Human Practices

Project journey
Integrated human practices
Survey

Project journey

In order to get a compelling and challenging project, we, the Aalto-Helsinki 2023 iGEM team, undertook a series of ideation sessions to come up with potential project ideas. Multiple proposals were laid on the table, and particular attention was given to those that generated more interest among the team members.

Subsequently, our list of feasible projects was narrowed down to two general themes: biological solutions for plastic pollution and the development of sustainable food production methods. Both challenges offered a wide range of possibilities for an iGEM project, thereby making it a challenging task to make a definitive choice.

We presented our ideas to our principal investigators (PIs), Heli Viskari (University lecturer in the Department of Bioproducts and Biosystems at Aalto University) and Markus Linder (Professor at Aalto University, Vice head of the Department of Bioproducts and Biosystems), who graciously provided us with highly valuable contacts to assist in shaping our project.

  • “We decided to start contacting researchers with expertise in our areas of interest. We addressed to Rahul Mangayil and Christopher Landowski, both of whom kindly discussed our ideas and offered us valuable suggestions.”
    31.3.23
    Rahul Mangayil, an Academy Research Fellow at Aalto University, has made significant contributions to our project. He specialises in engineering microbes to produce biomaterials. Rahul suggested using enzymatic plastic depolymerization as an innovative approach. He introduced PETase, an enzyme capable of breaking down PET, and proposed engineering microbes to produce it. The process involves PET binding to the enzyme, resulting in the production of several monomers. To achieve a complete PET conversion, another enzyme called MHETase is needed.
    Rahul proposed enhancing the enzymes’ activity by using the SpyCatcher/SpyTag system, a protein ligation method. This technology can efficiently bring the enzymes together and improve the PET depolymerization process.
  • 31.3.23
    Christopher Landowski, a molecular biologist and Co-founder/CTO of Onego Bio startup, agreed to meet our team. During our visit to Onego Bio's laboratories, we discussed the advantages of microbial agriculture. It offers precise component cultivation for the food industry, utilising less space and resources while ensuring a stable supply of high-quality products. Inspired by their success, Christopher proposed exploring animal-free egg yolk for vegan mayonnaise, suggesting Pichia pastoris and Escherichia coli for fats, flavours, and aromas. He also connected us with a fat production specialist at VTT to support our ideation process.
  • “Both projects had significant interest and held the potential for a substantial iGEM project; it was very complicated for each member to select an option. Subsequently, after incorporating the experts’ advice, we refined our ideas and presented them in a more precise and articulated manner to our PIs and supervisors at Aalto University and University of Helsinki.”
    11.4.23
    In a meeting with our project advisors, Sesilja Aranko and Ville Paavilainen, we presented our proposals and sought guidance. They acknowledged the value of our plastic degradation project, particularly in optimising PETase enzyme production alongside Rahoul Mangayil's research. Sesilja's expertise in SpyCatcher-SpyTag technology offers potential enhancements for the depolymerization process. The proposal for producing a vegan egg yolk with microorganisms was considered challenging due to high fat content. Moreover, our team would need bioreactors and comprehensive compound analysis methods.
  • “At that point, our team remained impartial without a definitive preference, as both projects presented distinct, exciting challenges and goals. However, upon having a meeting with Nesli Sözer, a new perspective was given to the plan.”
    25.4.23
    During our meeting with research professor Nesli Sözer, an expert in industrial biotechnology and food production, we presented our ideas and recommendations gathered from previous discussions. Nesli, recognizing the potential of both projects - enzymatic plastic degradation and sustainable food production - proposed a merger, suggesting that the product derived from plastic depolymerization could be applied to microorganisms to convert it into a nutritionally valuable molecule. This proposal ignited genuine motivation within our team, as it presented a challenging and innovative opportunity. Nesli provided valuable guidance on the selection of suitable microorganisms, recommending bacterial strains over yeast for our project and highlighting the potential of Pseudomonas putida due to its natural ability to metabolise PET monomers without genetic modification. Furthermore, we learned that our focus should be on obtaining protein-rich end products, as they are more likely to be achieved through bacterial metabolic pathways.
  • “We wanted to explore the potential applications of our end product, hence we reached out to Shameer Kodambiyakamenna, who is one of our project advisors, seeking his valuable insights on the matter.”
    10.5.23
    We had the privilege of engaging with Shameer Kodambiyakamenna, a postdoctoral researcher in Agricultural Sciences at the University of Helsinki, whose expertise proved invaluable in exploring the potential applications of our project's end product.
    Due to European restrictions on genetically modified organisms for human nutrition, our project would have regulatory constraints. Shammeer proposed alternative applications for the protein we aim to extract. He shared his ongoing research on harvesting parasitoids to serve as natural pesticides. A compelling synergy emerged from our discussions, whereby our protein-rich biomass could be utilised as feed for these insects involved in the production. This approach ensures that our product is directed towards a practical and beneficial purpose, rather than human nutrition. Shameer's expertise and ongoing research provide a valuable foundation for exploring this innovative application.
  • “In order to perform correctly our work in the lab, we needed to seek for advisors to help us with the laboratory protocols and lab troubleshooting”
    22.5.2023
    Our team contacted Natalia Kakko, a Doctoral researcher in the Department of Bioproducts and Biosystems at Aalto University. Natalia has high expertise in synthetic biology and she is familiarised with the cloning process. Hence, she agreed to become our supervisor for wet lab troubleshooting. For the same to meet, we contacted Salla Koskela, a Postdoctoral researcher in the Department of Bioproducts and Biosystems at Aalto University. Salla is highly familiarised with Aalto University laboratories and cloning protocols. Consequently, our team asked her for support as an advisor within the lab.
  • 29.5.23
    Alesia Levanova, a postdoctoral researcher in the Molecular and Integrative Biosciences Research Programme, became our advisor via a course on mentoring, leadership, and project management. She agreed on helping us to refine our protocols and the designs of the plasmids to use. We had a couple of meetings and went over the cloning protocol, making the required adjustments for getting a better outcome. She has also provided the team with the media needed for the transformation step within the protocol.
