon="One of our favorite application was the idea of modelling financial systems using microbial communities. We came across the following papers. We researched and determined that natural systems’ adaptation processes might reveal methods for streamlining bank interconnections and maintaining diversity in order to protect against global financial crisis. We thought that we could simulate “big bank” and “small bank” populations thanks to our G.E.A.R. population control system. This artificial consortia could then be used to study how the damage on one population (or node) is propagated through the system, and therefore study how the system’s robustness varies as a function of diversity." data-balloon-pos="up">
 
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<button class="btn-link" data-balloon-length="large" data-balloon=" Our body is inhabited by about 100 trillions micro-organisms(2) , they form what is called our human microbiome and it has been shown that they have huge implications on our health. For example Clostridium difficile is responsible for the death of approximately 29,000 death in the US per year(3)  C. difficile is an opportunistic bacteria that colonise your gut microbiome when it is in dysbiosis. Hence we thought that we could use our system to monitor and modulate the proportion of microbiome species and determine whether they are at the healthy ratio or not. If not our system would produce a response to restore healthy ratio through drug production, or growth regulation, etc." data-balloon-pos="up"> 
 
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<button class="btn-link" data-balloon-length="large" data-balloon=" This idea emerged from one of our visit to the Royal College of Art final project presentations. We wanted to design a novel material with exciting properties, so we imagined a novel material which would be light-moldable. The material would be generated by two populations of positive and negative phototaxis cells. These two populations would produce an adhesive protein, which would allow the cells to aggregate. By stipulating the ratio of the negative and positive phototaxis cells we could control the thickness of the material. The different properties of the populations could be exploited to create customisable household items." data-balloon-pos="up">
 
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  <button class="btn-link" data-balloon-length="large" data-balloon=" After reading the Weber paper Synthetic ecosystems based on airborne inter- and intrakingdom communication(1) we thought about using our circuit to create an artificial healthy microbiome on spacecrafts. This would be possible as the papers shows that microbes can communicate through an airborne system. The symbiotic microbes will prevent pathogenic bacteria from overtaking the spacecraft. And therefore our system would keep the astronauts and space travellers happy and healthy on their flights to outer space." data-balloon-pos="up"> 
 
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  <button class="btn-link" data-balloon-length="large" data-balloon=" We imagined a technology which would provide a more sustainable way of consuming/producing probiotics. The user would buy a vial of pre-mixed cells equipped with our Genetically Engineered Artificial Ratio (G.E.A.R.) system. The user would then grow the purchased cells in his or her mini bioreactor, just like growing herbs and spices in your backyard. The G.E.A.R. system would control the cells ratio, allowing the production of probiotics of a desired composition." data-balloon-pos="up">
 
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  <button class="btn-link" data-balloon-length="large" data-balloon="Natural materials, like skin, have intricate material and structural properties which are hard to mimic in artificial materials. We could use microbes as biological 3D printers that can manufacture materials that these unique properties. Using our microbial consortia, we could 3D print composite materials at a microscopic level. Different material properties can be controlled by different ratios of the consortia population." data-balloon-pos="up">
 
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  <button class="btn-link" data-balloon-length="large" data-balloon="Microbial fuel cells can treat wastewater and simultaneously generate energy.  We can use consortia of Geobacter sulfurreducens and E. coli for a more efficient microbial fuel cell.  G. sulfurreducens work best in anaerobic conditions.  E. coli scavenge oxygen which is toxic to G. sulfurreducens enabling MFC operation in aerobic conditions.  However, production of succinate by E. coli reduced the efficiency of the fuel cell.  We can control the ratio of E. coli and G. sulfurreducens to optimise oxygen scavenging and maximising energy production while minimising succinate production" data-balloon-pos="up">
 
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<p>To determine why more labs aren’t using co-cultures, we visited Imperial College London's Centre for Synthetic Biology. We found that many researchers find it too difficult to determine conditions under which multiple cell types would survive. Different cell types grow best at different conditions and there are few established protocols that tell you exactly how you should grow them together. If conditions are not carefully balanced, one cell type tends to out-compete the other. Currently, this problem is addressed by various population control methods, such as auxotrophic cross-feeding and toxin-antitoxin systems. However, these techniques are neither robust, nor portable across different organisms, nor  do they allow precise ratiometric control of the different populations of cells.
 
