Young PIs in action: an interview with Dr Kyle Lauersen

On our latest “Young PIs” interview series, I had the pleasure the interview Kyle Lauersen who recently got appointed as an Assistant Professor. Even though we have many common connections (and we have published a paper together), we never had the chance to discuss, so I listened to his answers on my questions with great interest. I hope you enjoy the post as much as I did!

Kostas Vavitsas: Where did you start your research and how did you land your current position?

Kyle J. Lauersen: I moved to Saudi Arabia a about a month ago. I left Germany a few days before that to spend some time with my family and friends in Canada before making the exciting transition to KAUST.

I did my undergraduate, teaching degree and Master’s at Queen’s University in Kingston, Canada. I then moved to Germany to do my doctorate at Bielefeld University, under the supervision of Prof. Dr. Olaf Kruse in the Center for Biotechnology (CeBiTec) facilitated by the CLIB Graduate Cluster scholarship program (part of Horizon 2020). By the end of my doctorate, I was able to demonstrate that a specific strategy of spreading an enhancing intron throughout codon optimized transgenes to minimize exon lengths and mimic native regulation machinery could mediate their reliable overexpression from the nuclear genome of the alga Chlamydomonas reinhardtii. This technology enabled us over the last 5 years to demonstrate numerous examples of heterologous gene overexpression and the first examples of concerted metabolic engineering with this eukaryotic algal host, work which was largely done during my Post-Doc time also at the CeBiTec with Prof. Dr. Kruse. See our patchoulol paper.

I was very fortunate that I was able to act in a sense as a junior group leader and had the good fortune of working with (now Drs.) Thomas Baier and Julian Wichmann in an intimate team dynamic with complementing skills and expertise. Together with several bachelors and masters students we could focus on algal biotechnology and produce some solid work over the last 5 years.

How did I get my position? I think I came at the right time as KAUST is expanding towards impact focussed and translational research, so our work using algae as green cell factories really tweaked local interest. I was invited to KAUST to give a presentation on collaborative work with Prof. Salim Al Babili and at the end of the talk several senior faculty members approached me to ask if I would be interested in coming to KAUST. At that point I was applying for other faculty positions, so I accepted an interview and was back within a short time to conduct an intense week-long interview. At the end, I was informed that the Division was interested in hiring me as a Faculty member and, after a very short amount of thinking about it, decided it would be an excellent place to grow an algal biotechnology hub.


Kostas: What are the particular challenges of working with photosynthetic microbes?

Kyle: I always wanted to work with photosynthetic organisms. My undergrad research background is in plant biotechnology, I was engineering trees during my undergrad with Prof. Sharon Regan at Queen’s and engineering grass and ice binding proteins with Prof. VIrginia Walker during my Master’s. Also, I am a green guy, I didn’t want to work, say, with animal models. Compared to these systems (or most heterotrophic microbes), algae are easy to handle, smell better, and illuminated cultivation spaces full of green things (or other colours) are generally nice to look at. 

I think we are reaching a point where we need to be resource efficient. We like eating, we like breathing, being mobile, wearing clothes… all theses things are powered, at their origin, by photosynthesis, which makes working with photosynthetic organisms an incredibly logical pursuit.

The biggest challenge of working with algae is the limited investment in research time compared to heterotrophs. So generally, although we build on our predecessors, we are often doing work which has long been figured out in other hosts. Algae research used to be a small part of bigger plant-oriented labs, and only recently we see more dedicated algae research groups.

Now some needed technology is emerging, mainly overcoming technical challenges of how to grow algae in industrial settings or overcome the space-time yield limitations of phototrophic cultivation. Reactor designs like the Subitec system and those coming from CellDeg with CO2 permeabilization membranes are generating truly impressive yields of biomass from CO2. Aggressive and systematic handling of phototrophic microbes is also improving our ability to engineer novel traits. We can get Chlamydomonas transformants in a very short time (4-5 days) and combined with the MoClo or our (pOptimized) modular vector systems, I think we’re at a golden age of possibilities for engineering improvements in these hosts. Also the amenability to microbial handling with robotic systems will improve our turnover rates, which is a great advantage.

Kostas: Sustainability as a common theme among the researchers we interview. Will it bear fruit?

