iGEM 2014: UC Santa Cruz BioE Project

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Project Summary of the iGEM 2014 team: UC Santa Cruz BioE. A microbial fuel cell (MFC) uses bacteria to break down organic compounds found in waste water and generate an electric current. My colleagues and I intend to genetically engineering the bacteria Shewanella oneidensis in ways that will make it will participate in energy production more efficiently. We will be on of the two teams competing in the international genetically engineered machines (iGEM) competition to represent the UC Santa Cruz Banana Slugs on an international stage! Please consider helping us fund this project and/or giving it exposure. http://tinyurl.com/ne9cqbb

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iGEM 2014: UC-Santa Cruz-BioE Sai Edara - Biomolecular Eng Aaron Maloney - Bioelectronics Eng Marshall Porter - Biomolecular Eng David Dillon - Biomolecular Eng Christian Pettet - Biomolecular Eng Arjun Sandhu - Biomolecular Eng Ansley Tanoto Alex Ng Undergraduate Synthetic Biology Competition put on by MIT. What is iGEM? Undergraduate Synthetic Biology Competition put on by MIT. Students from universities around the world work with a kit of biological parts, and parts they design, to build biological systems What is iGEM? Undergraduate Synthetic Biology Competition put on by MIT. Students from universities around the world work with a kit of biological parts, and parts they design, to build biological systems What is iGEM? UCSCs debut year at the jamboree held in Boston, MA will be "...the largest single event in the history of the iGEM (International Genetically Engineered Machines) competition and synthetic biology " This year we are working with the bacteria Shewanella oneidensis to increase efficiency of a microbial fuel cell, a technology capable of turning waste water treatment into a power generating process. iGEM at UC Santa Cruz Waste-water treatment Treatment of waste water can be divided into three main steps 1. Heavy and light materials are removed by separation in a holding tank 2. Microorganisms are used to break down organic matter 3. Water is disinfected to be reintroduced to environment http://en.wikipedia. The bacteria Shewanella oneidensis Can live in both environments with or without oxygen Can reduce poisonous heavy metal Has electrogenic properties allowing it to generate electricity in a Microbial Fuel Cell (MFC). http://www.newscientist.com/article/dn9526- bacteria-made-to-sprout-conducting- nanowires.html#.U9gp_LEzCM0 What is an MFC? Microbes Break down Carbohydrates Transfers electrons to anode, which then flow to the cathode Protons pass through permeable membrane Protons and electrons react with oxygen to make clean water Can be implemented into secondary treatment of waste- water to allow for power generation[5] http://www.sciencebuddies. org/Files/3665/5/Energy_img033.jpg Physical Design A lot of previous research has looked at structural design Two main points Large surface area on electrode Close distance between electrodes 3D Model for casing Designed by iGEM 2013 Team Bielefeld (Germany). Files converted into 3D printer files and printed. http://2013.igem.org/Team:Bielefeld- Germany/Project/MFC Our Project We believe the bacteria which drive the power generation of an MFC can be genetically engineered to create more power Design MFC with increased efficiency by Altering metabolism of our electrogenic bacteria Modifying growth pattern of biofilm formation Two pronged approach, each with potential to improve efficiency alone Energy Balance and Coulombic Efficiency The process of metabolism and electron transfer is complex. The cell itself uses up some of the energy in other processes One such process is metabolite generation, which reduces coulombic efficiency.[1] We plan to redirect metabolism toward a pathway capable of harvesting the lost energy When Shewanella is grown without oxygen, it generates the metabolite acetate from acetyl- coa Acetate generation [3] gate keeper to the TCA cycle Converts acetyl-CoA and Oxaloacetate to Citrate Diverts Acetyl-CoA from being converted to Acetate (metabolite) Citrate Synthase (GltA) Under anaerobic conditions Shewanella is capable of using the oxidative branch of TCA, which will produce more energy lost by metabolite generation Use of oxidative branch is reliant on Citrate Synthase activity Oxidative branch Oxidative branch of TCA Citrate Synthase Under the anaerobic conditions citrate synthase activity reduced by over one half due to downregulation of the gltA gene coding for citrate synthase [3] In our project we will recover this activity using an expression plasmid (gene deletion) [3] Magnitude of electron transfer reliant on surface area of the anode More surface area allows more bacteria to transfer electrons Growth of bacteria in biofilm allows for a dense community to grown in one area Growth of Shewanella in anaerobic conditions leads of down regulation of biofilm production, and biofilm density is lost Biofilm Steps in Biofilm growth: 1 2 3 4 5 http://en.wikipedia.org/wiki/Biofilm Biofilm Biofilm formation in Shewanella is controlled by the gene mxdA, which regulates levels of c-di-GMP Upon deletion of mxdA, biofilm biomass decreases (fig A, mxdA) Biomass also decreases when switching from oxic to anoxic growth (fig B, control) but is retained when a gene similar to mxdA is expressed (fig B, VCA0956)[7] We hope to express mxdA in anoxic conditions [3] to increase biofilm density A B [7] Citations 1. Korneel Rabaey, ed. Bioelectrochemical systems: from extracellular electron transfer to biotechnological application. IWA publishing, 2010. 2. Franks, Ashley E., and Kelly P. Nevin. "Microbial fuel cells, a current review." Energies 3.5 (2010): 899-919. 3. Brutinel ED, Gralnick JA. Anomalies of the anaerobic tricarboxylic acid cycle in Shewanella oneidensis revealed by Tn-seq. Mol Microbiol. 2012 Oct;86(2):273-83. doi: 10.1111/j.1365-2958.2012.08196.x. Epub 2012 Aug 27. PubMed PMID: 22925268. 4. Papagianni M. Recent advances in engineering the central carbon metabolism of industrially important bacteria. Microb Cell Fact. 2012 Apr 30;11:50. doi: 10.1186/1475-2859-11-50. Review. PubMed PMID: 22545791; PubMed Central PMCID: PMC3461431 5. Rabaey K, Verstraete W. Microbial fuel cells: novel biotechnology for energy generation. Trends Biotechnol. 2005 Jun;23 (6):291-8. Review. PubMed PMID: 15922081. 6. Beliaev, Alex S., et al. "Gene and protein expression profiles of Shewanella oneidensis during anaerobic growth with different electron acceptors." Omics: a journal of integrative biology 6.1 (2002): 39-60. 7. Thormann, Kai M., et al. "Control of formation and cellular detachment from Shewanella oneidensis MR-1 biofilms by cyclic di-GMP." Journal of Bacteriology 188.7 (2006): 2681-2691.

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