Project Description
The Problem: Today's bioproduction is expensive, inaccessible and unsustainable.
Let's play a riddle: what is in your birthday cake, the clothes you wear, and the medication you take? The answer is molecules made through bioproduction. Bioproduction is powerful, but it rarely runs smoothly. The production of these molecules is not as easy and reliable as it could be: the productivity is unstable, as metabolic bottlenecks prevent constant outputs. Further intermediates and by-products accumulate because feedback control in cells is limited, especially for complex, multi-step pathways. It is hard to fine-tune expression levels in complex pathways, since even minor imbalances can derail the entire metabolic flux. The consequence is a high metabolic burden for the producing cells, forcing them to use large amounts of energy and resources away from growth and maintenance (Kim et al., 2024, Mao et al., 2024). In practice, this means that the very cells we rely on for production become less efficient over time, which limits yields and, consequently, profits. Further, the hardware required to study and optimize these processes is costly and complicated. This makes it inaccessible to many iGEM teams and smaller research groups. As a result, research is slowed down, datasets remain small and inconsistent, and true high-throughput optimization of bioproduction remains out of reach.
We tackle this at a time, where bioproduction is more relevant than ever. Bioproduction promises a much needed sustainable alternative to a lot of current production methods. On one hand the chemical industry uses a lot of unsustainable practices, such as high water consumption, the contamination of our environment through substances such as lead and enormous CO2 emissions. On the other hand natural resources are more limited than ever, therefore they can not always achieve the high yields necessary to meet worldwide demand.
Our exemplary molecule vanillin is a perfect example for these issues connecting to common production methods. Vanillin is the most used aroma compound globally and a product that almost everyone encounters in their everyday life. Growing and harvesting vanilla beans comes with a huge downside due to it taking up a lot of space. The area harvested in 2022 was 92,066 ha big - this equals to 112,964.4 football fields (FAOSTAT) - or 8.74 times the size of Paris. Additionally there is no sustainable alternative in chemical production, as it is based on fossil hydrocarbons (Ni et al., 2015). The solution for these challenges is bioproduction which allows vanillin production from natural substrates using microbes - this is a sustainable and scalable alternative!
Another great example for the future of bioproduction is kaempferol. Kaempferol is a promising product, as it possesses a great range of health benefits, including cardioprotective, neuroprotective and anti-diabetic effects (Alrumaihi et al., 2024). While it is currently extracted from plants, the yields are very low, requiring lots of biomass and labor (Tartik et al., 2023). Therefore its production could be increased drastically with bioproduction.
Our approach: STREAMlining bioproduction
With STREAM, we aim to solve these problems. We combine the construction of hardware and molecular ratiometric biosensors to stabilise the microbial production of high-value biomolecules.
Wetlab: Ratiometric Biosensor
We introduce the production pathway for the target molecule into the E. coli Marionette strain. To demonstrate our construct, we use two exemplary molecules: kaempferol and vanillin, although these can be replaced by other desired metabolites. To monitor the concentration of the produced molecule, we implement a transcription-based ratiometric biosensor, yielding a fluorescent output. To understand how the biosensor works, refer to the animation below: If the repressor binds the promotor, there is only the baseline fluorescent output (here mCherry). Once the target molecule comes into play, it binds the repressor and thereby activates a second fluorescence - here shown with sfGFP. Based on the ratio of these two fluorescent outputs, we react: If the output is very low, we can intervene by adding inducers to upregulate expression levels of specific enzymatic steps, thereby overcoming bottlenecks in production consistency.
Drylab: Modular, low-cost chemostat
Our modular, sensor-integrated chemostat system enables continuous, automated control of microbial cultures. By maintaining cultures at steady state, the system ensures reproducible growth conditions while allowing fine control over the dilution rate, the key parameter that balances nutrient supply with culture growth. We integrate off-the-shelf sensors to monitor key variables with remote control tools. This allows constant oversight of cultures, early detection of deviations from normal growth patterns, and identification of optimal intervention points, all while keeping the costs minimal. Further, we support the hardware by an easily handable software, which allows direct observation and evaluation of measurements to adjust parameters in real time. The result is a bioreactor platform that supports long-term, reproducible experiments - without the need for expensive infrastructure or extensive manual labor.
By integrating the biosensor with our chemostat, we create a closed-loop monitoring and control strategy. The complementary model calculates optimized inducer concentrations and timepoints. Our approach achieves constant high output and yield of the end product, with no intermediate accumulation and reduced (toxic) by-product accumulation, resulting in reduced metabolic stress, fewer metabolic bottlenecks, and an expanded bacterial lifespan.
Full STREAM ahead - how STREAM helps you to shape the future
The future looks bright and the tools we developed can help you shape it. The chemostat opens perspectives for affordable bioproduction experiments, especially for other iGEM teams and smaller research groups. Explore the potentials our project holds:
Click to expand future directions
Plug and play principle
Our system allows for easy modulation of used sensors and probes, and offers many great opportunities for further adaptation and development. In the long term, this system could even be scaled up to perform large experiments without prohibitive costs.
Control strategy
While we currently employ a Proportional–integral-controller for the pH, future collaborators could focus on implementing a broader strategy that involves the control of further parameters and machine-learning-supported control mechanisms.
Ratiometric Biosensor
Our work serves as a proof of concept that can be adapted to various other pathways. A natural next step is the induction of more than two key enzymatic step, to further increase stability and reduce unwanted complications.
References
Alrumaihi, F., Almatroodi, S. A., Alharbi, H. O. A., Alwanian, W. M., Alharbi, F. A., Almatroudi, A., & Rahmani, A. H. (2024). Pharmacological Potential of Kaempferol, a Flavonoid in the Management of Pathogenesis via Modulation of Inflammation and Other Biological Activities. Molecules, 29(9), 2007. https://doi.org/10.3390/molecules29092007
FAOSTAT. Accessed October 5, 2025. https://www.fao.org/faostat/en/#data/QCL
Kim, D. H., Hwang, H. G., Ye, D., & Jung, G. Y. (2024). Transcriptional and translational flux optimization at the key regulatory node for enhanced production of naringenin using acetate in engineered Escherichia coli. Journal of Industrial Microbiology and Biotechnology, 51, kuae006. https://doi.org/10.1093/jimb/kuae006
Mao, J., Zhang, H., Chen, Y., Wei, L., Liu, J., Nielsen, J., Chen, Y., & Xu, N. (2024). Relieving metabolic burden to improve robustness and bioproduction by industrial microorganisms. Biotechnology Advances, 74, 108401. https://doi.org/10.1016/j.biotechadv.2024.108401
Ni, J., Tao, F., Du, H. et al. Mimicking a natural pathway for de novo biosynthesis: natural vanillin production from accessible carbon sources. Sci Rep 5, 13670 (2015). https://doi.org/10.1038/srep13670
Tartik, M., Liu, J., Mohedano, M. T., Mao, J., & Chen, Y. (2023). Optimizing yeast for high-level production of kaempferol and quercetin. Microbial cell factories, 22(1), 74. https://doi.org/10.1186/s12934-023-02084-4