Kyle: I’m very optimistic that what we’re doing will have a real impact in the future. Photosynthetic biotechnology is one part of the greater spectrum of sustainable technologies that are rapidly needed, advancing and coming to market. One good applied example of this is on-site carbon capture, algae can sequester CO2 in the same facility where it is emitted. And keep in mind that there is no faster way to produce photosynthetic biomass than by algae (in the right cultivation set-ups).

The sustainability argument is of course not limited to photosynthetic organisms. And any applications need to be combined with other technologies to become more efficient. What we’re doing will not ‘save the world’ on its own, but it is one incredibly important piece of the puzzle. So we need to boost translational research within the field and increase communication between stakeholders at all levels. This is something that we are seeing now in many EU consortia, like the Horizon 2020 project Photofuel (among many others), which I had the pleasure to take part in 

Kostas: How can we better do translational research with algae? 

Kyle: There was a lot of hype in the field through the last 20 years that has made investors more skeptical on the commercial value of algal applications. But this means that we need to make our argument stronger and provide real practical examples of where algal systems really shine. For example, Algenuity is working on interesting applications of reduced pigment content Chlorella, a great protein source. I think important goals now are to produce a lot more clean biomass of high quality, and demonstrate engineered algae in high-density cultivation concepts that compete with yeast or prokaryotic counterparts. And we need to engage more with material scientists, process designers, and chemists to tailor our lab-scale practices to be amenable to outdoor, or large scale cultivation.

Kostas: When is the right time to apply for a PI position?

Kyle: I don’t have any real wisdom to offer on the topic. I was lucky, I applied only to 10 other positions, which in itself was an emotional rollercoaster over the course of a year..

The process is a bit of a soul-searching exercise. I am Canadian, and I had obtained permanent residency in Germany, had culturally adapted, and had a very good working environment at the CeBiTec. Whenever I was applying to a position I had to imagine myself living there. And when you move to a new place it is a mental hurdle to readapt. Mobility for scientists is both a blessing and a curse: you get to travel but you have a nomadic lifestyle.

Coming back to the question, when to apply… it depends where you are and the support you have. In my case, although I had a great network around the world and Bielefeld was a fantastic incubator, I couldn’t materialize my connections into a permanent position as my network consisted largely of algae people and their institutes weren’t looking for more. What I should have likely done is go to more generalist conferences or meetings and advertise myself to a broader audience.

Kostas: When did you decide you wanted to become a PI? Have you contemplated other roles?

Kyle: I can tell you exactly when. It was during my research internship the summer before my fourth year BSc honours thesis. I walked into Sharon Regan’s lab, and I saw miniature poplar trees growing in sterile magenta boxes in MS agar medium. This was the first time I had seen anything like this and I was instantly hooked, I knew I wanted to stay in such an environment and become a PI. I tried some business and entrepreneurial projects, but academic research always pulled me back.


Kostas: What is the advice you would give to early career researchers in synthetic biology?

Kyle: You have to be self aware and surround yourself with people that can teach your things and work with you to a shared goal. In your younger years you should explore as many things as possible, later during your PhD you should focus, build a team around you and get alot of output.

Put your emotions aside and work really really hard, be consistent, get up everyday, go to work, and be very vocal. Communicate your expectations and work.

Keep networking. Career progression in science is to a large extent merit based, but it will only take you so far. For example, I don’t have any Science or Nature papers (yet?), but the team in Bielefeld and I have produced some pretty solid research. And don’t forget to develop your soft skills!


Short bio

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Dr. Kyle J. Lauersen is currently an Assistant Professor at King Abdullah University of Science and Technology (KAUST) in Thuwal, Saudi Arabia. His group focuses on Sustainable and Synthetic Biotechnology with their main research focussed on engineering algae to be green cell factories. You can also find him on Twitter, LinkedIn, and ResearchGate.

ODYSSEE: A modular platform for field diagnosis of Tuberculosis

By iGEM Thessaly 2019

 

Can you imagine a world where everyone has unlimited access to healthcare? A world where equal opportunities are guaranteed, despite economic, social or political status, through the collaboration among countries?

