Team:Charlottesville RS/Project

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''' Junk Food '''
''' Junk Food '''
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In Albemarle County, most of the waste water goes and is processed through the Moore's Creek Wastewater Treatment Plant (WWTP), which is managed by the Rivanna Water & Sewer Authority. Each year, the plant purchases 250,000 dollars worth of glycerin, which is then used as “food” for bacteria which degrade the other organic matter in the water.
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In Albemarle county, most of the waste water goes and is processed through the Moores Creek Wastewater Treatment Plant (WWTP), which is managed by the Rivanna Water & Sewer Authority. Each year, the plant purchases 250,000 dollars worth of glycerin, which is then used as “food” for bacteria which degrade the other organic matter in the water.  
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The UVa iGEM team from 2008 created a part that, when added, enables E Coli to produce polyhydroxybutyrate, a biodegradable, bio-derived plastic. Our idea is to make a type of E Coli that produces this plastic, with a reporter which fluoresces when the bacteria is successfully producing.  
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The UVA iGEM team from 2008 created a part that, when added, enables E Coli to produce polyhydroxybutyrate, a biodegradable, bio-derived plastic. Our idea is to make a type of E Coli that produces this plastic, with a reporter which fluoresces when the bacteria is successfully producing.
The plant could then use this bacteria to create polyhydroxybutyrate, filter out the E Coli, and then use the plastic as an alternative food source for their bacteria, saving them 250,000 dollars per year, as well as giving them a renewable energy source for their plant.
The plant could then use this bacteria to create polyhydroxybutyrate, filter out the E Coli, and then use the plastic as an alternative food source for their bacteria, saving them 250,000 dollars per year, as well as giving them a renewable energy source for their plant.
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== Importance of Nitrogen==
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Nitrate (NO3) is a compound found in most water. It is colorless and odorless, so it can only be detected with testing. Nitrate itself isn’t bad in water, but it causes eutrophic effects if there is too much, which causes a chain reaction of harmful effects (explained further in the next section). Every living organism (as well as non-living organisms such as viruses) needs nitrogen to live; it helps to form the structure of proteins and DNA and many other molecules in cells that are essential for plants and animals to live. The majority of nitrogen in the world is held in the atmosphere, but not in its usable form. For nitrogen to be usable by organisms, it must be in the form of either ammonium or nitrate, which it is converted to in either abiotic or biotic fixation. Animals (like ourselves) get nitrogen from eating plants and other animals. Plants get nitrogen not by consuming other organisms but by absorbing it through the soil in its fixated forms.
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| [[Image:HD_Sensing.png|left|thumb|300px| Fig. 1: A killer cell senses the specific signal of autoinducer-2 secreted by prey cells and chases them with chemotaxis]] || [[Image:Fig_0-System.png|left|thumb|300px|Fig. 2: Once the preys are caught the killer cells are induced by a second specific signal, autoinducer-1, to activate a killer mechanism.]]
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[[https://2008.igem.org/Team:Heidelberg/Project back]]
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You are very interested in this exciting project, but did not understand all details? Well then come on with me on my guiding tour [[Team:Heidelberg/Human_Practice/Phips_the_Phage/General_Backround|'''... follow phips to step 3''']] [[Image:Phips_3.png|middle|50px]]  [ [[Team:Heidelberg/Team| ''... back to step 2'']] ]
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<div> ''Deutsche Übersetzung:''
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'''Ecolizenz zum Töten: Eine ''E. coli''-Konstruktion zum gezielten Angriff auf pathogene Mikroorganismen'''