  • “Since our proposal is quite innovative, our team needed to specify some details such as defining the optimal method of obtaining protein from the resulting product of depolymerization. Through some research, we came across the BioPROTEIN project, which shared certain similarities with our workflow. Consequently, in order to get guidance towards the last phase of our project, we decided to reach out to the person responsible for overseeing the project.”
    7.6.23
    Stephen Techtmann, an associate professor at Michigan Technological University, and his team have launched the BioPROTEIN project, which involves converting plastic waste into protein powder. They use byproducts from plastic breakdown to feed microorganisms, resulting in a protein-rich microbial biomass. In discussions with Stephen, we shared our plastic depolymerization enzyme system, while he offered insights into his chemical methods. We learned about suitable bacterial strains for transforming plastic monomers into valuable compounds. He shared details about his microorganism system, optimal conditions, and analysis techniques for the resulting biomass.
    “After Stephen’s meeting, our team agreed to incorporate Rhodococcus opacus and Pseudomonas putida as the responsible strains for metabolising TPA and EG.”
  • “During the whole process, we have been open to any kind of advice regarding the iGEM competition, hence, it was a great opportunity meeting with Marcel, who was visiting Helsinki in the summer.”
    18.6.23
    Marcel Zimmeck, a visiting team advisor from the iGEM team Hamburg, also joined us for a meeting. With his extensive experience of participating in the iGEM competition twice, Marcel provided us with practical guidelines and invaluable feedback on our current project. His insights have been instrumental in refining our approach.
  • “After some research, we considered it very important to gather information regarding bioethics. In that sense, we could implement bioethics within our project.”
    28.6.23
    In a highly informative meeting with Heikki Saxen, a distinguished bioethicist, critical topics related to reducing plastic waste and genetically modified food were discussed. We delved into the pressing issue of plastic waste, emphasising the need to prioritise safety, sustainability, and waste reduction. His valuable input guided us in understanding the potential exacerbation of the problem and the importance of comprehensive public engagement to address public concerns. Furthermore, he shared his expert opinions on the risks and benefits associated with genetically modified organisms and GMO-modified food. His valuable guidance highlighted the significance of transparent risk assessments, health and safety protocols, and a balanced approach that considers both potential benefits and drawbacks. With his input, we discussed the development of ethical guidelines to ensure responsible usage of GMOs.
  • “In order to get advice to orient the product of our project towards human consumption, we decided to contact a company which sells a product made out of very similar sources.”
    6.7.23
    Solar Foods is a Finnish startup of food technology which is aiming to produce edible food out of air using electricity to produce hydrogen. They use naturally occurring microorganisms to avoid the use of GMOs due to EU regulations. We contacted Susanna Mäkinen, head of the Biology department, and Juha-Pekka Pitkänen, CEO of the company. We shared a long discussion regarding EU regulations and GMO uses as well as the public vision towards a product produced by microbes. They shared the story of the company with us, because it has been recently founded and the work they are currently doing. Our Human Practices team presented our project, hence we received valuable feedback.
  • “We implemented in our protocols some advice received and oriented our aims towards a specific public. Consequently, we decided to contact a responsible in the Environment department in Bangladesh to discuss about waste management and our product idea.”
    18.7.23
    Shafiullah Bhuiyan, a worker in the Department of Environment in Bangladesh agreed to meet with us for discussing the challenges of waste management. It is important to be conscious of different environmental scenarios so that we can adapt the model of implementation and bear in mind problems that we weren’t considering due to the fact that they are not present in Finland. Shafiullah guided us through the different complications Bangladesh faces regarding plastic waste pollution and how it is regulated there. He shared very interesting insights and recommendations for us to implement so that we can describe a more feasible project.
  • “We were considering different ways of improving our project. One possibility was engineering the bacteria that were producing our biomass so that they could utilise the monomers given in a more efficient way. We contacted a professor with whom Stephen Techtmann put us in contact.”
    21.7.23
    We had a Zoom meeting with Ting Lu, a professor of Bioengineering at the University of Illinois at Urbana-Champaign. Research into engineering synthetic pathways into Pseudomonas putida to utilize PET monomers has recently been conducted in his group. He gave us valuable advice on the do's and don'ts of pathway engineering and also suggested that we should additionally look into conducting an adaptive laboratory evolution study, due to the time constraints.
    “After this meeting, we started to consider candidate strains and design the possible parts to be used in the engineering process.”
  • 28.7.23 - 30.7.23
    At the Nordic conference held by SDU in Odense, Denmark from 28.7-30.7 we gained valuable insights on improving our Wiki and presentation skills. Thøger Jensen Krogh, an iGEM alumni and now iGEM mentor with a Ph.D. in Biochemistry, Molecular Biology & Bioinformatics, emphasised writing the Wiki text early on and refining it later to ensure a clear and concise representation of our work. The presentation tips from Christian Eiming focused on creating a clear storyline by following the "Hollywood script formula" which includes suspense and conflict to engage the audience effectively. We intend to take all his valuable advice to heart and improve both our presentation slides and the way in which we present. Towards the end of the conference, all the teams presented their projects in front of a judging panel. Feedback from the judges highlighted the need to envision our end product more clearly for the audience. Then the Alumni panel shared their iGEM experiences, emphasising the importance of building team spirit, setting achievable milestones, and celebrating small victories. They also mentioned valuable skills like maintaining a "keep pushing” mentality, taking on responsibility, effective delegation, and setting realistic work goals. These lessons will guide us in enhancing our project's human practices and overall success.
  • 4.8.23 - 6.8.23

