Safety is a foundational pillar of our iGEM project. From initial planning to final execution, we have implemented comprehensive biosafety and biosecurity strategies. According to the new iGEM 2025 regulations, all experimental materials should strictly follow the "white list" specification (only officially certified low-risk biological components and microorganisms are used), and the experimental operations of the high school group should be completed in laboratories that meet the Biological Safety Level 1 standard or lower risk. These practices ensure the safety of our team, the surrounding environment, and any external stakeholders. Furthermore, our framework aims to provide replicable safety protocols for future iGEM teams.

Experimental Materials

All the experimental materials we have selected comply with the iGEM safety policies, as follows:

Host Microorganisms: The experiment utilizes standard non-pathogenic Escherichia coli (E. coli) laboratory strains (such as DH5α or BL21), which are classified as BSL-1 microorganisms. According to the iGEM white list regulations, these strains remain low in pathogenicity even after genetic engineering modifications and carry no risk of antibiotic resistance marker dissemination [1]. Research indicates that commonly used engineered strains such as E. coli BL21(DE3) can be safely employed under strict protocols.

Genetic Components: The experiment involves standard synthetic biology components—such as promoters, RBS (ribosome binding sites), and terminators—all of which are sourced from the iGEM official Parts Registry whitelist and contain no toxic or allergenic coding sequences [2].

Major Potential Risks

In addition to adhering to the contents mentioned in the safety policy, we have also identified some potential security risks that may exist. We are willing to remind everyone that we need to be extremely cautious when conducting experiments, as follows:

Biological Safety Risks: Improper handling of E. coli cultures (e.g., non-vertical pipetting or prolonged opening of culture dishes) may lead to aerosol dispersion; skin contact could cause minor localized infections (though BSL-1 strains generally do not result in severe illness).

Chemical Reagent Risks: The experiment involves the use of LB medium (routine nutritional components), Antibiotics (such as ampicillin, used solely for selection and without human exposure pathways), Ethanol (for disinfection), and organic solvents (e.g., Acetophenone, if applicable, which must be strictly limited in quantity).

Equipment Operation Risks: Improper balancing of the centrifuge may cause mechanical injury; mishandling of the autoclave may result in scalding or pressure leakage.

Personal Protection

All participants had completed the laboratory safety training courses and passed the knowledge assessment before entering the laboratory, including emergency treatment procedures, chemical operation specifications, and personal protective equipment (PPE) standards. The basic safety requirements were as follows: wearing a laboratory coat, disposable gloves, goggles, and toe-covered shoes during the whole experiment. Hair was also tied up, no dangling jewelry was worn, and countertops were sterilized before and after each experiment. In addition, protective equipment can be selected according to the specific operation - an ultraviolet protection mask should be worn when using UV lamps, and an activated carbon mask should be worn when touching organic solvents. At the same time, the instructor assisted the team members in operating the experiment safely (Figure 1).

Figure 1.The instructor is instructing the members to operate the centrifuge safely and correctly.

Laboratory Safety

The experimental place is a safe laboratory that meets the Biological Safety Level 1 and is equipped with fire-fighting facilities (fire extinguisher, sprinkler device), an emergency eye-washing device, and a special recycling bucket for biological waste; Obvious warning signs and functional zoning marks were set up in the experimental area [3]. At the same time, all equipment (centrifuge, constant temperature incubator, sterilization pot) should be completed by professional personnel (laboratory manager or instructor) before use to ensure trough-free operation, in which the centrifuge rotor should be regularly calibrated, sterilization pot pressure gauge should be regularly checked; In addition, safety warnings were clearly marked on all equipment, standard operating procedures (Sops) were provided for high-risk tools, emergency stopping mechanisms and power cut off devices were emphasized in the initial training, students were required to wear basic protective equipment (lab coats, gloves, toe-covered shoes, and goggles) throughout the experiment, and safety signs were prominently posted in all work areas to strengthen risk warnings.

Figure 2. Safe and clean environment in the laboratory.

Equipment Safety

To be able to use the equipment safely, we also developed some safe operation methods for the safe use of critical equipment：

Waste Management

To safely handle all the waste generated during our experiments, we have formulated relevant strategies by category to ensure that our waste is safely disposed：

Biological Waste： Culture media, plates, and centrifuge tubes containing viable bacteria must be sterilized at 121℃ for 30 minutes via autoclaving before being disposed of as "general biological waste" (prohibited from direct drainage). Contaminated items such as gloves, masks, and lab coats should be placed in dedicated yellow biohazard bags, sealed securely, and processed uniformly by the school laboratory.

​Chemical Waste： Low-concentration organic solvents (e.g., ethyl acetate waste liquid) must be collected in labeled, airtight containers and handed over to professional institutions for disposal [4]. Antibiotic solutions (e.g., ampicillin waste liquid) should be diluted and sterilized prior to discharge to avoid environmental contamination.

