Team:CIDEB-UANL Mexico/project abstract

iGEM CIDEB 2014 team’s project consists on a bacterial bio-filter, in which sodium ions are filtered out of salty water. In order to achieve this objective, the project was divided into 4 modules; each one consisting on a separate construction in order to separately test the effectiveness of the genes used. These modules are named after their function, and the name of the project “E.CARU” was based in them. The Capture module uses a new gen “nhaS” registered by the team, to remove sodium ions from water. The Aroma module is in charge of producing a “Winter green” odor to report if the filter is working. The resistance module allows E. coli to withstand the salinity of the environment in which it is required to work, and finally, the Union module causes the bacteria to join to silica pearls, facilitating their removal from the filter. The project’s modularity was used for testing the genes; in further development, the pieces are planned to be joined in a smaller, possibly single construction.

Water has been always known as a source of life, but nowadays there is not enough usable fresh water available in the world. The global lack of fresh water is an issue that presents a dangerous problem to our future. Only a small portion of our planet's water is actually usable. Ninety-seven percent of the world's water is too salty for consumption or agricultural use. Furthermore, much of the rest is held in ice caps or other unattainable sources. This leaves approximately one percent of the global water as liquid and fresh; ninety-eight percent of which is groundwater (Bouwer, 2002).

For solving this problem different methods have been developed. One of them is desalination, converting sea water (rich in salts) into usable water, but this method is very expensive due to the great use of electrical energy and the extraction process produces dangerous wastes to the environment (Cotruvo, 2010).

For that reason our project is focused on developing a biological machine capable of performing desalination, reducing costs and avoiding dangerous wastes during the process. For making this possible, E. coli needed to capture Na⁺ ions in saline environments to be removed from water after performing its task.

Before E. coli could be able to remove Na⁺ ions from water, it needed to aquire a resistance to adverse conditions, in particular excess salt. This could be possible through a protein called “IrrE”, which makes E. coli resistant to saline environments as well as UV rays and temperature changes.
The protein NhaS (a new part), was used to give E. coli the ability to bind and capture Na+ ions. Also, the optimized version of the reporter “BSMT1”, a protein able to react with salicylic acid and release a Wintergreen odor, was used to know if nhaS was expressed.
The final task that E. coli should perform was to bind its membrane to silica pearls, in order to be able to be removed from the water after sequestering Na⁺ ions. In order to do this, a fusion protein named L2+AIDA was used. L2 gives E. coli the ability to bind silica, and AIDA acts as a tag for making L2 a membrane protein. With this ability E. coli could be removed from water through a bio-filter, made up of silica.
The complete circuit is shown in figure 1. BSMT1 opt acts as a reporter for nhaS which is regulated by UV (to have a control over the NhaS expression), and IrrE with L2+AIDA are continuously produced.

But we realized E. coli could have a genetic overload because the circuit was too big (approximately 5000 bp). Also the time we had to finish it was not enough, as well as most of the proteins we wanted to produce were putative or untested. So for a better understanding and for determine if each piece works we divided the project into four modules: capture, union, aroma and resistance, but the project is the result of their correlation. In fact our E. coli was named E. CARU (each letter by each module).

Escherichia coli Capture Aroma Resistance Union

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