Ensuring safety is the priority of our project and laboratory work. Throughout the entire iGEM cycle, we have strictly followed both iGEM safety policies and national biosafety regulations. Our project does not involve any high-risk organisms, human or animal samples, or activities requiring advanced approval. All experimental work was performed under Biosafety Level 1 or 2(BSL-1, BSL-2) conditions, and every team member underwent comprehensive safety training before starting laboratory operations.
Our project involves engineering strains of Saccharomyces cerevisiae BY4741, Saccharomyces cerevisiae S288C derivant and Saccharomyces cerevisiae CEN.PK 113-7D derivant to produce natural antimicrobial peptide LL-37 under controllable conditions. Saccharomyces cerevisiae is a GRAS(Generally Recognized As Safe) organizm. According to ATCC(Fig. 1), these strains belong to the first category in biosafety, which is almost not pathogenic to humans. We deliberately designed our project to avoid any elements that could increase antimicrobial resistance or pose ecological risks. All parts used are on the IGEM White List.
Fig. 1 Strain information of Saccharomyces cerevisiae from ATCC
Fig. 2 Strain information of E.coli DH5α from ATCC
Fig. 3 Strain information of S. aureus from ATCC
For cloning and bacterial inhibition assays, we employed Escherichia coli DH5α, also classified as RG1 (Fig. 2). In addition, the bacterial inhibition assays invloves Staphylococcus aureus, which is classified as a RG2(Fig. 3). All experiments involving S. aureus were conducted strictly in a certified BSL-2 laboratory(Fig. 4), with enhanced biosafety measures and under the supervision of trained personnel. Plasmids pESC, p426, and pY26 were used as vectors, and all synthetic DNA (provided by Tsingke Biotech) was verified for biosafety compliance. Experiments were conducted exclusively in a BSL-1 laboratory. At the end of each experiment, all cultures and waste were sterilized immediately, and intracellular DNA was degraded to prevent accidental release into the environment.
Fig. 4 the BSL-2 laboratory
Our engineered strains are designed for controlled laboratory use only, with no intention of environmental release.
The 2025 Jiangnan-China's team carries out their experiments in the Co-culture Fermentation Joint Innovation Laboratory,which is a Biosafety Level 1 (BSL-1) laboratory and includes emergency exits, fire-fighting cabinets, and first-aid kits (Fig. 4), and the laboratory is equipped with emergency eyewash and shower equipment. Also, the required signs are posted in the laboratory (Fig. 5). Our laboratory building also contains BSL-2 laboratory for us if there is further experiment.
All our aseptic operations are carried out on the ultra-clean bench. All the experiements waste can be handled properly. The used medium needs to be sterilized and placed in the garbage can for unified treatment. There are special collection and storage places for organic waste liquid, broken glass products and sharps such as syringes, which professional personnel will deal with them in a unified manner and at a regular time.
Fig. 5 Safety instruments
Fig. 6 Safety signs
Before entering the laboratory, all wet lab members completed a safety training program, including chemical hazard awareness, emergency response procedures, and a laboratory skills assessment. During operations, team members wore appropriate personal protective equipment (PPE), including lab coats, gloves, and masks. Work surfaces and equipment were disinfected with approved agents before and after use.
All hazardous waste, including liquid reagents and broken glass, was disposed of following institutional biosafety protocols. Before performing any formal experiments, the laboratory manager trained members in the safe use of essential instruments, including autoclaves, ensuring all procedures were conducted safely and reliably.
Our project involved several hazardous chemicals, including:
• Toxicant: formaldehyde, Super Red(DNA stain)
• Highly flammable chemicals: ethanol, isopropanol, methanol, acetonitrile
• Acids and corrosives: hydrochloric acid (HCl)
• Antibiotics: kanamycin, ampicillin
• Other hazardous reagents: sodium dodecyl sulfate (SDS), TEMED, APS
All chemicals were handled according to their Safety Data Sheets (SDS), using proper PPE and fume hoods when necessary. UV sterilization was employed where appropriate, and antibiotics were used strictly for selection purposes with risk mitigation measures in place.
All experiments adhered to the Biosafety Law of the People’s Republic of China and iGEM’s safety policies. We followed institutional guidelines for handling data, biological materials, and waste management.
In addition, during our human practices activities in primary and secondary schools, we actively promoted laboratory safety awareness. We educated students on the risks of microbial leakage and methods for preventing accidental spread after experiments, contributing to broader biosafety literacy.