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Pathogene Mikroorganismen stellen eine große Herausforderung sowohl für die Medizin als auch für die Industrie dar. Gemeinschaften aus Mikroorganismen, die in der Fachsprache als Biofilme bezeichnet werden, haben sich als äußerst resistent gegenüber konventionellen Therapien erwiesen [1]. Um deren Wachstum zu bekämpfen bedarf es daher alternativen Ansätzen. Die Entstehung von Biofilmen, genauso wie die der Virulenz, die Produktion von Antibiotika oder die Bildung von Sporen werden durch Kommunikations-Schaltkreise reguliert, die auf kleinen Signalmolekülen - den sogennanten Autoinduzierern (AI) - basieren. Unser Ziel ist es, diese Kommunikationsmechanismen in der Weise zu integrieren, dass synthetische Bakterien potenziell gefährliche, AI-ausschüttende Spezies aufspüren und töten können. Mit ''E. coli'' als Modellorganismus beabsichtigen wir zwei separate Stämme - einen Killer- und einen Beutestamm - zu entwerfen. Das Projekt gliedert sich in zwei ergänzende Module: Die Signalerkennung umfasst die Modifizierung des Chemotaxis-Systems von E. coli [2], um Killer-Zellen zu den Beute-Zellen schwimmen zu lassen. Dafür ist es notwendig, dass ein Erkennungsmodul einen Beute-spezifischen Stimulus an die Signalkaskade für Chemotaxis weiterleitet, so dass die Killer-Zellen entgegen des Gradienten des Stimulus schwimmen. Das gerichtete Schwimmen kommt nur zum Erliegen, wenn sich die Killer-Zellen in der unmittelbaren Umgebung der Beute-Zellen befinden - dort, wo die Stimuluskonzentration maximal ist. Das Killer-Modul gewährt, dass - einmal in der Nähe der Beute angekommen - ein bakterizider Mechanismus aktiviert wird. Computermodelle werden ergänzend zeigen, wie sich die beiden Modelle im Zusammenspiel verhalten, und wie die Effizienz des Gesamtsystems in seiner räumlichen Umgebung einzuschätzen ist. In Zukunft - sicherlich außerhalb der Reichweite unseres Projekts - könnte die Spezifität auf relevante Pathogene, genauso wie auf andere Organismen, Gewebe oder sogar Krebszellen erweitert werden. </div>
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[[https://2008.igem.org/Team:Heidelberg/Project back]]
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== Project Realization==
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Have you ever wondered, why numbers of iGEM team members are limited to a maximum of twelve students? Apart from financial resources, this limit is only natural: efficient work with larger team sizes requires a subtle organization and project management, in particular when the team is yet green! On top, team formation and project work have to be conducted within a short time interval and under liabilities to the academic curriculum - both on the student and the advisor sides. The social abilities for reliable team formation, in particular communication, team integration, and consequence to work towards common team goals, must therefore be substantial and dedicated to compensate these circumstances. When you visit our portraits, however, you will realize that we are not really green and that each of us has an active past often intricately connected with scientific work. In our project description we provide a sincere answer to the apparent question, if 16 "greenhorns" can push forward in only four months to contribute research on an international level!
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In order to streamline our horse-power, we divide the project into four columns, which are simultaneously developed by us and can merge in the end with a synergistic effect to a complex population system (modeling inclusive). At the same time the global progress is robust towards each column, consequently a delay or failure in one column will not lead to a breakdown of the overall project. In accordance with current scientific standards demanded in systems and synthetic biology, we pursue an interdisciplinary approach combining mathematics, experimental, and quantitative biology [3].
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The first column ([[2008.igem.org:Team_Heidelberg/Project/Sensing|Project - Sensing]]) comprises the design and development of a sensing module. Chemotaxis is a very effective mechanism of bacteria to move towards nutrients. Using the natural signals for bacterial quorum communication, we re-design a chemotactic receptor for detection of the quorum signal (AI-2) and its transmission to the chemotaxis signaling network. A prey strain, which is to be found via the sensing mechanism, will be engineered to secrete two prey specific quorum signals (AI-1 and AI-2). We dedicate two columns to the development of a killing mechanism to secure a higher implementation confidence: In one column, the second ([[2008.igem.org:Team_Heidelberg/Project/Killin_I|Project - Killing I/Phages]]), a killing mechanism is developed based on bacteriophages, which are conjugated into prey cells, once the killers are in their vicinity. In prey cells, however, the phage becomes lytic due to the missing regulation of the cI transcription factor; the phage is thereby released and infects other prey cells in a domino effect. The killing module in the third column ([[2008.igem.org:Team_Heidelberg/Project/Killing_II|Project - Killing II/Colicins]]) can be considered as a mechanism for recognition of molecular