 

Well, this is just not wishful thinking. These are some of the goals set by the UN (Sustainable Development Goals) for a better world by 2030. iGEM Thessaly decided to work on contributing to the effort made for the achievement of these goals.

 

iGEM Thessaly is the first team from the Thessaly area of central Greece to participate in the iGEM competition.  We are ten students from different Departments of the University of Thessaly. Our project “OdysSEE” aims for the fight against the communicable disease Tuberculosis (TB), a major threat for populations affected by crises such as refugees.

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Refugees and migrants are entitled to the same universal human rights and fundamental freedoms as all people, which must always be respected, protected, and fulfilled. More than 85% of refugees flee from and stay in countries with a high burden of TB (Kimbourgh et al, 2012). 

 

Despite increases in notifications of TB, progress in closing detection and treatment gaps is slow and large gaps remain. The goals of the World Health Organization’s End TB strategy will not be achieved without new tools to fight TB. 

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For this reason, we are developing “OdysSEE”, a rapid, reliable and safe test for early diagnosis of Tuberculosis that would be applied in refugee camps in Greece, as well as worldwide, wherever is needed. OdysSEE reflects the challenging journey that refugees are going through and our logo contains a migratory bird every piece of which represents a unique part of our project.

 

The test will work on urine samples. Once the Mycobacterium tuberculosis, that causes the disease, dies in a patient’s lung, it releases DNA fragments (cell-free DNA - cfDNA) into the blood as it breaks down. cfDNA’s small size allows for it to cross the kidney barrier and appear in the urine (Fernαndez-Carballo et al., 2018). The biomarker we selected is the IS6110 gene (1355 bp), which is located in the genome of the Mycobacterium Tuberculosis (MTB) and encodes for a putative transposase. IS6110 belongs to the family of insertion sequences (IS) of the IS3 category and is most commonly used for the detection of MTB because it is highly conserved (Thierry et al., 1990, Thabet S. & Souissi N., 2016).

 

The detection workflow contains 4 steps of amplification of the target gene. It begins with isothermal DNA amplification of the MTB DNA fragment, with the incorporation of two universal sequences, at 5’ and 3’ end respectively. An in vitro transcription of the amplicon follows with the combination of these two steps enabling addition and amplification of a universal trigger sequence, which is transcribed to RNA.

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This trigger RNA enables the in vitro translation of a toehold switch, a biosensor that encodes for a b-lactamase. b-lactamase is an enzyme that hydrolizes cephalosporins including nitrocefin, which then turns from yellow to red. The colorimetric readout will enable naked eye detection of the result. 


Tuberculosis detection is just the beginning. We aim to create a universal tool able to identify other communicable diseases as well. The key component to achieve this is the trigger RNA that is designed by the team’s wet lab and added to the reverse primer for the first step amplification. This can be achieved by just changing the primer set, while keeping the overhangs that contain the universal trigger, as well as the following path the same, thus targeting different pathogenic agents.

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The ultimate goal is to supplement conventional diagnostics by providing a modular, universal diagnostic platform for various diseases so that all patients have access to innovative tools and services for rapid diagnosis and care.

Join our journey and stay in touch with us and our project by following us on Facebook, Instagram and Twitter and also by visiting our website. Any feedback for our project is welcomed and can be addressed to us via email at igem.thessaly@gmail.com.

You can support our effort through our crowdfunding platform here.

 iGEM Thessaly’s research project is supported by the research infrastructure Omic-Engine, the State Scholarships Foundation (ΙΚΥ), the Research Committee of the University of Thessaly, Hellenic Petroleum, and ELPEN.

  

References

Kimbrough, W., Saliba, V., Dahab, M., Haskew, C., & Checchi, F. (2012). The burden of tuberculosis in crisis-affected populations: A systematic review. The Lancet Infectious Diseases, 12(12), 950–965.

D Thierry, A Brisson-Noël, V Vincent-Lévy-Frébault, S Nguyen, J L Guesdon, B Gicquel, Characterization of a Mycobacterium tuberculosis insertion sequence, IS6110, and its application in diagnosis (1990), Journal of Clinical Microbiology

Thabet, S., & Souissi, N. (2016). Transposition mechanism, molecular characterization and evolution of IS6110, the specific evolutionary marker of Mycobacterium tuberculosis complex. Molecular Biology Reports, 44(1), 25–34.