    The Junior Jam was held by the Münster iGEM team in Münster, Germany. In addition to defending our project presentation and poster to other iGEM teams, the Junior Jam Conference offered us the opportunity to attend very interesting workshops on the matter of human practices. The workshops presented a great opportunity to enhance your skills in talks and learn more about founding start-ups. They were offered by the REACH EUREGIO Start-up Center.

    How to pitch your start-up: during this session, the participants rehearsed a 3-minute pitch about the project, aiming to improve presentation skills. Several techniques were shared in order to learn how to keep the audience engaged.

    From Science to start-up: In this workshop, it was presented the mechanisms and resources needed in order to build a start-up from an idea, and we received very useful examples. After a very insightful presentation, we applied The Business Model Canvas to our own project. Within this activity, we had to identify our potential stakeholders, key partners, and other resources.

  • 7.8.2023
    Our team participated in the Lab seminar held in the School of Chemical Engineering at Aalto University by the Department of Bioproducts and Biosystems. We gave a presentation about our project and current results to all the researchers, receiving very useful feedback. We got to do troubleshooting and received nice suggestions in regards to how to utilise our end product and different alternatives we could offer to the public. Regarding our bacterial strains, several researchers suggested different modifications to perform in our media in order to get enhanced bacterial growth. Hence, we took into consideration alternative techniques for the analysis of the biomass as well.
    “Presenting the project to a group of experts in the field of interest was a highly valuable activity for our team. After getting feedback, we arranged a meeting with a company involved in plastic packaging management to determine the usability of PET in different sustainable strategies.”
  • 14.8.23
    We conducted a meeting with Suomen Uusiomuovi company. This corresponds to Finnish Plastics Recycling Ltd, an authorised extended producer responsibility organisation in Finland, in control of the implementation of the legislation for plastic production responsibility in companies. We presented our presentation as an alternative outcome for PET waste in order to get feedback on our idea from their point of view.
  • “We also needed professional feedback from the food production field in order to complete our revision for both sides of the project targets.”
    6.10.23
    Our team recontacted Nesli Sözer, the expert in industrial biotechnology and food production from VTT we contacted in the spring. She helped us to define our project, hence we presented our final version and the results obtained. She provided fruitful insights on how to present our idea in order to implement it as an alternative food production methodology.
    “After this meeting, we concluded our human practices work. All the meetings were very helpful throughout the whole process of our iGEM project. Keeping our project in touch with the “real world” helped us to refine and redirect ourselves and our objectives.”

Integrated Human Practices

Methodology

Understanding the problem

The plastic pollution crisis is a global environmental issue characterised by the widespread and unsustainable production, use, and disposal of plastic materials. It has reached alarming proportions due to several key factors such as overproduction and consumption, single-use plastics, inefficient recycling, persistence in the environment, its impact on ecosystems, health concerns due to the presence of microplastics in the food chain, its global reach worldwide and the environmental costs it implies. To tackle this crisis, a multifaceted approach is needed and it should include a reduction in plastic production, an improved recycling strategy, innovation, regulatory policies, and education and awareness to the public

In parallel, food security is a global challenge due to inadequate access to safe, nutritious, and sufficient food for a significant portion of the world’s population. It encompasses several issues contributing to this problem like food insecurity, climate change, economic factors, waste and loss and non-sustainable agricultural practices. Addressing this issue requires a lot of effort which implicates policy and governance, sustainable development, climate resilience, emergency response, reducing food waste and global cooperation.

Defining a solution

As the Aalto-Helsinki 2023 iGEM team, we have reached several experts working in different aspects related to these matters in order to shape our project. We needed to define a feasible and integrated solution so that these challenges could be addressed from a realistic approach. Our aim has always been to make a real contribution, which could eventually end up being really impactful.

We have addressed different aspects of these problems. Our main objective has been defining the different steps within our project to achieve successful results. Our team needed to design an adequate process, which we have been constantly defining with trial and error approaches. The guidance received from the different experts acting as advisors to build a successful workflow of experiments has been crucial. Furthermore, we have scaled the project in order to design a feasible implementation. By consulting different experts in different areas of plastic waste management and novel food production, we were able to get a realistic approach on how to present our proposal to the public and define who are our stakeholders. Moreover, we have taken into account the bioethics part of the process, due to the importance of creating an impactful project without damaging any other areas. This is also important for the outreach and educational part of the project. An iGEM project commonly implies novelty, hence, we need to take care of letting the public know about the advantages and disadvantages of genetic engineering and the real impact our project would make in the world.

Academy Research Fellow in biomolecular materials at Aalto University.

Problem

Our team had a very preliminary idea for the project, we were considering approaching the plastic pollution problem with a biological solution. We intended to explore microbe-based alternatives. By doing some research, we came across the application of some microorganisms, such as Idonella sakaiensis, in plastic degrading systems due to their capability of enzymatically breaking down plastic.

Rahul Mangayil’s research focuses on engineering microbes to secrete proteins directed to biomaterial production. Since Rahul employs these plastic-degrading enzymes for his research, we decided to contact him to seek some guidance and try to get a more innovative idea.

Summary

Given our collective interest in exploring innovative approaches to plastic waste management, Rahul proposed focusing our efforts on enzymatic plastic depolymerization. Drawing from his expertise in enzyme expression through engineered bacteria, we could adopt a similar methodology to express enzymes of interest for our project.

Rahul introduced us to PETase, a hydrolytic enzyme capable of breaking down polyethylene terephthalate (PET). To obtain this enzyme, we would need to engineer a microbial strain so that it expresses the desired protein. Afterward, we would collect the enzyme and employ it in the process of plastic depolymerization. Through this process, PET polymers would bind to the hydrolase, resulting in the production of three distinct monomers: mono (2-hydroxyethyl) terephthalate (MHET), terephthalic acid (TPA), and bis(2-hydroxyethyl) terephthalate (BHET).

PETase enzyme is naturally expressed in the bacteria Ideonella sakaiensis, which utilises PET as its primary carbon source for growth. To achieve a complete conversion of PET into TPA and ethylene glycol (EG) monomers, another hydrolase named MHETase, also naturally expressed in Ideonella sakaiensis, is required. This enzyme catalyses the conversion of MHET into TPA and EG.

Rahul had many exciting suggestions regarding the utilisation of these enzymes. An intriguing approach for our project entailed enhancing the activity of these enzymes within a unified system to facilitate a more efficient depolymerization process. To accomplish this objective, he acquainted us with the potential application of the SpyCatcher-SpyTag system, a protein ligation method. Through the implementation of this cutting-edge technology, we would be able to successfully bring these proteins together, thus ensuring a heightened efficacy in PET depolymerization.

Implementation
  • Checking 2016 New Zealand's iGEM team project, in which they transform E. coli to produce PETase enzyme.
  • Researching on PETase and the depolymerization process
    PETase enzyme chemical structure and reaction products from depolymerization. MHETase enzyme is needed as well in order to get complete depolymerization. PET: polyethylene terephthalate, BHET: bis(2-hydroxyethyl) terephthalic acid, MHET: mono(2-hydroxyethyl) terephthalic acid, TPA: terephthalic acid, EG: ethylene glycol. https://www.mdpi.com/2073-4360/15/7/1779
  • Research on the SpyTag-SpyCatcher technology, which would introduce an innovative part to the process of obtaining PETase and MHETase enzymes from engineered bacteria since we would be optimising the depolymerizing process. The Dry lab team designed the constructs to engineer E. coli with this technology.
    Designed SpyCatcher-FAST-PETase fusion (BBa_K4701302), and SpyTag-MHETase fusion (BBa_K4701303) parts.

