The need for an innovative bioreactor design was born in conjunction with the concept of reducing perchlorate to oxygen on Mars. The genetically modified E.coli require an environment that would both optimise perchlorate reduction and protect them from the extremes of the Martian environment. Being located in the isolated, hostile environment of the Martian surface the bioreactor design had to be low maintenance, energy efficient and most of all: largely self sufficient.

Below, you will find a timeline of design steps that were taken in order to reach our final bioreactor.

Design 1

Initially we had two bioreactors, one for removing perchlorate from water on Earth and another for producing oxygen on Mars. They were to be identical apart from an attachment for Mars which would allow the input of regolith rather than water.
We wanted to improve upon the bioreactor designed by previous iGEM team, Leiden 2016, so this was where our bioreactor started. It was very simple with mobilised bacteria being mixed with perchlorate in a single chamber.

Design 2

We decided to try and have the two reactions (perchlorate to chlorite, and chlorite to chloride and oxygen) taking place in separate areas of the bioreactor. Because the operons involved the whole process is so long, we were hoping that this would allow us to insert less DNA into E. coli, making expression more likely.
However, this idea didn’t last long enough for us to come up with a design, as we discovered that chlorite is toxic to E. coli *source* and so it would all have to take place in one chamber and with one operon.

Design 3

After reading Biotechnological Applications of Microbial (Per)chlorate Reduction(Ouwei Wang and John D. Coates, 2017) we considered the idea of growing E. coli on a cathode in order to provide it with more electrons and speed up the reaction by up to 12%. The paper proposed a bioreactor in which DPRB (disparate perchlorate reducing bacteria) grew on a cathode, supplying ions that seemed to be a limiting factor in the reaction. Hydrogens would pass from the anode to the cathode through a cation selective membrane, aiding the reaction in the diagram (left). This would speed up the first half of the process, converting perchlorate to chlorite, as this is the slower half. We would simply replace the DPRB with modified E. coli.

Design 4

A meeting with Dr Mike Allen from the Plymouth Marine Laboratory introduced us to the concept of Swirl Flow Bioreactors (SFB), which would allow us to mix the perchlorate with the E. coli as well as separating the oxygen, all in one chamber. We took the design of a typical swirl flow and adapted it to fit our needs of having immobilized E. coli and a cathode.

Design 5

In a second meeting with Dr Mike Allen, he also told us valuable information about how our bioreactor would be received within global markets. He informed us of his experience with selling and developing his own bioreactor, and from that, he advised that we focus more on the Mars travel side of the the bioreactor as there wasn’t much room in the market for a bioreactor on Earth.
During this meeting, we decided to remove the cathode from our design as it would provide the opportunity for unexpected chemistry, as well as sparks which would react badly with the oxygen we are producing. This allowed our bioreactor design to simplify significantly and we started making quick progress.

Design 6

Our first SFB without cathodes involved four chambers, one to mix the regolith with water to extract the perchlorate, and three to consecutively reduce the perchlorate into oxygen until none is left. The vortex created in the SFB would sort the contents according to density, forcing the oxygen into the centre of the tube. A manifold at the end of the tunnel containing a small central tube would carry the oxygen out of the bioreactor while everything else would be carried into the next chamber via a larger tube surrounding the oxygen outlet.

Design 7

With a solid base, we began to tweak the design based on the needs of stakeholders *add specific stakeholders*.

3D printable to be made on Mars, and anything that couldn’t had to be light and easy to carry on a rocket.
Low power consumption in order not to drain the resources of the rest of the biodome.
Shielded from ionising radiation which would affect the E. coli. This shielding also needs to be light enough to take to Mars.
There must be a way to kill the bacteria in the bioreactor in the case of unwanted mutations or other unforeseen events.

Design 8

Our first remodel of the SFB involved cutting down the number of chambers requiring propellers in order to reduce the power consumption and resources needed.

Ouwei Wang, John D. Coates (2010) Biotechnological Applications of Microbial (Per)chlorate Reduction (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748585/)
Françoise Bichai, Benoit Barbeau (2006) Assessing the Disinfecting power of Chlorite in Drinking Water (https://www.researchgate.net/publication/255640211_Assessing_the_Disinfecting_Power_of_Chlorite_in_Drinking_Water)

Team:Exeter/Bioreactor Design