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<p>We have set out to engineer a genetic circuit that allows ratiometric control of populations in a co-culture to allow future synthetic biologists to realise the full potential of synthetic ecosystems. <br><br>
 
  
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Our genetic circuit employs three modules. The first is the communication module, which utilises two orthogonal quorum sensing systems to allow our <i>E. coli</i> populations to detect their own population density, as well as that of the other population. <br><br>
 
  
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In order to allow for different quorum sensing systems to be utilised in the circuit, we chose to work with four possible systems: Las, Rhl, Lux, and Cin. The Las, Rhl, and Lux transcriptional activators, LasR, RhlR, and LuxR, are fairly well characterised in the Biobrick registry. However, the Cin transcriptional activator is not. <br><br>As a result, crosstalk characterisation for LasR, RhlR, and LuxR did not include Cin 3O-C14 AHL. Therefore, to improve the characterisation of these parts, we performed cross talk experiments with Cin 3O-C14 AHL. Additionally, while a part for CinR does exist in the registry, it has an LVA tag and is uncharacterised. We made a new part for CinR without an LVA tag, and are working to characterise it, and perform crosstalk experiments for CinR with the AHLs associated with Las, Rhl, and Lux. <br><br></p>
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<p>Our second module, the comparator module, links quorum sensing signals to RNA logic so that the bacteria can compare their own population to the population of the other cell line.<br><br>
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The final module is a growth regulation module which allows our cell lines to respond to the signal relayed by the comparator module. If its population is too large, a growth inhibiting protein is expressed, allowing the population ratios to balance once again. <br><br></p>
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In addition to the circuit, we have produced a software tool, A.L.I.C.E, which helps scientists design their own co-culture experiments. We have worked with other iGEM teams to generate the preliminary data for A.L.I.C.E. Our project aims to provide a framework to advance the use of co-cultures in synthetic biology and in research of microbial consortia.
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                                <h1 class="fh5co-intro-lead animate-1 to-animate">Team AshesiGhana</h1>
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                                <h2 class="fh5co-intro-sub animate-2 to-animate">Introducing the Greatest Gold Miners in the World....Microbes</h2>
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                        <h2 class="fh5co-section-heading to-animate">Project Description<span class="fh5co-border"></span></h2>
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                                <p>For the 2017 IGEM, Team AshesiGhana is working on a bio-mining project. The objective of our project is to develop an organism capable of liberating gold from the refractory ore. We aim to provide not only an alternative, non-toxic approach for small scale mining, but also deliver an easy and fast bio-detection and quantification method of ore in the mining industry of our country. The availability of such a biosensor will allow routine monitoring of the ore before a mining endeavor is undertaken thus preventing the destruction of the environment. The project will engineer the typical environmental organism Acidithiobacillus ferroxidans with a FRET probe.</p>
 +
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                                <p>This new organism will primarily be capable of sensing and quantifying the amount of gold in the ore. This will be accomplished by using a two-part probe, a donor part which is made up of gold binding protein (golB) attached to a green fluorescent protein (nowGFP). The second part of the acceptor is also made up of a binding protein and a red fluorescent protein (mRuby2). In the presence of a high amount of free gold, the two parts would be in close proximity and energy transfer can take place and the red protein would be excited giving off a fluorescent signal. Using calibration experiments, we can relate the amount of fluorescence to the amount of gold present,liberated by the organism from the ore.</p>
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                                <p>As Acidithiobacillus ferroxidans is a difficult organism to grow in large quantities, we will engineer a strain of E coli to produce two of the main oxidizing enzymes for iron and sulphite which will liberate the gold from the ore. The organism will also be engineered with protective enzymes against the low pH, which is one of the bi-products of the gold liberation reaction, and metal toxicity. The same FRET biosensor part can be added to the E coli strain for the quantification of gold. This new organism can easily be grown in large batches and so can be used to extract gold from refractory ore without the need of any toxic treatment thus providing a safe alternative for small scale mining. </p>
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                                <p>Sugio, T., Taha, T., & Takeuchi, F. (2009). Ferrous Iron Production Mediated by Tetrathionate Hydrolase in Tetrathionate-, Sulfur-, and Iron-GrownAcidithiobacillus ferrooxidansATCC 23270 Cells. Bioscience, Biotechnology, And Biochemistry, 73(6), 1381-1386. http://dx.doi.org/10.1271/bbb.90036</p><p>
 +
Zeng, J., Jiang, H., Liu, Y., Liu, J., & Qiu, G. (2007). Expression, purification and characterization of a high potential iron–sulfur protein from Acidithiobacillus ferrooxidans. Biotechnology Letters, 30(5), 905-910. http://dx.doi.org/10.1007/s10529-007-9612-2
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<p>George Abraham, B., Sarkisyan, K., Mishin, A., Santala, V., Tkachenko, N., & Karp, M. (2015). Fluorescent Protein Based FRET Pairs with Improved Dynamic Range for Fluorescence Lifetime Measurements. PLOS ONE, 10(8). http://dx.doi.org/10.1371/journal.pone.0134436</p><p>
 +
Held, P. (2005). White Paper: An Introduction to Fluorescence Resonance Energy Transfer (FRET) Technology and its Application in Bioscience. Biotek.com. Retrieved 10 July 2017, from https://www.biotek.com/resources/white-papers/an-introduction-to-fluorescence-resonance-energy-transfer-fret-technology-and-its-application-in-bioscience/</p><p>
 +
Lam, A., St-Pierre, F., Gong, Y., Marshall, J., Cranfill, P., & Baird, M. et al. (2012). Improving FRET dynamic range with bright green and red fluorescent proteins. Nature Methods, 9(10), 1005-1012. http://dx.doi.org/10.1038/nmeth.2171</p><p>
 +
Lavdas, A. You May Not Know Theodor Förster but You Know His Work: FRET - Bitesize Bio. Bitesize Bio. Retrieved 16 July 2017, from http://bitesizebio.com/23012/you-may-not-know-theodor-forster-but-you-know-his-work-fret/</p>
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Latest revision as of 14:11, 1 November 2017