Molecular biologist, Co-founder, and CTO at Onego Bio startup, he works as a cellular agriculture trailblazer and is the inventor behind the Bioalbumen® technology. His work enables the production of egg white without the need for chickens. This is achieved by utilising precision fermentation technology, employing a microflora called Trichoderma reesei as a living factory.

Problem

The proposal for creating an innovative and sustainable food product was very preliminary. Our team wants to determine what is needed to actually obtain a nourishing product out of bacterial cultures. Moreover, GMO regulations are highly strict towards human nutrition. Hence, if we go for this approach, we would need to consider how to include an engineering cycle, which is required for the iGEM competition.

Summary

During our visit to their laboratories, we had a discussion regarding the advantages of microbial agriculture in comparison to traditional agricultural practices. Microbial agriculture offers the advantage of cultivating precise components specifically intended for the food industry, while requiring significantly less space and resources, providing a stable and controlled high-quality supply of products. Given the success in producing egg white, Christopher’s team suggested that we explore the possibility of obtaining an animal-free egg yolk, thereby enabling the creation of vegan mayonnaise. To achieve this, Christopher proposed the utilisation of Pichia pastoris (yeast) and/or Escherichia coli as tools for generating the necessary fats, flavours and aromas. He also provided us with a contact from VTT, who specialised in the production of fats in yeast, in order to facilitate our ideation process.

Implementation
  • Research about different yeast strains to use simultaneously to produce several recombinant components needed such as lipids, flavouring components, and proteins.
  • The downstream process ensures the safety of the final product and facilitates future marketing (like removing the cell debris).
  • Start considering which approach would be useful for our product, like a marketing strategy. A good idea would be to direct it towards spaceships, areas with limited agricultural resources, and then eventually become a staple in everyone’s daily diet.

Remaining problems: targeting the specific production of protein or other components and how to include the engineering cycle.

Sesilja Aranko, a staff scientist in the Department of Bioproducts and Biosystems at Aalto University, and Ville Paavilainen, the research director of the Institute of Biotechnology at the University of Helsinki. Sesilja and Ville helped us in project ideation early on.

Problem

After delineating the two project proposals, the team embarked on the task of knowing which option was more feasible to develop. We needed to consult with our advisors so that we could receive their feedback and check with them if there were suitable facilities. Given that the two proposals require different methodologies, it was necessary for us to determine if the approaches were doable as well as to check the availability of expert guidance during the experimental phase.

Summary

After reviewing our plastic degradation project, Sesilja and Ville recognized its potential value in contributing to the optimization of PETase enzyme production, particularly in synergy with Rahoul Mangayil’s research. Sesilja’s expertise in the SpyCatcher-SpyTag technology could significantly enhance the depolymerization process by facilitating the closure of the two enzymes required, PETase and MHETase. In the context of our proposal for vegan egg yolk production using microorganisms, Sesilja and Ville regarded it as a more challenging option. Egg yolks are characterised by a high fat content, while bacterial cultures typically produce proteins easier. Additionally, the production of such a product would need the use of a bioreactor. Moreover, a comprehensive method for extraction and analysis of the various compounds within the product would be indispensable.

Implementation
  • Define a more realistic approach for both proposals taking into account the materials that we would need.
  • Determine which ones would the sources of our input products for the project be.
  • Contacting Rahul Mangayil, and him becoming and advisor for the team in regards to his expertise.
  • Contact VTT in order to see if we are allowed to use a bioreactor.

Nesli Sözer, a research professor at VTT specialised in industrial biotechnology and food, with expertise in the production of nutritionally valuable molecules using cultured microorganisms.

Kari Koivuranta is a principal investigator at VTT working in the Production Host Engineering group in the Industrial Biotechnology business area.

Problem

Our different approaches for the iGEM project woke up a lot of interest in most of our members. The production of the vegan egg yolk project seemed very challenging taking into account the facilities we had available. We contacted Nesli to see if we had the possibility of adapting this option to a more feasible project.

Summary

Our team presented our ideas and recommendations gathered from previous meetings. Given the intriguing nature of both projects - enzymatic plastic degradation and sustainable food production - Nesli proposed a merger of these initiatives. Consequently, the resulting product derived from plastic depolymerization would be applied to microorganisms to convert it into a nutritious molecule. This proposal gave rise to genuine motivation within the team, as it presented a challenging and innovative opportunity.

Nesli and Kari provided us with valuable guidance regarding the most suitable options for microorganisms and the desired molecules to be obtained. We comprehended that, for our project, bacterial strains would be more appropriate than yeast, as yeast is incapable of utilising the resulting monomers from PET depolymerization. Another noteworthy recommendation was to consider Pseudomonas putida for acquiring our protein, as it can naturally metabolise PET monomers, eliminating the need for genetic modification. Additionally, our aim should be to obtain an end product rich in proteins, as they are less complex molecules and more likely to be obtained through bacterial metabolic pathways, rather than lipids or other compounds.

Implementation
  • Merge our project proposals into a whole process where we use PET plastic as the primary source to produce protein.
  • Do research on the PET depolymerisation process, the resulting monomers and how to transform them into protein.
  • Do research on P. putida and the metabolic pathways that would make possible the transformation of TPA and EG monomers into proteins.
  • Do research on other microbial strains and their ability of utilising TPA and EG as a carbon source.
  • Contact the people responsible for the Microbial Domain Biological Resource Centre (HAMBI) at the University of Helsinki in order to see which strains are available for us to use.
  • Division of the project in two parts:
    1. Plastic microbiological depolymerization, PETase and MHETase obtention by optimising the microbial production protocol for the production of these enzymes.
    2. Protein-rich biomass production from the depolymerization products utilising bacterial cultures.

Postdoctoral researcher in Agricultural Sciences at the University of Helsinki.

Problem

We were not familiarised with EU regulations towards GMO utilisation in the human nutrition field even though we are aiming for a nourishing end product. European regulations are quite strict regarding the utilisation of genetically modified organisms. We needed to consider an alternative application for the outcome of our project that could be implemented nowadays. We presented our project to Shameer, who offered us some suggestions on how to effectively utilise the biomass generated by our cultured bacteria.