Fernández-Carballo, B. L., Broger, T., Wyss, R., Banaei, N., & Denkinger, C. M. (2018). Toward the development of a circulating free DNA-Based in vitro diagnostic test for infectious diseases: A review of evidence for tuberculosis. Journal of Clinical Microbiology, 57(4), 1–9.

 

Young PIs in action: an interview with Julie Zedler

A few months ago, I was delighted to hear that my former colleague Julie Zedler secured a tenure-track position in Jena, Germany. I couldn’t miss the chance to interview her for our “Young PI” series; you can see the result of our chat below.

Kostas Vavitsas: The sustainability theme is pretty common among the fresh PIs we interview. Do you think synthetic biology research with photosynthetic organisms can really make a difference in the climate change front?

Julie Zedler: The answer is 100% yes – even though this will almost certainly take longer than we would like. Personally, I believe that photosynthetic organisms are an often neglected player in climate change discussions that we need to raise awareness for. I think the fact that there is a large number of young PIs working with sustainability in mind is very encouraging. However, in order to address big socioeconomic challenges like climate change, we need a critical mass of people and funding bodies to be onboard with the idea. Young scientists interested in making a difference is the first step to finding sustainable solutions but the support infrastructure also needs to be in place to maintain our forward momentum.

Kostas: What is the biggest advantage and the biggest disadvantage of working with photosynthetic microbes?

Julie: I would say the biggest disadvantage is at the same time one of the biggest opportunities: the field of photosynthetic microbes, especially when it comes to applications, is still very young. This makes it sometimes difficult to convince industrial partners and funding agencies to put their weight behind photosynthetic systems. At the same time, photosynthetic microbes have a huge, largely untapped potential. This is super exciting and probably also one of their biggest advantages. Another big advantage is that they are extremely visual and their benefits are intuitive – when talking to a non-scientific audience they almost speak for themselves.

Kostas: When is the right time to apply for a PI position?

Julie: I don’t believe that there is a “right time” to apply per se – often these things come down to the timing of available positions or funding application deadlines and you always need that little bit of luck (combined with hard work of course). I would recommend keeping your eyes out for funding calls and open positions and remember it is almost never too early to apply but it could easily be too late…

Kostas:  How do you feel about your new role? Where do you see yourself in the next few years?

Julie:I am very excited about my new role and I am very curious about what the future holds. Being a young PI, I find it quite important to be an appropriate role model for my students. I am currently building my group and kicking-off work on a number of ideas and research interests that I would not be able to cover on my own. I still do enjoy the occasional bench work though and I promised myself I would stay connected to the practical aspects of research for as long as possible. Looking forward, I think we need to make an increasing effort to communicate what synthetic biology is and how it can impact our everyday lives – it’s crucial to get the public to tag along with scientific progress and explain how and why we think this is the way forward.

Kostas: A comment on equity in science, do you think women have a longer or more challenging road to professorship?

Julie: Personally, as a female scientist, I have never felt disadvantaged. However, numbers speak for themselves and women are still clearly not equally represented (especially when we get to the PI stage), therefore, I am an active supporter of women in science. In all three countries I have worked in, UK, Denmark and Germany, there are efforts to ensure equal opportunities at universities. However, in this respect, Scandinavian countries might be a little bit ahead of the rest of Europe. Overall, I think the biggest problem, both for women and men, lies in the incompatibility between academic workloads and a healthy private life. We need more support and flexible, tailor-made solutions for individual career choices. I believe that nowadays equality challenges are largely of societal nature – when gender stereotypes are finally overcome this will also be reflected in the workplace.

Kostas:  What is the single most important piece of advice you would give to an Early Career Researcher in synthetic biology?

Julie: My students would probably say that I always tell them to do their controls – which I do say over and over again. Specifically concerning SynBio, I think we have to remember that this is a quite young, emerging field. There are high expectations but it is up to all of us to put the work in to make the field thrive. So grab your opportunities and make the most of them!

