Team:MADRID UCM/Human Practices

Cloning design - 4C_FUELS

All the 4C_Fuels team believes that Human Practices is an essential part of our project, which has been of tremendous importance throughout its entirety development. To approach this crucial part of any iGEM project, we have decided to use The Human Centered-Design Process. It is an approach that enables us to tackle nowadays’ complex problems and engage with the local community around us.

Our Approach to Human Practices

We aim to interact with the society we live in by promoting scientific communication and seeking advise from experts. In this way, we are capable of creating feedback loops where our key stakeholders are related to each other and it gives each segment the capacity and opportunity to contribute towards the overall change needed towards achieving a balance between social, technological and economical sustainably in order to change the world.

As Peter Carr said, “Human Practices is the study of how our work affects the world and how the world affects our work”, and in order to transmit our values, we have adopted the Human Centered-Design Process.

Human Practices Trhough Human Centered Design

The term Human Centered-Design (HCD) is relatively contemporary. It was first used in 1999 when the international norm ISO 13407 was published, but it was not until 2010 when the last version of the document was reviewed that it took considerable relevance in today’s transformation norms.

According to ISO 9241-210:2010, the Human Centered-Design process is an approach to design from a perspective of end-users, their needs and requirements, thus their satisfaction, by applying human factors, knowledge and techniques. Another definition depicts it as a philosophy that empowers a team to pursue the design of products, systems or experiences that address the core needs of those who are experiencing a need or problem.

For us at 4C_Fuels, HCD is a problem-solving approach that focuses primarily on deciphering the needs of a group of potential end users to then find the way within our team to reunite forces from the different disciplines we work with towards a common goal. In order to do that and following up with iGEM’s ideals, we want to create feedback loops and collaborations not only between us (creators) and our end-users but also between those who may be affected by our work, in order to find solutions together.

Overall, Human Centered-Design is a process, a philosophy and a framework that enables personal and professional growth for users and suppliers. It has been proven that when there exists a correct time management and the ultimate goals are broken down into smaller tasks, the productivity of the team increases and everyone is able to be creative and express their inputs in the design process obtaining an efficient work methodology. As a result, the user receives a product or service which is intuitive for them, supposes a competitive advantage and allows for the organization to reduce training and support costs, contributing to sustainable development. The projects that have introduced a Human Centered-design approach have a higher likelihood to be completed on time and within budget, moreover they are more prone to be successful and viable in the long-run compared to others.

While the Human Centered-Design has many forms, the model we use at 4C_Fuels includes 5 phases that are: Problem Identification, Inspiration, Ideation, Experimentation & Iteration and Results & Implementation which will be explained in more detail next.

The Challenge of a sustainable Society

Climate change is a real danger to humanity. We believe the way society operates nowadays is unbearable and unsustainable if nothing is done. There is currently still a lack of compromise to take care of the environment as a continuum, and so first of all a shift in society’s mentality is needed to transform our modus vivendi towards one where global sustainability lies at the top of the priorities. This is only possible if we understand sustainability holistically from its three main pillars: economic, environmental and social approaches. To know more about the three main aspects of sustainability, visit our sustainable development page .

The first step in addressing the problem is understanding who will be our final users and stakeholders that may help us and guide us along the way in order to be concise in our goals along this project. Research on the key stakeholders who may be affected by our idea of finding global sustainability or that their activity affects ours have been conducted initially and is of importance in this section.


Stakeholders are essentially groups of people that affect or are affected by the activity of 4C_Fuels. If we want to achieve a holistic change in society in terms of sustainable development, we need to take into consideration groups that relate to the traditional refinery, governments, the scientific community, students, logistics, biotechnology sector, etc. The ultimate aim is to raise awareness through public engagement of the need for a change towards a more sustainable society by transferring knowledge between the parts and promote innovation and research in novel areas. The key groups of interest that best match our project were identified and classified in 4 main groups: General Society, Industry and Producers, Institutions and R&D and End-users. We are proud to announce that we have established closer relationships with some entities belonging to more than one group, which have become partners or sponsors of our project.

Main Stakeholders

General Society Stakeholders

Young Population & Students: New generations to come. They could live in a society where their needs have been compromised due to the bad management we have done today, or in a healthier and more sustainable world.