Summary

The methodology we employ to acquire biomass comprising nutritionally valuable molecules involves the use of genetically modified organisms. Therefore, due to European restrictions on genetically modified organisms, our project cannot be directed toward human nutrition. Taking this regulatory constraint into consideration, Shameer wanted to share potential applications for the protein we intend to extract.

He acquainted our team with his ongoing research and the laboratory facilities at his disposal. Shameer is currently working on harvesting parasitoids of Goniozus nephantidis species, employing larvae of the rice moth (Corcyra cephalonica) as hosts. The purpose of this collection is to subsequently release these parasitoids into crops, where they serve as natural pesticides against the Opisina arenosella plague, a significant pest in coconut production systems. The larvae are fed using a diet comprising semolina mixed with other nourishing components.

Considering our project, an intriguing synergy emerges whereby our protein-rich biomass could be utilised as feed for these insects employed in the production of natural pesticides. This approach ensures that our product is not intended for human nutrition, but instead contributes to a practical and beneficial purpose.

Implementation

Consider the alternative possibility of offering our product for the natural pesticides field. We need to apply a realistic view to the project and check for other options that could be approved with the current European regulations.

Associate professor at Michigan Technological University, creator of the BioPROTEIN project: Turning Trash Into Treasure. This innovative endeavour involves obtaining a protein powder derived from plastics. The team employs byproducts obtained from plastic depolymerization to nourish a consortium of microorganisms, which metabolise these substrates and yield microbial biomass abundant in protein and other nutritional compounds. The primary objective of the project is to direct this product as a potential food alternative.

Problem

The second phase of our project needed some refinement. After doing some research on TPA and EG utilisation, we did not have an exact approach of the most optimal strain for us to use in the biomass production part. We came across Stephen's research and we decided to contact his group. Moreover, his product was presented as an alternative protein source, hence our group was intrigued about these matters.

Summary

During our meeting, we had the opportunity to exchange details regarding our respective projects with Stephen. He showed interest in our enzymatic system for plastic depolymerization, as he employs chemical methods in his research.

Conversely, we received valuable knowledge about the most suitable bacterial strains for effectively transforming TPA and EG monomers into valuable molecules, particularly protein. Stephen shared with our team the various microorganisms utilised in his system for obtaining biomass. We discussed the optimal conditions and combinations of different bacteria for metabolising the monomers. Additionally, he provided our team with guidance regarding the analysis of the resultant biomass, elucidating the methodologies employed and recommending useful bioinformatic tools for further analysis.

Implementation
  • Incorporate other bacterial strains in the biomass production phase. The more strains, the more possibilities we have to obtain a profitable biomass.
  • Research for Rhodococcus opacus, Comamonas testosteroni and Bacillus subtilis as candidate strains for TPA and EG utilisation. We need to determine if they can utilise the monomers and the conditions in which these strains would grow better.
  • Consider using disodium TPA for better incorporation in the minimal media.
  • Research on bioinformatic pipelines for the assessment of the security of our biomass.
  • Incorporation of the Bradford protein assay in order to determine the amount of protein that we obtain in our end product.
  • Consider engineering other strains in order to enhance monomer utilisation. In this matter, Stephen connected us to Ting Lu, an expert within the field.

Bioethics researcher, member of the board and founder of the Institute of Bioethics in Finland.

Problem

Our proposal showed some difficulties in terms of producing a product for human consumption. The reason is the utilisation of transformed bacteria for producing our enzymes, it implies using GMOs throughout the process. After our meeting with Shameer, new possibilities came across. We had to bear in mind all the possible regulations in every different option. According to iGEM regulations, there are several organisms which cannot be engineered, however, it was not clear to us if we were allowed to try our product obtained through a process that uses GMO bacteria in Corcyra cephalonica. Moreover, we wanted to get more insights about the ethical impact this idea can have as well as for the idea of using our protein-rich biomass in food intended for human consumption.

Since we are not familiarised with the ethical implications that our process can have in science and society, we decided to contact Heikki due to his expertise in the field.

Summary

During our meeting with bioethicist Heikki Saxen we received some very valuable inputs. One key suggestion was to consider the overall goal of our process and prioritise safety and sustainability. With this in mind, we made sure to evaluate the potential risks and benefits of genetically engineered organisms (GMOs) that we plan to use in our project. We conducted a thorough risk-benefit analysis, assessing the impact of these organisms on the ecosystem and the environment.

Our laboratory procedure and ambitions were kept in line with the philosophy of having more good effects than harmful ones. We also emphasised the importance of public engagement and trust, learning from previous experiences such as the COVID-19 vaccine rollout, to address potential concerns and communicate effectively with the public. By approaching the analysis with humility and patience, we aim to establish a solid foundation of ethical practices and ensure the safe usage of GMOs.

Stakeholder engagement was another crucial aspect highlighted by Heikki Saxen. Great emphasis was given to maintain a humbleness in our approach to allow for proper communication to our stakeholders.

He also emphasised the importance of transparency and public engagement throughout our project.

Furthermore, Heikki Saxen emphasised the importance of transparency and public engagement throughout our project. One of the key aspects to consider is that multinational companies dominate the GMO industry, which significantly contributes to the negative outlook of the common people about GMOs. A fresh perspective from academia, with the right level of education and science communication, has great potential and a relative advantage in gaining trust from potential consumers and the general public.

Implementation
  • Development of a comprehensive ethical guideline focused on responsible synthetic biology applications in waste management and alternative protein production. It provides best practices and ethical considerations regarding our project. It can contribute to responsible research and innovation in the scientific community. This guideline will be available for other teams to follow, adding up as an important contribution to the overall iGEM ecosystem.
  • By fostering a collaborative and inclusive approach, we aimed to address diverse viewpoints and incorporate them into our decision-making process effectively:
    1. Engaging relevant stakeholders in discussions and decision-making processes to gather all the different opinions and concerns.
    2. Take into consideration the interests and perspectives of non-profit academic institutions, ensuring their involvement and contribution to the project.
    By striving for a uniform point of view while considering different stakeholder opinions, we ensure that our project not only adheres to ethical guidelines but also fosters an inclusive and participatory environment.
  • Prioritising transparency and open communication, sharing risk assessments, health and safety protocols, and experimental procedures. We have provided access to our scientific information through our website and engaging transparently to the public in our work.