exudation patterns. Once killer cells are in the vicinity of preys, they detect the specific signal molecule AI-1, which leads to the production and release of colicins. The bactericide substance, to which the producing host is resistant, will then eliminate the prone prey. In a fourth column ([[2008.igem.org:Team_Heidelberg/Modeling|Modeling]]) we model the system behavior of these two bacterial strains - the killer and the prey - in a 2D landscape. One approach is based on partial differential equations and simulates chemotaxis and the colicin killing mechanism; the second approach, based on delay differential equations, analyzes the killing dynamics of conjugated bacteriophages. [[2008.igem.org:Team_Heidelberg/Project/Visualization|Visualization]] - the architrave - with microscopy is finally employed to quantify bacterial density landscapes in space and time - allowing a phenotypic connection and comparison with our modeling work. An experimental engagement would these days of course not be appreciable without the active responsibility towards the sake of the community under ethical considerations. We therefore actively engage in [https://2008.igem.org/Team:Heidelberg/Human_Practice/Project_Overview  human practice] ... and the pupils are enthusiastic!
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In Figure 3 you can see the essence of our project: The sensing column could be advanced significantly, though efficient chemotaxis towards AI-2 could not be established. The phages are conjugated efficiently, though a killing of the prey could not yet be experimentally implemented. The colicin column and the modeling, however, worked out in all respects, and the human practice work has found enthusiastic resonance. With seven new standardized Biobrick parts, all of them have been shown to work, four characterized new parts and one characterized old part, we will run a high pace for the hottest awards at the championship jamboree in one week!
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[[Image:HD_overview2_klein.png|left|650px|thumb|Fig. 3: Overview on project advancements. The colicin, the modeling, and the human practice columns are immaculate.]]
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[[https://2008.igem.org/Team:Heidelberg/Project back]]
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== Project Perspectives==
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This project is supposed to be a wet model for a mechanism of specific target finding and elimination, in particular of cancer cells. There is clear evidence that bacteria accumulate in tumor areas [4, 5]. One reason might be the immunosuppressive effect of the tumor. The possibility to discriminate cancer patients from breath samples [6, 7, 8] gives rise to the idea to use bacterial receptors for the recognition of specific molecular exudation patterns. The effect of bacterial accumulation might in this way be enhanced by the engineering of a pattern sensitive chemotaxis mechanism. Furthermore bacteria could be induced very specifically in the near environment of the tumor to metabolize a pre-curser into a tumor suppressive molecule. Both mechanisms, increased accumulation at tumor sites and tumor located drug metabolism, would lead to a potentiated selectivity for the elimination of cancer cells. If the toxin is then also cancer cell specific - as could be shown by Pastan ''et al.'' [9] - then three strong effects would fulminate into ... of course, all this is still music of the future - but we are sure: it will be the future!
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[[https://2008.igem.org/Team:Heidelberg/Project back]]
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== Literature ==
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== Why we Don't Want Nitrates in our Water ==
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Because everything needs nitrogen to live, we obviously want to have some in our water. The problem arises when there is just too much - because that causes eutrophication. This is a huge problem right now in the Chesapeake Bay, and since we are part of the watershed that flows into the Bay, we have a responsibility to keep it clean. The Moores Creek WWTP and other treatment plants in the area are now obligated under the law to filter enough nitrate out of the sewage water because of the condition of the Chesapeake Bay, but soon most treatment plants in the country will probably have to follow in their footsteps. The filtered supernatant of the sewage water that goes through Moores Creek will drain into the Bay, so it needs to be clean. If there is too much nitrate in this supernatant, it will further contribute to the decline of the Bay - when there is too much of a macronutrient like nitrate, everything grows too much. For our purposes, this is mostly