We have engaged with experts from different fields to make sure that our project and its execution can be responsible, safe and sustainable. In this page, we highlight specific interactions from stakeholders, in particular from the water industry, that have directly impacted and shaped different parts of our Phosphostore project. Click here to our silver Human Practice page to see the full list of different experts we engaged with!
1. Used real world phosphate data from Davyhulme Treatment Works to estimate the production cost of our bacteria and used Davyhulme as a real life scenario to estimate the cost of treating wastewater for a year
2. Determined the thermostability of our PPK enzyme in the lab to test the viability of our idea which was generated by the suggestion from one of our stakeholders
3. Created a business plan that is developed by exploring three areas of human practices:
legislation, scale-up and intellectual property
4. Used information and resources given by the Centre for Process Innovation to model a bacterial continuous culture system and estimate the production cost of our bacteria
Exploring the water industry taught us a lot on how great scientific ideas may not be feasible in the real world. From the start of our journey, we thought about the water industry as a potential market for the implementation of our project in real life. However, we simply did not know the current state of the UK water industry and whether or not synthetic biology that incorporates genetically modified organisms would be accepted as an innovative solution, given how controversial it can be.
We talked to seven different experts - five of which were from water companies - to learn and understand the current state of the water industry. The full list of our interactions can be found in our silver Human Practice page. The most important information that we gained from our interactions is that there are no insurmountable barriers to innovation using synthetic biology and that regulations in the water industry are now more flexible in accommodating new innovation. As long as we can prove that our project is cheaper, reliable and safe to use, water companies will be interested. We have summarized our findings in an infographic that can be found here.
Now that we know the current state of the industry, we need to know how our project works in real life and how it can fulfill the necessary criteria to make it appealing to water companies. So we had a site visit to Davyhulme Treatment Works where we were shown the basic process of water treatment (from separation of sludge to water treatment through biological means) through a tour around the site. Since Davyhulme does not treat phosphate, we asked for data about the wastewater they treated to provide us some realistic parameters on the expected concentration levels of phosphate.
After permission was granted, we were grateful to be given access to a small part of the data that is specifically about phosphate. We used this data in our continuous culture model to estimate the production cost of our bacteria and used Davyhulme as a real life scenario to estimate the cost of treating wastewater for a year. This allowed us to determine the economical feasibility of our project and also led to a discussion on alternative synthetic biology strategies that could be employed as a cost-reduction strategy.
Furthermore, we had a discussion with Sara Lyons, one of the technical managers of the site, in regards to the execution of our project in treating phosphate. Sara encouraged us to be more creative and to think about other ways our project can be used outside of the treatment plant. She explained to us how most of the phosphate usually originates from detergents and perhaps it might be more feasible if we can reduce the phosphate levels of the wastewater before it reaches the treatment plant and before it gets mixed up with other kinds of waste. We discussed this idea within the team and thought about incorporating our project within dishwashers and washing machines. In this case, Phosphostore must work in an environment in high temperature and pH. With this in mind, we investigated the heat stability of our three cgPPK2 constructs to assess the practicality of incorporating them into a device that removes phosphate from washing effluent.
Investigating the heat stability of our PPK constructs
At this point in the project, we had established that BBa_K2213004 was unable to be purified intact so we proceeded to investigate the heat stability of our two remaining PPK constructs: cgPPK2_His6 (BBa_K2213003) and PduD(1-20)_mCherry_cgPPK2_His6 (BBa_K2213005) through the use of a thermostability assay. The aim of this investigation was to know whether they would be functional in the warm water discharged from washing machines and dish washers. The result can be seen in the figure below:
From figure 2, cgPPK2_His6 (BBa_K2213003) showed maximum change in denatured protein between 29.8ºC - 30.6ºC; whereas the PduD(1-20)_mCherry_cgPPK2_His6 (BBa_K2213005) showed peaks at 32.2ºC - 32.4ºC. This suggests that the addition of a PduD tag and mCherry protein slightly increases the heat stability of the cgPPK2 protein. The tag-mCherry-PPK construct also consistently showed a second peak at 95.2ºC. Using Imperial2011's experience with BBa_I13521, we believe this peak is likely to be caused by the unfolding of the mCherry domain. For more information on how we completed this thermal shift assay, please check out our protocol.