A  Finnish food technology company revolutionising global food production with their approach, which implies renewable energy and bioprocess engineering. In this way, food production is detached from agriculture. The final product is a protein made with cellular agriculture. Microbes are fed with air (carbon, hydrogen, oxygen and nitrogen) and electricity so that they perform fermentation.

Susanna Mäkinen is the head of the Biology department, she is in charge of the experiments and of discovering strains, and Juha-Pekka Pitkänen, CEO of the company, answered our questions.

Problem

During the ideation process, we have been having doubts about the final use of our end-product. One of the problems we aim to tackle is food security. We already had some alternatives in mind after Shameer Kodambiyakamenna’s meeting in order to not use our product for human consumption. However, we wanted to gather information and advice to acknowledge what steps are needed in order to drive our project towards food for human consumption. Moreover, we considered it interesting to know their approach to reach people and get them to know their work and product.

Summary

Juha-Pekka, the CEO, was already working with biomass production for different purposes. Then he came up with the idea of biomass aiming at food production. Their team screens microorganisms from the Finnish nature to find appropriate microbes that occur naturally, avoiding GMOs.

Susanna pointed out that an important factor for adapting the bacteria to an alternative carbon source is the conditions in which the microbes grow. It needs to be considered that providing the monomers is not enough, it needs to be checked if any inhibition can happen within the microbes metabolism or in combination with other compounds in the media. Moreover, we could perform the laboratory evolution assay, picking up the best strains and the best colonies growing in specific conditions. By culturing the bacteria several rounds, they improve their tolerance to the new environment.

She also shared their procedures regarding safety procedures and analyses to ensure the security of human consumption products.

Juha-Pekka remarked to our team the importance of safety testing, especially for legal regulations. Most of the studies required for approving a project of these characteristics are related to the microorganism used. Furthermore, in terms of public perception, Juha recommended us to be as transparent as possible and to demonstrate energy and time-wise that our proposal implies a real improvement.

Implementation
  • Research and metabolic flux analysis of our microbial strains and TPA and EG. In this way we predict expected results towards the growth of the strains with the different monomers provided in the media. Moreover, this helps us to determine which metabolic pathways need to be taken into account in order to design new plasmids for genetically engineering the strains. We perform this in an attempt to get full profit out of the monomer utilisation.
  • Performing laboratory evolution assay in order to improve the tolerance of our strains towards TPA and EG monomers. In that way, our strains consume the monomers naturally and we can avoid GMOs.
  • Calculations for the energy consumption of our whole process. We need to determine how much it would cost for us to produce protein in order to write a project implementation plan.
Growth of R. opacus after the laboratory evolution assay in a minimum media with TPA (1) and EG (2) as carbon sources.

He is a waste management engineer in the Department of Environment in Bangladesh, where they control and organise plastic waste management.

Problem

Since one of our team members is from Bangladesh, we were very conscious of the problems plastic contamination drives in the country. It is the cause of severe clogging that contributes to floods in the city. Besides, this type of pollution and its direct consequences are factors contributing to illnesses or physical accidents. Our team then assumed that our approach could be something highly interesting to address in other countries as an alternative to plastic waste management. We were also interested in getting feedback on the feasibility of the project from a person who faces a very different reality than in Finland regarding plastic waste.

Summary

Bangladesh faces severe plastic pollution problems. It is a highly populated country and it lacks proper waste management systems. We presented our project and asked him several questions about the current situation and future prospects of plastic waste management in Bangladesh. More than 90% of plastic waste in Bangladesh is single-use plastic, which poses a major challenge for degradation and recycling. However, plastic waste is not separated at its source in Bangladesh, but mixed with other household materials. This makes it difficult to collect and sort plastic waste for further treatment. The government of Bangladesh is aware of the plastic pollution issue and is working on a new policy to control plastic waste production and disposal. They aim towards rules on plastic manufacturers to justify the disposal of the products and promote biodegradable alternatives for plastic items. Feasible projects that aim for plastic degradation are of huge value in countries like Bangladesh, the waste management sector is open to new ideas based on biotechnology or synthetic biology. However, the circular economy applied to plastic waste management in Bangladesh is not very feasible given the high demand and low supply of plastic materials. The focus is rather on reducing, reusing, or recycling plastic.

Implementation
  • Refinement of our problem statement and value proposition to emphasise the need and potential impact of our solution for managing PET plastic accumulation in countries like Bangladesh.
  • Taking into account how our project could be implemented for other kinds of plastics, single-use plastics, since all of them contribute to the plastic waste crisis.
  • Adjust our design criteria and specifications to account for the challenges of collecting and sorting mixed plastic waste in low-resource settings. In the future, when implementing PET-2-Protein, it is highly important that the treated plastic is isolated from other kinds of waste.
  • Exploring the ethical, social, and environmental implications of our solution for different stakeholders, such as consumers, producers, regulators, and communities in Bangladesh.
  • Identifying potential partners and collaborators who could help us test and implement our solution in Bangladesh, such as universities, NGOs, or local authorities.
  • Developing a communication strategy to raise awareness and educate people about the benefits and limitations of our solution, as well as the importance of responsible plastic consumption and disposal. It is important to refine the way of communicating the objective of enzymatic depolymerization so that the population is willing to collaborate in terms of waste sorting.

Professor at the University of Illinois Urbana-Champaign whose primary research area is synthetic bioengineering.

Problem

The second phase of our project implies the utilisation of TPA and EG monomers by microbes, obtaining nutrient-rich biomass. After exploring the metabolic pathways of different candidate strains, we concluded that, out of the strains that are available for us to employ, Rhodococcus opacus and Comamonas testosteroni are the microorganisms we should use for TPA utilisation. Both can naturally use the monomer. However, we intend to optimise the metabolic pathways needed in order to obtain an enriched end product. Hence, we consider Escherichia coli and Pseudomonas putida to achieve high utilisation of both monomers after engineering modifications. To get advice regarding this process, we contacted Ting Lu, whose research focuses on microbial synthetic biology applied to plastic degradation.

Summary

In his engineering process, Ting aims for microbial utilisation of TPA and EG monomers, as well as for enhancing amino acid or fatty acid content in the resulting biomass. Enhancing the biomass is the result of several trial and error assays after coming up with the modelling pathway. Based on the consumption analysis, his team has refined the utilisation of the monomers and the composition of the resulting product. He recommends our team to dig into the literature, so we can either build our own consumption pathways or improve existing ones.