Logistic-workers: Directly affected by the production of biofuels since most of the heavy logistic applications are not fully electrificable and lag behind the current sustainable development goals

General society: There is a need for mass conviction that a greener future is possible and it can potentially be done now. Only if the majority of our society is convinced of this premise, will it work as they will be the ones who with their vote could press governments to act as soon as possible.

Stakeholders in Industry and Manufactuiring sectors

Biotechnology & Microalgae industry: New generations to come. They could live in a society where their needs have been compromised due to the bad management we have done today, or in a healthier and more sustainable world.

Biorefinery Hub : Biorefinery concept aims to establish a more biological centered economy. Aiming to decentralize industrial production exploiting the availability of local biological resources.

Renewable Tech Companies: Whose business operates in favor of energy efficiency and new renewable technologies, such as Acciona and develops projects related to sustainability.

Stakeholders in Research & Academic Institutions

Research institutions Supporting innovation and discovery. Helping developing other projects from the field and contributing with new information or data not known.

Educative Centers: Universities, schools and the people involved in them are a crucial part of sustainability, accounting to promote critical thinking, raise awareness and transfer the required knowledge for the development of our society.

Guvernamental Institutions: Take the general concerns and ideas of the general population and shape them in the form of regulations and laws. Only understanding and collaborating with them a truly technological implementation could be achieved.

End Users as Stakeholders

Chemical Industry A traditional industry that evolves when new technologies on more sustainable processes are developed, but sometimes it does at a slow pace. It benefits from the research done by scientists and is granted with subsidies from governments.

Transports and Logistics Direct impact could potentially be achieved if this sector operated fully on renewable fuels and a more sustainable approach. Multinationals are starting to improve these processes.

Automotive Industry: Especially, the automotive sector of non-electriphicable vehicles. It is important to provide solutions in the short-term such as the usage of renewable fuels in motor vehicles to compensate for the incapability of some individuals to access electricity if electric cars were the norm.

Aviation sector: Boom industry in recent years that could allow for some room for improvement in terms of shifting from fueling planes with chemicals derived from petroleum towards biofuels.

Team building & Inspiration

4C_Fuels was born as an idea to transform the conventional biotech and chemical industry into one that operates based on sustainable development. The creator of this idea was initially Jorge, our Project Coordinator, alongside Álvaro, our instructor and Co-coordinator, and soon enough they came together with the rest of the team. What characterizes us is that we come from different backgrounds: there are biologists, biotechnologists, chemists, chemical engineers, designers and computer scientists which enriches the steps towards the goal of this project.

The diversity and inclusivity all contributing parts have shown towards the project itself and the rest of the colleagues has been crucial for the Human Centered-Design process to be successful on its initial stages. There have been sessions where each of the members would present their ideas on a certain topic and a debate was always generated amongst the others. This allowed us to step out of our comfort zones and think outside our own box with an open mind as we were not sure which direction the project was desired to take

But we were not alone on this road, we began a knowledge-seeking process as a decision on the topic was needed as soon as possible. In order to reach that point, we began to make contacts in the industry… We started contacting biotech companies based in Madrid, where the majority of the team resides nowadays, and in Spain in the interest of collecting as much information from the local community as possible to get a more realistic view of the situation at a regional level from the beginning. Many phone calls and unread emails later,at the end of october 2020, the Association of Biotechnologists of Madrid (AsBioMad) provided us with great support and advice on how to begin with the project itself. Thanks to them, we gained our first visibility on social media and received tips on how to start with our scientific dissemination online which we initiated right afterwards.

Another meeting took place on 20th of november 2020 between Fundación DRO, Compluemprende and AsBioMad in the desire of the latter to provide help to students and young researchers in the biotech industry. Compluemprende is an entrepreneurship unit of Universidad Complutense de Madrid that provides training, support, entrepreneurship programmes, company incubation and contests that aligns with the objectives of Fundación Damián Rodríguez Olivares (DRO). A more concise view of the purpose of the project was provided and next steps evaluated. These meetings, especially with AsBioMad caused an impact on the companies we were able to reach as they served as mediators between us and high ranks from that moment onwards.