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Julie is a Junior Professor for Synthetic Biology of Photosynthetic Organisms at the Friedrich Schiller University in Jena, Germany. After her studies at the University of Konstanz (Germany), she pursued a PhD with the Colin Robinson group at the University of Kent in Canterbury (UK). As part of the Marie-Curie International Training Network PHOTO.COMM, she worked on chloroplast engineering in the green microalga Chlamydomonas reinhardtii. After a one-year postdoc at the University of Kent, she obtained a Marie Skłodowska-Curie individual fellowship to work at the University of Copenhagen (Denmark) in the group of Poul Erik Jensen. Her project “Cynthetica” (funded by the European Union’s Horizon 2020 research and innovation programme) focused on the development of synthetic biology tools for cyanobacteria. She has now obtained a tenure track position in Jena and is building her group around the theme of “green systems” and synthetic biology with a current focus on cyanobacteria. You can find her on ResearchGate and Linkedin or follow her lab on twitter (@SynBioJazz).

Optimization of cell-free biosensors for synthetic biology

By Amir Pandi and Olivier Borkowski

Cell-free synthetic biology recently became a branch of synthetic biology with dedicated research groups and conferences. Cell-free systems present a great potential for synthetic biology, allowing for quick in vitro transcription-translation from circular or linear DNA. The most common cell-free systems nowadays are lysate-based cell-free systems which are made by combining cell extract plus reaction buffers. These systems were initially used for fundamental discoveries in molecular biology (study of the genetic code and translation process) and later on to produce recombinant protein. In the past few years, cell-free attracted synthetic biologists’ attention as a platform for high-throughput characterization and prototyping of natural and synthetic biological circuitry. As advantages of cell-free systems we can be list: Non-GMO hosts, absence of growth dependent challenges, lower level of noise, less susceptibility to toxicity, simple cloning as genes can be cloned separately or possibility of using linear DNA (PCR product), high adjustability by varying the concentration of DNA parts or buffer elements.

However, there are still obstacles to use cell-free systems in synthetic biology. A major challenge is inefficient repression behavior. Since many bacterial regulatory elements rely on repression (i.e. most of transcription factors building blocks used in synthetic gene circuits), cell-free synthetic biology has been further developed for metabolic engineering applications than gene circuits development.

Composition and functioning of biosensors in transcription-translation cell-free systems .  (a)  A Cell-free biosensor is composed of the cell-free reaction mix (cell lysate and reaction buffers) plus the DNA.  (b)  The addition of the chemical (inducer) produces GFP. In this case, the inducer de-represses the promoter.

Composition and functioning of biosensors in transcription-translation cell-free systems. (a) A Cell-free biosensor is composed of the cell-free reaction mix (cell lysate and reaction buffers) plus the DNA. (b) The addition of the chemical (inducer) produces GFP. In this case, the inducer de-represses the promoter.


In a recent study published in ACS synthetic biology, we explored different optimization strategies to improve repression in a cell-free system. We designed a simple biosensor responding to D-psicose: psiR, a transcription factor (TF) actuates the expression of gfp from ppsiA promoter.

Sampling a wide range of concentrations for both plasmids expressing TF and GFP reporter is crucial. By trying random concentrations, you will likely not be able to see any GFP production and give up on the experiment. Initially, we only measured a very weak signal with the maximum concentration of the TF and low concentration of reporter DNA. At its best, our first experiment, based on variation of the 2 plasmid concentrations, led to an inefficient cell-free biosensor (very low fold change in the signal).

Optimization strategies applied to improve the fold change of a cell-free biosensor functioning through a transcriptional repressor. (a)  Doping,  (b)  Preincubation, and  (c)  reinitiation of (two-step) reaction. Adapted from  Pandi et al. 2019,  ACS synthetic biology  .

Optimization strategies applied to improve the fold change of a cell-free biosensor functioning through a transcriptional repressor. (a) Doping, (b) Preincubation, and (c) reinitiation of (two-step) reaction. Adapted from Pandi et al. 2019, ACS synthetic biology.

Then, we applied three strategies to overcome the issue of our low fold change.

The first strategy is using a TF-doped extract: the lysate is prepared from cells harboring a constitutive TF-expressing vector so the lysate already contains TF proteins. The cell-free reaction starts with the TF ready to repress its cognate promoter in the absence of inducer. Adding the inducer derepresses the promoter and produces GFP.