bad in reference to algae. The huge algae blooms that eutrophication causes not only block out sunlight to the organisms living under the water, but they eventually lead to anoxic zones in the water (meaning that there is no oxygen there). This happens when the algae dies and floats to the bottom of the water, and is then eaten by bacteria. Because there is so much algae to eat, the bacteria community flourishes, and they end up using all the oxygen in the water. Animals like fish and any others that preform cellular respiration cannot live under these circumstances because they need oxygen - this often leads to large groups of organisms dying. Right now, eutrophication is probably the biggest problem for the Chesapeake Bay, and plenty of other bodies of water in the world, It's very important that waste water treatment plants do their job  efficiently and without too much money spent; and using polyhydroxybutyrate instead of glycerin will help to do that.
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|[1] || width=500px |K. Brenner ''et al.'', Engineered bidirectional communication mediates a consensus in a microbial biofilm consortium, ''PNAS'', Vol. 104 (44), pp. 17300-17304 (2007)
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|[2] || G. H. Wadhams and J. P. Armitage, Making sense of it all: bacterial chemotaxis, ''Nat. Rev. Mol. Cel. Bio.'', Vol. 5, pp. 1024-1037 (2004)
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|[3] || N. Wingreen and D. Botstein, Back to future: education for systems-level biologists, ''Nat. Rev. Mol. Cel. Bio.'', Vol. 7(11), pp. 829-832
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|[4] || J. M. Pawelek ''et al.'', Bacteria as tumour-targeting vectors, ''The Lancet Oncology'', Vol. 4, pp. 548-546 (2003)
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|[5] || H. Loessner ''et al.'', Remote control of tumour-targeted ''Salmonella enterica'' serovar Typhimurium by the use of L-arabinose as inducer of bacterial gene expression ''in vivo'', ''Cellular Microbiology'', Vol 9 (6), pp. 1529-1537 (2007)
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|[6] || C. M. Willis ''et al.'', Olfactory detection of human bladder cancer by dogs: proof of principle study, ''Biomedical Journal'', Vol. 329 (2009)
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|[7] || M. Phillips ''et al.'', Detection of Lung Cancer with Volatile Markers in the Breath, ''Chest'', Vol. 123, pp. 2115-2123 (2003)
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|[8] || M. Phillips ''et al.'', Volatile organic compounds in breath as markers of lung cancer: a cross-sectional study, ''The Lancet'', Vol. 353, pp. 1930-1934 (1999)
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|[9] || I. Pastan and D. Fitzgerald, Recombinant Toxins for Cancer Treatment, ''Science'', Vol. , pp. 1173-1177 (1991)
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[[https://2008.igem.org/Team:Heidelberg/Project back]]
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== Polyhydroxybutyrate ==
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Polyhydroxybutyrate (PHB) is a polymer of glucose and a biodegradable plastic. It is a special type of polyester called a polyhydroxyalkanoate. PHB is produced by microorganisms such as Ralstonia eutrophus or Bacillus megaterium in response to conditions of physiological stress, particularly conditions in which nutrients are limited. PHB can also serve as food for other bacteria. 
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[[image:Phips_phage.PNG|middle|200px|Phips the Phage]]
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In waste water treatment, bacteria are required to dispose of waste in the water, but feeding them is very expensive for treatment plans, and for the Rivanna Water Treatment Plant, the closest such facility to our school, it costs the city up to $250,000 a month to simply feed these bacteria. 
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Hi, you are new here and hear about synthetic Biology for the first time? Perfect! I am Phips the Phage and will be your guide to this exciting field of biological research and if you like I will explain the background synthetic biology and gentic engineering as well as of this project to you. Just follow me[[Team:Heidelberg/Human_Practice/Phips_the_Phage/|'''... follow Phips''']]
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Our idea for the project involves using bacteria in order to produce PHB as food for the bacteria that are involved in waste water treatment. Since PHB is only produced when nutrients are limited, it would actually involve feeding the PHB-producing bacteria less than the waste water treatment bacteria would be fed. We believe our idea has the potential to drastically reduce the costs of waste water treatment plants around the globe.