In relation to the topic that was taking shape from these conversations, we started to think about microalgae as a potential microorganism in our project. We asked for an opinion on this topic to ALGASOL RENEWABLES via phone call. Our understanding of the situation was that the microalgae sector is focusing primarily on biomass production and the biggest challenge relies on finding cheap cultivation systems. On top of that, we got to know that regulations are pretty strict and there are very limited grants for this type of project in our region. What we gained from this talk was that we wanted to promote this industry and raise awareness about its importance in sustainable development. The company itself inspired us to follow its path, developing photobioreactors and producing High Value Products (HVP) and next generation biofuels. We would like to take the microorganism at the centre of our research and find alternatives that allow us to cut costs and resources by choosing the correct strain.

Along the road of this project, we have maintained contact with AsBioMad throughout its entirety, they have always served as effective guidance for us and we would usually update them with the progress made.


It was not until the presentation of our first project proposal at Compluemprende’s Hackaton in late november 2020, when the iGEM competition began to be a reality for us. This moment was key to believe in our idea and we began to find real solutions to the societal problem that kick-started us and motivated us to take part in this competition. We successfully defended our idea and obtained the first prize, winning the contest, which allowed us to be granted a scholarship for the initial development of the project. The hackathon served as a door to get to be known for our clear business plan and innovative idea. What was honored the most was a model for photosynthetic production of added value products from microorganisms such as microalgae or cyanobacteria.

We continued investigating our chosen group of microorganisms but wanted to know more about these tiny “bugs” and so in January 2021, we reached Maria Juana Llorens Navarro from the department of Biochemistry and Molecular Biology at Universidad Complutense de Madrid. Specifically, we wanted to know how external factors could affect cyanobacteria. The most frequent factors that have a considerable impact on microalgae’s growth are that these are slow processes, there can easily occur any contamination in the medium they live in and they are sensitive to purification processes as well. Therefore, our conclusion from this meeting was that the choice of strain to work with would depend on our ultimate aim, but without forgetting to take into consideration these factors. Ideally, for time management purposes and envisioning the months ahead, we would choose a rapid growth strain.

February 2021 was a month full of energy: the team was starting to bond together, there existed a naive idea of the direction our project would follow and we had made contacts with some organizations and institutions… but there was an important question we hadn’t yet found an answer to and it was, what is our main product? We wanted to look for some answers on this topic and gladly found two advisors: BioCar and Exolum.

BioCar is a research group that studies advanced biofuels and bioproducts at CIEMAT. We had the pleasure to talk with some of their members who validated our project idea and forged the idea of the necessity of new commercially viable and efficient tools to adopt by the traditional biorefinery sector. It came to our mind that phototrophs could play an important role in this change. During the session, a collaborative effort was made to decide the product our project was going to focus on and a holistic evaluation of several biofuels was considered. We confirmed with them our desire to focus on n-butanol due to its numerous advantages.

On the other hand, Exolum, the entrepreneurship spin-off of CLH: a large liquid fuels company, agreed to have a meeting with us. Felix Gómez Cuenca enlightened us with the great potential our idea had but advised us in the complications n-butanol faces as a liquid fuel. A vivid discussion took place around this topic, concluding that n-butanol was a feasible fuel for light vehicles and trucks, but may need some upgrading steps for its use in heavy transportation such as ships or planes due to its low energetic density. We were left to investigate other options but finally decided to direct our project towards the production of n-butanol because it plays a part in relatively simple and sustainable processes which can be improved further and it serves as feedstock for multiple chemical processes and synthesis but is only obtained from petroleum up to now.

The result of gathering information from experts in the field and after some brainstorming done regularly with the team, we decided to offer industrial solutions to fight for climate change. These solutions include providing the necessary infrastructure and genetic tools for the production of high value chemicals and biofuels, focusing on n-butanol, for the decarbonization of intensive and traditional processes. In this sense, it will account for an increase in the share of green compounds towards the overall production of those affecting the biotechnology and chemical industry, the aviation sector, transportation and logistics, amongst others industries. Ultimately, we seek to implement a model of circular bioeconomy.