Regarding the strains used in this process, it is known that Rhodococcus opacus and Comamonas testosteroni can utilise the monomers naturally, therefore, this natural capacity only needs to be enhanced, it is an easier process.

Regarding the engineering target, Ting’s team is centered on enzyme engineering and metabolic network topology, so that the production of biomass is optimised. He recommended us to look for the bottlenecks of the metabolic networks that are activating with our input and engineer the enzyme to alleviate it or engineer a parallel pathway. These bottlenecks can be identified in several ways. HPLC analysis is the most simple approach. By checking the availability of the different metabolites along the metabolic pathway you can determine which are some conflictive points and see the different rates and accumulation of metabolites.

A different feasible option is to model the growth medium itself in order to provide to the microorganism sources that could be helpful for the assimilation of the monomers. When tackling specific nutritional products, Ting’s team focuses on fatty acids and some amino acids. The choice of those is a combination of the demand for the macronutrient and the feasibility of producing them. His recommendation is to identify the potential molecule that we aim for based on these criteria and try to get it. A perspective to give to our project is to scale it down into an “all together” process, where one step goes after the other automatically.

Implementation

Designing the parts that we would use for engineering P. putida and E. coli in order to enhance monomer utilisation and protein production.

As explained here we designed parts that should in principle allow P. putida and E. coli to utilize TPA and EG as carbon sources respectively. The design of both plasmids was carefully done based on research for improving the assimilation pathways of both of the monomers.

During the Junior Jam conference, in Münster, several members of our team attended a workshop organised by the REACH EUREGIO Start-up Center.

Problem

Within the iGEM process, it is needed to have a clear idea of who the stakeholders interested in our product would be. As a team, we had not previously any concrete thoughts regarding this matter since it was complicated for us to establish the stakeholders interested in our product in the real world.

Summary

During the presentation of the workshop, the organisers presented the mechanisms and steps included in the development of a start-up, starting from the ideation phase to its implementation in the real world. In order to build a start-up, several aspects need to be taken into account. They described them to us and proposed a very useful activity for describing different factors that our project would need in order to present it as a valuable idea to build a start-up from. They guided us throughout the activity and we were able to clarify several doubts regarding the stakeholders related to our project. Listed below is the outcome of our workshop:

  • Key partners (motivations for partnerships: optimization and economy, reduction of risk and uncertainty, acquisition of particular resources and activities)
    1. Universities
    2. Vegan industries
    3. Plastic waste management companies or organisations
    4. Key partners would provide equipment and financing
  • Key activities (what key activities do our value propositions require? Which are our distribution channels? Customer relationships?)
    1. Value propositions requirements: Up-scaling our project, Research & Design, Enzyme development
    2. Distribution channels: we would be provided with PET plastic waste from industries and distribute our product to other industries related to food production (vegan industries for example).
  • Key resources
    1. PET plastic
    2. Financing
    3. Laboratory equipment
  • Cost structure (what are the most important costs inherent in our business model? Which key resources are most expensive? Which key activities are most expensive?)
    1. The cost structure would be mainly directed towards lab equipment, upkeep of the idea, maintenance of the facilities and salaries.
  • Value propositions (what value do we deliver to the customer? Which one of our customers' problems are we helping to solve? What bundles of products and services are we offering to each customer? Which customer needs are we satisfying?)
    1. Values: sustainable plastic recycling, green responsibility for companies, addressing the climate crisis, social awareness, innovation and technology
    2. Customer needs/offered to customers: non-meat protein source
  • Customer relationships (what type of relationship does each of our customer segments expect us to establish and maintain with them?)
    1. Automated service
    2. Push-pull logistics
    3. We still have to scale up the system
  • Channels (how do we raise awareness about our company's products and services? How do we help customers evaluate our organisation's values?)
    1. We have started by organising educational activities to raise awareness of genetic engineering.
    2. Community outreach can be done by social media.
    3. We would attend company fairs since our main target of our end product would be vegan companies
  • Customer segments (for whom are we creating value? Who are our most important customers)
    1. Food-producing companies that look for meat alternatives
    2. Biotechnology companies
    3. Food industry in general
    4. Waste management industries/companies so that they can provide our main source.
    5. Research institutions
    6. The green conscious public in general
Implementation
  • Specific determination of our values and stakeholders in order to present our idea as if it was a proposal for a start-up.

A producers' association in the packaging industry that promotes packaging recycling

Problem

One of the main focuses of our project is plastic waste reduction in order to support the environment. As a team that is developing a project with the aim of implementing it in the real world, we need to make sure that our proposal is feasible, realistic and that it will contribute somehow to the problem that we are trying to solve. The Suomen Uusimuovi company is in charge of the producer responsibility in Finland, which is mandated by both Finnish and European laws. This principle ensures producers are responsible for their products' lifecycle and the related costs.

Summary

We delved into plastic recycling, understanding the mechanical processes and the emerging trend of chemical recycling. They explained how different plastics are sorted separately before turning them into small pallets as well as the two different recycling techniques that are more common, chemical and mechanical. Our team introduced our project and they remarked that in Finland, PET bottles have an impressive 95% recycling rate, making them some of the most recycled products globally. Our conversation also explored public perceptions of plastic packaging and the potential of replicating Finland's efficient waste management systems in countries with unique challenges. We wrapped up with insights into global partnerships like EPRO and the Circular Plastics Alliance.

Implementation
  • One of the key insights was that the use of single-use plastics was also really important for Finland. So for our long-term plan, we also kept in consideration how in the future our project can be used for the degradation of single-use plastics.
  • Taking into account that legislation has clearly defined how recycling should be done. European laws do not take into account chemical recycling as recycling. Consequently, our enzymatic degradation could participate as a clean and sustainable alternative for recycling. Hence, we could further explore how our project can contribute to Finland's ambitious target of a 50% recycling rate by 2025.
  • Understanding the concept of producer responsibility as mandated by Finnish and EU laws.
  • Discussing public perceptions of plastic packaging and the potential for replicating Finland's waste management systems in other countries.
  • Global Sustainability Alignment: The project's focus on circular economy aligns with global sustainability benchmarks. By addressing the challenges of plastic waste and food production, the project contributes to several United Nations Sustainable Development Goals (SDGs).