Latest revision as of 15:13, 20 June 2014

Project Idea

Junk Food

In Albemarle County, most of the waste water goes and is processed through the Moore's Creek Wastewater Treatment Plant (WWTP), which is managed by the Rivanna Water & Sewer Authority. Each year, the plant purchases 250,000 dollars worth of glycerin, which is then used as “food” for bacteria which degrade the other organic matter in the water.

The UVA iGEM team from 2008 created a part that, when added, enables E Coli to produce polyhydroxybutyrate, a biodegradable, bio-derived plastic. Our idea is to make a type of E Coli that produces this plastic, with a reporter which fluoresces when the bacteria is successfully producing.

The plant could then use this bacteria to create polyhydroxybutyrate, filter out the E Coli, and then use the plastic as an alternative food source for their bacteria, saving them 250,000 dollars per year, as well as giving them a renewable energy source for their plant.

Importance of Nitrogen

Nitrate (NO3) is a compound found in most water. It is colorless and odorless, so it can only be detected with testing. Nitrate itself isn’t bad in water, but it causes eutrophic effects if there is too much, which causes a chain reaction of harmful effects (explained further in the next section). Every living organism (as well as non-living organisms such as viruses) needs nitrogen to live; it helps to form the structure of proteins and DNA and many other molecules in cells that are essential for plants and animals to live. The majority of nitrogen in the world is held in the atmosphere, but not in its usable form. For nitrogen to be usable by organisms, it must be in the form of either ammonium or nitrate, which it is converted to in either abiotic or biotic fixation. Animals (like ourselves) get nitrogen from eating plants and other animals. Plants get nitrogen not by consuming other organisms but by absorbing it through the soil in its fixated forms.

Why we Don't Want Nitrates in our Water

Because everything needs nitrogen to live, we obviously want to have some in our water. The problem arises when there is just too much - because that causes eutrophication. This is a huge problem right now in the Chesapeake Bay, and since we are part of the watershed that flows into the Bay, we have a responsibility to keep it clean. The Moores Creek WWTP and other treatment plants in the area are now obligated under the law to filter enough nitrate out of the sewage water because of the condition of the Chesapeake Bay, but soon most treatment plants in the country will probably have to follow in their footsteps. The filtered supernatant of the sewage water that goes through Moores Creek will drain into the Bay, so it needs to be clean. If there is too much nitrate in this supernatant, it will further contribute to the decline of the Bay - when there is too much of a macronutrient like nitrate, everything grows too much. For our purposes, this is mostly bad in reference to algae. The huge algae blooms that eutrophication causes not only block out sunlight to the organisms living under the water, but they eventually lead to anoxic zones in the water (meaning that there is no oxygen there). This happens when the algae dies and floats to the bottom of the water, and is then eaten by bacteria. Because there is so much algae to eat, the bacteria community flourishes, and they end up using all the oxygen in the water. Animals like fish and any others that preform cellular respiration cannot live under these circumstances because they need oxygen - this often leads to large groups of organisms dying. Right now, eutrophication is probably the biggest problem for the Chesapeake Bay, and plenty of other bodies of water in the world, It's very important that waste water treatment plants do their job efficiently and without too much money spent; and using polyhydroxybutyrate instead of glycerin will help to do that.

Polyhydroxybutyrate

Polyhydroxybutyrate (PHB) is a polymer of glucose and a biodegradable plastic. It is a special type of polyester called a polyhydroxyalkanoate. PHB is produced by microorganisms such as Ralstonia eutrophus or Bacillus megaterium in response to conditions of physiological stress, particularly conditions in which nutrients are limited. PHB can also serve as food for other bacteria.

In waste water treatment, bacteria are required to dispose of waste in the water, but feeding them is very expensive for treatment plans, and for the Rivanna Water Treatment Plant, the closest such facility to our school, it costs the city up to $250,000 a month to simply feed these bacteria.

Our idea for the project involves using bacteria in order to produce PHB as food for the bacteria that are involved in waste water treatment. Since PHB is only produced when nutrients are limited, it would actually involve feeding the PHB-producing bacteria less than the waste water treatment bacteria would be fed. We believe our idea has the potential to drastically reduce the costs of waste water treatment plants around the globe.