Experimentation & Iteration

After realizing the challenge we wanted to work with, identifying the relevant stakeholders and gathering all the required knowledge for shaping the project , we were ready to start working in the iteration phase. At this time we focused on the judging rubrics of iGEM’s competition and decided to frame our project in the manufacturing track. Our project at 4C_Fuels has five main lines of action: synthetic biology, encapsulation, design, social media and bioprocess engineering. Each of them was assigned a coordinator which is a leader in their area and organizes next steps towards the completion of main goals.

During this stage, continuous conversations with experts and professionals took place in order to shape and reshape our project repeatedly. Great ideas need action but without the proper means, some have to be modified in order to meet necessities and most importantly seek specific results. We were also curious about how other iGEM teams were functioning. To do so, our team both organized and joined several meetups in order to be able to receive constructive feedback and provide it to others, creating internal feedback loops beneficial for both teams, attending to iGEM’s values and reinforcing a sense of community.

The synthetic biology team focuses on metabolic engineering as well as secretion of our main product: n-butanol. We made use of the installations and labs provided by Universidad Complutense de Madrid, Centro Nacional de Biotecnología (CNB) and Instituto de Ciencia de Materiales de Madrid (ICMM) for the experimentation in this field from summer 2021 onwards. Research was conducted and experts consulted for the election of the desired strain for our lab experimentation. Ideally, a fast-growing strain that could potentially grow with atmospheric CO2 and cheap nutrients under ambient conditions was desired. More information can be found on the phototrophs synthetic biology page regarding this topic.

From the beginning of the experimentation phase, especially regarding the area of synthetic biology and dealing with such unique microorganisms, we found we needed an exclusive community. There was a great need for a phototrophs community and we, along with many other teams helped establish it. It would not have been possible without Marburg and Bielefield teams as they have been key in the organization of events and invitation of attendees and speakers to each session. Very interesting conversations regarding phototrophic microorganisms took place and it was a space to ask for help if any procedure or part was not working as expected. On top of that, we agreed to take part in the Phototrophs Handbook, a guide for the founders of the phototrophs community and for future iGEM teams dealing with phototrophs on parts and metabolic engineering.

Design at 4C_Fuels has primed from the beginning of this project. A branding document was created from scratch and was cautiously followed by each member of the team in any official documents or presentations done. But design for us goes further than that. A prototype of a photobioreactor was built piece by piece by us on a self-taught act. Its purpose was to be used when the cyanobacteria were on point for the production of n-butanol, to be grown and thus gather real data from our operating technology. Another option suggested by the department of chemical engineering of Universidad Complutense de Madrid was to implement our PBR into a setup owned by the university when the process was optimized and proven its feasibility.

Regarding the bioprocess engineering section, various meetings took place with different groups at the faculty of Chemistry of Universidad Complutense de Madrid. We got to finally meet virtually with Victoria Eugenia Santos Mazorra and Miguel Ladero Galán who with their expertise in biorefinery could perfectly understand and express their views on the escalation of units and optimization of growth and operation conditions. Their suggestion lied The second meeting allowed us to meet experts in butanol purification Vicente Ismael Águeda and Marcos Larriba where we dealt with the available methodologies and systems for the purification of n-butanol. They instructed us about the requirements the industry demands and the limitations due to very low liquid concentrations of the n-butanol produced, which has been crucial for our development of the page Implementation and Proof of concept .

As part of the great social media campaign we launched from the early beginnings of this project, we have been active and present in several platforms: Instagram, Twitter and Linkedin mainly. A considerable percentage of the efforts made to attain goals set in the diverse ideation meetings within the 4C_Fuels team was dedicated to nurturing our socials moved by the desire to promote scientific communication and public engagement with our project and the synbio and biotech world in general. For this purpose, graphic design of Instagram posts and divulgation articles on topics that had some relation to our main line of research were published. On top of that, we attempted to create a series of podcasts that broadened the content created in other platforms in order to promote our own visibility from the different platforms and in different artistic disciplines. We intended to create discussions and conversations with our followers and the network in general, by expressing our opinion on some controversial topics with the aim to convince the general population of scientific facts rather than gibberish. In this field, Promega Iberica, one of our partners and sponsors, had an enormous impact on the way we distributed tasks within the team, created and generated interesting pieces of information to share and how we spaced the uploads to create even more engagement with our audience. As a result of their advice, we created our Landing Page, in order to keep valuable and interesting information at hand for anyone that wanted to access it prior to the creation of this wiki and secondly, with the intention of attracting potential investment. More information about this can be found in the Scientific Communication & Education page .