​Sharps Management： Inoculation loops, disposable pipette tips, and other sharp objects must be deposited in puncture-resistant sharps containers. Under no circumstances should sharps be discarded randomly to prevent injuries.

​Emergency Response Procedures

During the experiment, many unexpected situations may arise. To ensure the safe conduct of the experiment, we have anticipated possible unexpected circumstances in advance and formulated solutions to address them：​

Skin Contact: In case of accidental contact with E. coli cultures, immediately disinfect the affected area with 75% ethanol and rinse thoroughly with running water for 15 minutes. If redness, swelling, or pain persists, seek medical attention promptly and report the incident to the supervising teacher.

Eye Exposure: If solution splashes into the eyes, flush with an eyewash station for at least 15 minutes without rubbing. Avoid touching the eyes and seek medical help immediately.

Aerosol Exposure: In the event of broken culture plates or leaking centrifuge tubes during operations, stop the experiment immediately, shut down the biosafety cabinet/supernatant hood, and cover the contaminated area with a 0.5% sodium hypochlorite solution for 30 minutes before cleanup. Personnel should wear N95 masks and evacuate to a ventilated area for observation.

​Fire/Equipment Failure: Laboratories are equipped with dry powder or carbon dioxide fire extinguishers. In emergencies, contact the campus safety office via the posted emergency number and evacuate following designated routes.

We particularly focused on the safety of gene expression. In this project, when verifying whether the gene expression is correct or whether our target product can be obtained, we will strictly abide by and guarantee the whole process of microbial confined space operation, leakage risk prevention, and inactivation treatment. Microbial environment control: all microorganisms (including recombinant bacteria) involved in gene expression should be operated in a closed space (such as a biosafety cabinet) or a closed fermentation system (such as a fermenter) to ensure their physical isolation from the external environment. During the fermentation process, the plasmid loss was inhibited by regulating parameters (such as temperature, pH, and dissolved oxygen) to ensure the stability of microbial growth. To prevent the leakage risk, the experimental operation should strictly abide by the aseptic technology to avoid the generation of aerosol; It is forbidden to take recombinant microorganisms out of the laboratory, and all wastes (such as culture media and bacteria) should be sterilized before being discarded to prevent gene spread. When not in use in inactivation treatment, after the end of the experiment, the remaining recombinant microorganisms need to be sterilized by pressure steam, completely inactivated, and treated according to the biological waste standards to ensure that no live bacteria remain. These measures together ensure the safety of the gene expression process.

To promote biosafety awareness beyond the lab, our team organized a biosecurity workshop at our school. Participants received safety handbooks and attended an introductory seminar on synthetic biology risks.

We also submitted a draft framework for biosecurity legislation, suggesting enhanced regulations for genetically modified organisms in educational institutions. We aimed to contribute meaningfully to public discussion on synthetic biology oversight.

All the research was conducted in a closed experimental environment. The experimental results will not be used for the production and sale of food, medicine, or health supplements, nor will they be made available to the public. At this stage, the experiments are only carried out in a strictly enclosed laboratory environment. In future applications, the engineered bacteria will be used only in factory-based fermentation processes. After fermentation, the bacteria will undergo complete sterilization, and only their products will be extracted for downstream applications. These downstream products will be entirely free of any engineered bacteria.

During the project discussion at The Chinese University of Hong Kong, Shenzhen, we happened to coincide with their "Biological Safety Week" (Figure 3). During this period, we systematically learned about the overview of laboratory biological safety, the requirements for using biological safety laboratories and equipment, as well as the safety management regulations for hazardous chemicals. At the same time, we also watched the sharing of biological safety accident cases and precautions. After returning to the team, we organized another discussion, reflecting on how to better implement the concept of biological safety based on our own projects. This experience not only deepened our understanding of laboratory safety but also made us more aware of the importance of standardized operations and risk prevention.

Figure 3. The "Biological Safety Week" of The Chinese University of Hong Kong, Shenzhen.

Through robust design, responsible practices, and proactive outreach, our team has integrated safety into every phase of our iGEM project. We believe our documentation and implementation reflect high standards and a commitment to improving biosafety practices in the iGEM community.

[1] iGEM. iGEM Safety Policies (2025 Edition) [EB/OL]. [2025-09-10]. https://responsibility.igem.org/safety-policies

[2] iGEM Registry. Help: White List [EB/OL]. [2025-09-10]. https://responsibility.igem.org/guidance/white-list

[3] Standardization Administration of China. GB 19489-2008 Laboratories - General requirements for biosafety [S]. Beijing: Standards Press of China, 2008. https://openstd.samr.gov.cn/bzgk/gb/newGbInfo?hcno=EB3B94B543F6E4CD18C044DE6AB64CEC&refer=outter

[4] Public Health Agency of Canada. Canadian Biosafety Standard[M]. 2nd ed. Ottawa: Her Majesty the Queen in Right of Canada, 2015.