Ashesi University
College

Project Description

For the 2017 IGEM, Team AshesiGhana is working on a bio-mining project. The objective of our project is to develop an organism capable of liberating gold from the refractory ore. We aim to provide not only an alternative, non-toxic approach for small scale mining, but also deliver an easy and fast bio-detection and quantification method of ore in the mining industry of our country. The availability of such a biosensor will allow routine monitoring of the ore before a mining endeavor is undertaken thus preventing the destruction of the environment. The project will engineer the typical environmental organism Acidithiobacillus ferroxidans with a FRET probe.

This new organism will primarily be capable of sensing and quantifying the amount of gold in the ore. This will be accomplished by using a two-part probe, a donor part which is made up of gold binding protein (golB) attached to a green fluorescent protein (nowGFP). The second part of the acceptor is also made up of a binding protein and a red fluorescent protein (mRuby2). In the presence of a high amount of free gold, the two parts would be in close proximity and energy transfer can take place and the red protein would be excited giving off a fluorescent signal. Using calibration experiments, we can relate the amount of fluorescence to the amount of gold present,liberated by the organism from the ore.

As Acidithiobacillus ferroxidans is a difficult organism to grow in large quantities, we will engineer a strain of E coli to produce two of the main oxidizing enzymes for iron and sulphite which will liberate the gold from the ore. The organism will also be engineered with protective enzymes against the low pH, which is one of the bi-products of the gold liberation reaction, and metal toxicity. The same FRET biosensor part can be added to the E coli strain for the quantification of gold. This new organism can easily be grown in large batches and so can be used to extract gold from refractory ore without the need of any toxic treatment thus providing a safe alternative for small scale mining.

Sugio, T., Taha, T., & Takeuchi, F. (2009). Ferrous Iron Production Mediated by Tetrathionate Hydrolase in Tetrathionate-, Sulfur-, and Iron-GrownAcidithiobacillus ferrooxidansATCC 23270 Cells. Bioscience, Biotechnology, And Biochemistry, 73(6), 1381-1386. http://dx.doi.org/10.1271/bbb.90036

Zeng, J., Jiang, H., Liu, Y., Liu, J., & Qiu, G. (2007). Expression, purification and characterization of a high potential iron–sulfur protein from Acidithiobacillus ferrooxidans. Biotechnology Letters, 30(5), 905-910. http://dx.doi.org/10.1007/s10529-007-9612-2

George Abraham, B., Sarkisyan, K., Mishin, A., Santala, V., Tkachenko, N., & Karp, M. (2015). Fluorescent Protein Based FRET Pairs with Improved Dynamic Range for Fluorescence Lifetime Measurements. PLOS ONE, 10(8). http://dx.doi.org/10.1371/journal.pone.0134436

Held, P. (2005). White Paper: An Introduction to Fluorescence Resonance Energy Transfer (FRET) Technology and its Application in Bioscience. Biotek.com. Retrieved 10 July 2017, from https://www.biotek.com/resources/white-papers/an-introduction-to-fluorescence-resonance-energy-transfer-fret-technology-and-its-application-in-bioscience/

Lam, A., St-Pierre, F., Gong, Y., Marshall, J., Cranfill, P., & Baird, M. et al. (2012). Improving FRET dynamic range with bright green and red fluorescent proteins. Nature Methods, 9(10), 1005-1012. http://dx.doi.org/10.1038/nmeth.2171

Lavdas, A. You May Not Know Theodor Förster but You Know His Work: FRET - Bitesize Bio. Bitesize Bio. Retrieved 16 July 2017, from http://bitesizebio.com/23012/you-may-not-know-theodor-forster-but-you-know-his-work-fret/