Through previous meetings and interviews done that can be seen on our social media channels or at the 4C_FUels webpage (explained in further detail at the Scientific Communication & Education page ), we have understood that there exists a responsibility in us in divulging science for all. We believe that the general population needs to understand first and then create a sense of necessity of change towards a more sustainable society. Only with this transformation that the population will ultimately demand, an additional cash flow by public and private investors will be created and projects like ours will become a reality in the near future. We were lucky to receive training on this concept thanks to Manu from Bioemprender a few months ago. A future business plan was ideated

In a parallel scenario, we got involved in events, lectures or guides for the promotion of education in synthetic biology mainly. In june 2021, our team assembled a 3D-printed genetic model inspired by the Marburg Collection of Parts that was shared with us at the very beginning of this project. With this tool and promoting the completion of a quick quiz by students, we could see what was the impact of scientific dissemination on young generations. We could gather data from the students through a short question and multiple-choice survey and the results obtained from it are depicted in the section of Education & Scientific Communication. During the summer break, a group of campers came to our labs and we showed them our work and installations, organized by our PI. You can read more about this activities in the Scientific Communication & Education page .

Interview with Algaenergy

Also, during late spring there was a match between Bioemprender and us, so we organized a new meeting with Mónica Gutierrez Salazar (relación FEBiotec) and them, who gave us an insight on the viability of our project from the perspective of the biotech industry. What surprises us the most is when they were the ones who promoted a future call with AlgaEnergy.

The 15th of June we met with María Segura from ALGAENERGY, the biggest microalgae company in our region. First we discussed with María the general situation of microalgae indsutry in our region.

Microalgae industry is a very active sector in our region, but the high fixed costs (CAPEX) are the main obstacle to large-scale development. On the other hand, the lack of generalized knowledge of the technology by society increases the risk of investing in it. This is also the main difference with respect to other technologies such as classical renewables.

According to Maria, the main advantage of this sector is the possibility of cultivation on land with no agricultural value, the high capacity to fix solar energy and the use of atmospheric CO2 as well as CO2 from other industries.

On the other hand, the biggest challenges are the operating costs, derived from biomass harvesting. At the national level, the viability of these companies depends very much on the typology of each company and their business models.

We also took the opportunity to ask about what was initially their main field of action: biofuel production. The main conclusions obtained were that from an economic point of view the production of biofuels derived from the use of microalgae or cyanobacteria presents important viability problems. The main problem is the operating costs (OPEX). This is mainly due to the fact that the approximate cost of production is made up of 60-70% for downstream operations and product conditioning, and only 20-30% for cultivation at both the inoculum and industrial scale.

María confirmed how, after several research and development projects, they reoriented the business model towards other applications with direct market outlets such as fertilizers, nutrition or cosmetics.

Secondly, we take this interview as an opportunity to discuss about more technical issues related to the cultivation of phototrophic microorganisms. Obtaining valuable information on current industrial practices, the reaction systems and cultivation methodologies used and the main challenges faced by this industry.

After the great ammount of information received, we have thoughtfully considered all of it to shape or technological design, specially at the bioprocess scale. ALl of the achieved conclusions are deeply detailed within the Implementation and Proof of concept pages.

Results & Implementation

After thoughtfully designing all the project contents and workplan, our team started working in all of its aspects. The results of all this work are furtherly detailed in the Results page, as well as in ever page of this wiki. However, a brief summary is explained below

Human Practices & Social Aspects

As we have deeply explained within our sustainability page . 4C_Fuels was born to achieve a more sustaibable society, and this could only be achieved considering not only the technoeconomic aspect of sustainability, but also the social one.

With this idea in mind, we have developed various contents with the aim to promote social awareness about sustainability and raise the interest of general society for the potential of science and technology for solving our most pressing challenges. However, we also aimes to transmit the idea that technology alone is useless, unless people is aware about how its opinion, behavior and ideas are crucial for the definition of how technology is developed, and how it can be applied to transform our world.

