Project overview: The team is engineering Saccharomyces cerevisiae (strain W303) to enhance extracellular electron transfer and NADH availability for incorporation of created yeast strains in microbial fuel cells (MFCs). This involves CRISPR–Cas9 gene deletions and heterologous expression of cellobiose dehydrogenase (CDH). Work is conducted in E. coli (NEB® Turbo) and yeast Saccharomyces cerevisiae strain W303 and its derivatives under BSL-1 conditions.
Our project involves several potential safety and risk concerns, which can be grouped into chemical, physical, biological, environmental, and AI-related categories. Chemical risks primarily involve the use of ethanol, which is highly flammable, and Safestain, a DNA staining reagent that could cause harm if ingested or mishandled. Physical risks include the possibility of cuts when using scalpels and exposure to UV lamps, which could result in skin or eye injury. Biological risks arise from handling E. coli strains containing antibiotic resistance genes, which could potentially transfer resistance to other organisms, as well as the genetic modification of Saccharomyces cerevisiae yeast and the theoretical risk of accidental release. Environmental risks are linked to the possibility of modified yeast escaping into the wild or transferring genetic material to environmental organisms. Finally, while our use of AI tools is limited to visualization and writing tasks, we remain cautious to avoid providing any sensitive or personal data to these tools.
Risks were recognized through project design review, institutional biosafety guidance (University of Tartu), and discussions with the building safety officer and supervisors (Dr. Villu Kasari and Prof. Andres Merits). The team followed iGEM’s safety framework and EU biosafety regulations.
To mitigate these risks, all experimental work is conducted in certified BSL-1 laboratories, with access strictly controlled and supervised at all times. Experiments are carried out using biosafety cabinets, and no genetically modified organisms (GMOs) are released outside the lab. Our yeast strains have been engineered with auxotrophy and a genetic kill switch to prevent survival in the environment, and the final device is designed as a sealed “biological battery,” further ensuring containment.
All team members undergo mandatory safety and security training before participating in any lab work. This training covers proper use of personal protective equipment (PPE), biosafety levels, emergency procedures, and chemical and fire safety. Students always work under supervision, and unsupervised access to the laboratory is prohibited. Institutional compliance is maintained at all times, including proper waste treatment and inactivation following the University of Tartu guidelines. Procedural safety measures, such as the use of PPE, accident reporting systems, inventory management, and physical access controls, are rigorously enforced. Any new experimental procedures are reviewed in consultation with safety officers to ensure that additional risks are identified and mitigated.
The final device presents minimal risk to people, as it is fully enclosed and designed to prevent contact with living cells. The engineered yeast cannot survive outside the device due to the combined effects of auxotrophy and the kill switch. Risks to the device or infrastructure mainly involve potential mechanical failure, such as leakage or breakage of the enclosure, or reduced device functionality if the kill switch is triggered prematurely or yeast viability is compromised. To address these possibilities, we conduct rigorous testing and simulations of real-use conditions, ensuring both the structural integrity of the device and the effectiveness of biocontainment measures.