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From 2010.igem.org

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In this ever more energy-dependent world, where fossil fuel resources become scarce and raise environmental issues, the search for green energy sources is a growing concern in both civil and scientific communities. In this context, hydrogen turns out to be an interesting alternative. Indeed, hydrogen fuel cells do not affect the carbon footprint since their only side effect is a production of water. Although the hydrogen is the most abundant element on the planet, it is quite difficult to produce it. Current hydrogen production relies mostly on chemical processes, such as petroleum cracking or water electrolysis. Those processes require a lot of energy: heat for the chemical way or electricity for water electrolysis.
In order to develop greener and more energy-efficient processes, the use of micro-organisms as biocatalysts for hydrogen production has been studied for many years. While no industrial application has yet been achieved, nowadays the scientific and technological advances allow further developments and opportunities in this field.
A major problem of this approach lays in the fact that the use of dark fermentation to produce hydrogen attains very low yields, compared to other fermentative biofuel synthesis, e.g. methane or ethanol. We propose to design a genetically engineered E. Coli, Hydrocoli, with an improved natural hydrogen production pathway, using the organic compounds found in wastewaters as substrate.
In addition, we planned to implement various features to enable the strain to perform other tasks related to wastewater treatment, such as signaling metallic contamination, eliminating nitrogen compounds, or hindering hydrogen consumption by methanogenic bacteria. We will also set up a planned death system in order to prevent its proliferation outside the wastewater treatment plant.
Turning to wastewaters as a substrate is convenient in several ways: wastewater treatment plants exist almost everywhere and the E. Coli bacteria is readily available in wastewaters. Moreover this approach may result into a reduction of sludge which otherwise would need to be treated. Besides that, the use of genetically modified organism in waste water treatment has never been achieved and could potentially increase the efficiency of the process. It could also show people that genetically modified organisms are a safe and easy way to improve our environment. The ethic and economic issues of our project and of synthetic biology in general will be also discussed.
In order to tackle this ambitious project in a relatively short time, we have decided to divide the approach in several modules:

• Hydrogen production
• Bacteria planned death system
• Detection of metallic contamination

For each module we proceeded to a computerized model in order to analyze the reaction kinetics prior to the manipulations.

The hydrogen production module leaded to another module which we called homologous recombination. In order to increase the production rate of hydrogen in our Hydrocoli, we will have to delete several genes. We did those deletions, using a new method based on the λ phage Red recombinase developed by Datsenko and Wanner. We planned to construct all the tools needed in order to make any deletion of any genes using this method.