In addition, we truly believe in the potential of iGEM to fuel this sustainable transition our world needs. Because of this we have taken an active role within iGEM community (know more about in the Collaborations page ), as well as promoted the iGEM concept within our region. Besides huge collaborative projects, like the participation in the iGEM PHototrophs Community, we have also offered various lectures in our region and created divulgative contents explaining what iGEM is. Our aim with all of these activities is not only to settle the foundations for the emergence of future iGEM teams in our region, but also to raise interest about synthetic biology in the society. We want people to understand how synthetic biology is a powerful tool that goe beyond the GMO debate and can actually offer solutions for many problems nowadays still unsolved.

Wet Lab Results

After trial and error a few times without clear and promising results, it is easy to give up but we did try until the end. The work at the lab has been conducted iteratively, solving small problems till reaching the next one. Even though the results expected were not a reality yet, we have validated part of our theorethical design and documented all the issues we have found. We believe that our theorethical design can still be validated by future iGEM teams, which will take all of our work one step further to reality. Because of this, all the theorethical design has been extensively explained in this wiki, to ease the work of future scientist in the development of sustainable biomanufacturing technologies.

Nevertheless all our wet lab work has yileded some interesting outcomes.

  • Cell Encapsulation.On the one hand we have managed to study multiple procedures for cyanobacteria encapsulation in biohybrid materials. Our aim was the intial screening for the future development of cell-encapsulated photobiocatalyst. As a result of this research we have identified a promising silica-based nanomaterial where cyanobacterial cells can be kept alive for more than 30 days without losing its metabolic ability. All of this work has been extensively document in the encapsulation page .

  • Cyanobacteria Synthetic Biology We believe in the need to expand the available toolbox for cyanobacteria synthetic biology applications. In this aim we have proposed the theoretical design of an efficient Unmarking System for cyanobacteria, as well as created extensive documentation about cyanobacterial metabolic engineering. In addition, we have also documented our experience accessing novel fast-growin strains, which may pose important advantages to speed up the development of cyanobacteria metabolic engineering field. All of this contents can be found within our contribution to the Phototrophs Handbook as well as in thePhototrophs Synthetic Biology and Engineering pages.

  • Part Collection and Cloning design Despite we have been unable to succesfully test our expression constructs within a cyanobacteria. We managed to succesfully assemble and sequence-verify a set of useful regulatory and coding parts, which are furtherly detailed within the Contribution Page . In addition, because of our specific need to express a large number of enzymes we have adapted specific cloning procedures which are compatible with the utilization of the Marburg Collection MoClo Toolbox . All of this strategies are furtherly explained at the Cloning Design page

Dry Lab Results

Besides woring in the wet lab, we have also employed all tools in our hands to adress the needs of our project. In this regard, we have worked in three main ambits.

  • Hardware Inspired by the work of previous iGEM teams we have commited ourselves to the task of optimizing and adapting for our needs a previous open-source photobioreactor system. In this regard, we have worked in the construction of a device which will help us to cultivate our cyanobacteria in tightly controlled conditions, as well as automatically record the relevant growth parameters. All this hardware work lead to the succesful construction of an improved version of the OpenPBR system designed by Humboldt_Berlin 2019 iGEM team . To know more about our findings and improvements you can visit our Hardware page

  • Software Bioinformatics is gaining more and more importance for the development of any kind of work within synthetic and molecular biology. In fact we found ourselves in the need of identifying new integration sites for the novel fast-growing cyanobacterial strain we were planning to transform. To achieve this task we have developed a python script: NSfinder. This software piece is capable of identifying putative neutral integration sites in any prokaryotik genome, requiring only genomic sequence information. Our aim with the development of this tool, was to either fulfill one of our project needs as well as to provide synthetic biology community with an additional resource for easing the utilization of non-conventional strains during their research. To now more about this you can visit our Software page

  • Modelling Eventually we have also used computational tools in order to model and simulate the behavior of our design. First, we have colaborated with MiamiU_OH 2019 iGEM Team to model our metabolic design and study how our synthetic pathways will perform in the cyanobacteria.

    Secondly, we have deveoped an intiial engineering design to model and simulate a complete photosynthetic n-butanol production bioprocess, considering current performance of n-butanol producing engineered cyanobacteria. The results of this simulation validated the techno-economic viability of our proposed technology. Further information about all of this results can be found at the Modelling page and Proof Of Concept Page .