With the data collected from this experiment, we realised that even BBa_K2213005 would not be functional in dish washer effluent, which are often above 50ºC. There could, however, be an application for Phosphostore in cold washes from washing machines, as these can be as low as 20ºC. With ever-advancing technology of washing powder, the number of people using cold washes may increase; however, at this time we believe this not to be the biggest market for Phosphostore.
While exploring the water industry, we decided to explore the wider entrepreneurial aspects of our project. We wanted to see how and where our project can be implemented in real life and the factors that we will have to address if we want to move our work beyond the proof of concept. We decided to tackle three factors which together comprised our business plan: intellectual property, scale-up and legislation. The full report of our business plan can be found here.
Intellectual Property: Our Phosphostore project is based on a publication that cited a granted patent for a critical part of the technology. Since none of our team members had any knowledge on patent laws, we did not know how this may influence our project or our entry to the iGEM competition. We contacted Dr. Rick Watson from the University of Manchester Intellectual Property Office and Dr. Linda Kahl from the BioBrick Foundation to learn and discuss patent laws in the context of our project and the iGEM competition. From our discussion, we were able to understand our patent situation and learned how the BioBrick Foundation is addressing issues regarding intellectual property rights for the development of synthetic biology. We also researched further and evaluated the viability of patenting our project. We compiled all of our findings in a report which can be found here. From our exploration of intellectual property, we came to the conclusion to explore other potential markets outside of the EU and US in our business plan. This is because a big majority of biotechnology-associated patents are granted in the EU and US, including patents that are associated with our project. Thus, we would not have as much freedom to operate in these two territories.
Scale-Up: In order to implement Phosphostore in real life, we would have to consider how to produce our product at an industrial scale to meet the large demand. Since the product of our project is the bacteria itself, we researched industrial bacterial culture methods and contacted John Liddell from the Centre for Process Innovation to understand how it works in real life. One of the main things we learned is that there are a lot of different factors that will have to be considered in scaling up our project: the type and size of the bioreactor, the type and amount of medium used, additional supplements needed for growth (such as trace metals), growth rate of organism and various utility cost (cost to sterilize and maintain). Thus, predicting the production cost of our project to determine its economic feasibility would be a difficult challenge. Nonetheless, John helped us by providing important information on common factors used by companies, such as molasses and glycerol as a typical carbon source in industrial cell cultures. This information allowed us to make a rough cost estimate for the production of our phosphate storing bacteria in a continuous culture system which can be found here. In addition, he also referred us to some sources that contained cost estimation on the equipment and various utility/maintenance cost. We used these sources to estimate the implementation cost of our project in our business plan which can be found here.
Legislation: Addressing safety regulation and legislation is inevitable for all synthetic biology applications that is going to placed in the real world. We wanted to explore the legislation requirements on the commercialization of our synthetic biology approach, especially as the water industry applications of Phosphostore might involve a release of bacteria to the environment. Therefore, we contacted the Department of Environment, Food and Rural Affairs (DEFRA) where the EU application procedures for any use of GMOs were explained to us. However, our exploration on intellectual property had directed us to explore other international markets as well, some of which may or may not have a GMO legislation (if they are emerging economies). We discussed this with DEFRA which told to us that it may be possible that these countries may require the applicant to gain approval in the EU first, which has a well-respected GMO regulation. We later realized that researching GMO legislation of other international countries would be a huge burden to our group. Therefore, we initiated a collaboration with other iGEM teams from international countries where each team explored the GMO legislation in their respective countries. We then compiled all of our findings into a global overview of GMO legislation around the world and reflected on this collaboration. The results of this collaborative analysis can be found here.