A  research professor at VTT specialised in industrial biotechnology and food, with expertise in the production of nutritionally valuable molecules using cultured microorganisms.

Problem

To wrap up our project as a cycle, we got in touch again with Nesli, who agreed on revising our overview of the project, since she guided us towards our definitive project idea. Nesli is an expert in the novel food production field, therefore her advice could be very valuable in terms of our second problem to tackle: food security. We needed to define how our product can contribute to the field and how feasible it is to implement our idea in the real world, targeting novel food industries. Moreover, our team wanted to get her advice on how to present our idea to the public to make it more attractive and friendly as a sustainable alternative to food production.

Summary

During the meeting, we presented Nesli the final version of the process of our project. From her point of view, it is a proposal that adds up to the problems we are tackling. However, she pointed out that it is important to exclude E. coli as a strain for producing our biomass, since some strains can cause human disease. Moreover, she suggested specifying how to analyse our biomass so that it does not contain any microplastics and then downstream the process to the purifying or extraction steps of our protein product so that there is no harm when it comes to protein production. Regarding the techno economics of the project, our team should quantify the amount of plastic that can be converted into protein to prove the sustainability of our proposal. Then, suggest alternatives on the uses, for example feeding animals and how to include it into the food chain.

We also discussed with Nesli about the European regulations since we have been working with GMOs. It is important that we show how we isolate our modified bacterial strains from the final product. Furthermore, she showed us examples from other countries than Finland, where they are more flexible with these kinds of approaches. The regulations are very heavy, we are lagging behind other countries such as the United States. They already produce several products that require genetic modification, like the Impossible Burger, which is already commercialised and they are made with heme, a GMO soy ingredient produced in yeast. Besides, Singapore has available to consumers the first cultured meat and they are also going to start commercialising with the Solar Foods ice cream, which is made with bacterial cultures. Another example given was the United Kingdom, where they are getting more flexible with the genetic modification regulations in plants. We should embrace these examples and remark that is not an immediate approach but it shows very positive points and can really support and improve the food security problem.

Nesli provided some advice regarding our project presentation for the Grand Jamboree, remarking the importance of showing the impact that our proposal can make. All in all, it was a very fruitful and useful meeting.

Implementation
  • Give our project the sustainability data in order to prove that it can really make a difference.
  • Point out the advantages of microbial food production and the use of Genetically Modified Organisms. The changes are within the process but the end result, hence, the food, remains equal and safe.
  • Checking the “food grade” of the bacterial strains that we are using, for instance, proving that the organisms that we employ are safe for human consumption and meet specific regulatory standards. R. opacus and C. testosteroni meet these requirements, as well as P. putida. However, E. coli does not, consequently we need to propose alternatives. We have used this organism because of the low availability of different strains within our facilities and due to the fact that it needs to be a fast process for us to get results. But when presenting it to the public, we can propose alternatives since we would have more time to try our process in other strains which are more acceptable in this field.
  • Prepare an impactful presentation pointing out the importance of the problems that we are addressing.

Survey

In order to get a better picture of the public’s view on plastic and genetically modified organisms (GMOs) we created a survey and translated it into 10 languages; English, Finnish, Swedish, Spanish, Bengali, Persian, Indonesian, Vietnamese, Urdu and Portuguese. By translating the survey we hoped to reach a wider public and make it more inclusive. The objective was to collect insights on plastic usage, food production, and consumer behavior. The survey covered five main areas: Demographics, PET Plastic Awareness, Understanding Food Production Challenges, Acceptance of Innovative Foods, and Research on Converting Plastic Monomers into Microbial Edible Biomass, comprising a total of 19 questions.

We received a total of 253 responses. Most of the respondents (58.9%) were aged between 18 to 24, with a majority being female (62.5%). Furthermore, a significant portion (82.2%) of the respondents had either a bachelor's or master's education level. It is important to note that the survey may have a bias towards a certain age group, gender, and education level.

The survey results are available through this link. On the first page, you will find what people think about plastic, and the second page, indicates their thoughts on food production. Almost everyone (96.94%) knows about different types of plastic, showing good awareness. When asked about the reasons why plastic is perceived as a problem in their respective countries, a significant 93% emphasized environmental pollution, underlining a collective concern for the planet. Moreover, 65% and 64% acknowledged health risks and negative societal impact, respectively, reflecting an awareness of broader implications beyond just environmental concerns.

A substantial 89% expressed a notable level of concern, ranging from moderate to extreme, regarding the plastic's impact. This indicates a strong desire for positive change and a willingness to address the challenges posed by plastic. Furthermore, it is encouraging to see that 62.4% of respondents consistently recycle plastic, indicating a proactive effort toward sustainable practices. However, 4% said they never recycle plastic, potentially due to a lack of recycling facilities and knowledge. This suggests a critical need for improved infrastructure and education to encourage sustainable habits. On the other hand, for those who do recycle, the key motivations are waste reduction and a sense of personal responsibility. This implies a growing understanding of individual roles in mitigating the plastic problem and the desire to contribute positively to the environment.

In the second part of the survey, we looked into concerns about food production. The top three worries were the environmental impact, societal effects, and the cost of food. A significant 70.7% of respondents expressed a notable level of concern about how food production affects the environment. This shows a clear understanding of the broader impact of food production beyond just personal consumption.

We also wanted to know what factors guide people when buying food. The top four factors were hygiene, flavor, nutrition, and affordability. This suggests that people value both the safety and taste of food, as well as its nutritional value and cost.

When it comes to GMOs as food, 58% of respondents were either neutral or not familiar with the concept. This indicates that there's a need for more information and education about GMOs and their role in food production. Interestingly, safety emerged as a major concern for people regarding microbial food, with 80% stating it would influence their willingness to consume it. Safety is a critical factor for gaining public acceptance and trust in innovative food technologies.

Lastly, it is worth noting that 44.54% of respondents felt hesitant but open to trying food products derived from plastic. This indicates a level of curiosity and potential openness to innovative solutions for addressing food production challenges.

Overall, the survey offers a clear view of what people think about both plastic and food production. It shows that people care about the environment and want food that's safe and good for them. Interestingly, they're open to trying food made from unexpected things like plastic. This highlights the need to educate people about innovative food technologies and make recycling plastic easier.