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

1. 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

2. ● Undergraduate Synthetic Biology Competition put on by MIT.
What is iGEM?

3. ● 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?

4. ● 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?
● UCSC’s 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
"

5. ● 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

6. 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.

7. 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

8. 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

9. 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

10. 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

11. 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

12. ● When Shewanella is grown
without oxygen, it generates the
metabolite acetate from acetyl-
coa
Acetate generation
[3]

13. ● “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)

14. ● 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

15. 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]

16. ● 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

17. 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]

18. 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.