The second strategy is using preincubation: first, the cell-free reaction is performed only with the TF plasmid to produce the TF protein (preincubation). Then the reporter plasmid and the inducer are added to the mix before the reaction runs out resources to produce protein. The biosensor efficiency depends on the preincubation time modifying the balance between the amount of expressed TF (increases over time) and the available resources for GFP production (decreases over time). After 8 hours of preincubation, the repression of the promoter is at its highest level but there are not enough resources left for GFP production. Gene expression in cell-free drastically diminishes after 8-10 hours.

The third strategy is using the reinitiation of the cell-free reaction (two-step reaction): first, we preincubated the TF for 8 hours. Then we added the reporter plasmid plus fresh cell-free mix (lysate plus buffers) to reinitiate the cell-free reaction. We saw an improvement in the biosensor efficiency when either 15 or 30 µl were added with the reporter DNA.

Eventually, we compared the unoptimized biosensor as well as two different optimized biosensors to monitor the enzymatic production of D-psicose from fructose. With the optimized biosensors, we were able to quantify D-psicose production. The same preincubation or reinitiation approaches can be used to monitor the prototyping of multi-enzyme pathways in a faster and more efficient design-build-test cycle. Our strategies can be applied to optimize cell-free biosensors and gene circuits that mostly function through repressors and so generalize the use of cell-free systems in synthetic biology. 

Short bios

Amir Pandi:

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I am a PhD student in synthetic biology at Micalis institute, INRA, University of Paris-Saclay. With a bachelor's in the cell and molecular biology from the University of Tehran and a master of systems and synthetic biology from Paris-Saclay, I also participated in iGEM competition as a member (2016), as an advisor (2017), and a mentor (2019). In my PhD, I have been working on the development of biosensors and analog metabolic circuits in whole-cell and cell-free systems.

 

Olivier Borkowski:

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I am a Research Associate at Genoscope, located in Paris area. My research focus is on the relationship between protein production and host physiology. I work both with living cells and cell-free to understand the mechanisms behind the optimization of protein production and resource competition. Currently I am using approaches coupling cell-free technology and machine learning to optimize metabolic pathways.

EUSynBioS becomes a registered non-profit organization!

Dear EUSynBioS members,

It has been a great pleasure for EUSynBioS to participate and promote contents of Synthetic Biology for over 3 years now. We have started our journey as a small student organization and today we have over 400 members in all over the Europe.

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To date, as EUSynBioS we have organized admirable events such as London-UK 2016, Madrid-Spain 2017, Toulouse-France 2018 and eventually this year Brno-Czechia 2019 with the participation of stakeholder, representatives of the field and greatest keynotes driving Synthetic Biology not only in Europe but globally. Together with annual meetings and conferences, our website also serves as a meeting platform for the synbio community, with exciting interviews, research and events coverage, and opinion articles on synthetic biology.

With all this experience and uninterrupted support we get from you, we thought it was time to break our shell and create a legally recognized society and continue serving synthetic biology with new missions and responsibilities that are put over our shoulders to raise this field.

Today, as the president of EUSynBioS I am pleased to inform you about a very recent development in EUSynBioS. Since July 28, 2019 we are recognized as a legal entity by the French authorities under the law for associations of July 1, 1901 and decree of August 16, 1901, and entitled as "European Synthetic Biology Society" at the physical address 128 rue la Boétie, 75008, PARIS. Our new name shows our new perspective: embrace every aspect of synthetic biology; and embrace everyone in synthetic biology. With our new legal status and new motto we are going to be in your disposal as always we have been.

What does it mean for you? In the first instance nothing changes. You continue being members of EUSynBioS. With our new legal status though we are able to create more value for our members. Some of these ways are going to be discussed in our Annual General Assembly that is taking place during our Brno symposium. So I urge you to join us to discuss and decide by vote the future of EUSynBioS

Our activities will continue non-stop such as conferences, collecting and distributing up to date news on the field, building up the network of investigators, academics, and industry representatives, and promoting synthetic biology in every related platform.

 As we have done so far, the rest of the Steering Committee members and I will keep serving this community with enthusiasm and pleasure, at the best of our efforts.


Sincerely,

Huseyin Tas

President of EUSynBioS