Many iGEM teams today choose suicide genes to reach environmental biosafety in their systems. Normal suicide genes may experience gene expression leakage or dysfunction, resulting in overkill or underkill of engineered bacteria. In addition, expressing suicide genes is a waste of energy for engineered strains. The mechanism of suicide gene shutdown causes bacteria to produce poisons and antidotes just to stay alive at any given time. The production process consumes a lot of energy and material processes, which reduces the efficiency of bacterial function.
However, limiting the spread of bacteria through hardware containers may, firstly, increase the complexity and cost of the project, requiring specialised equipment and techniques to implement, which may not be applicable in some resource-limited application scenarios. Secondly, the bacterial filter membranes commonly used in hardware solutions are not 100% reliable, there is a risk of failure, and the risk of genetic leakage still exists if a failure occurs.

We use microbial cements in an innovative way that is not only efficient but also cost effective in limiting the spread of strains without the need to introduce expensive hardware equipment and without putting too much extra burden on the strains. Our team uses microbial cement to prevent genetic leakage.
usage：UreABC was cloned into the pET23b vector.  After verification, the recombinant plasmid was transformed into E. coli Rosetta.  The bacteria were cultured in LB medium supplemented with 100 μg/mL ampicillin for 16 hours at 37°C. 

1. environmentally friendly method that does not require large amounts of energy or resources to run, helping to reduce carbon emissions and resource waste
2. the application of microbial cement provides a powerful security tool for genetic engineering and biotechnology research. By preventing the uncontrolled spread of microbial strains, we can reduce the risk of genetic leakage and ensure the safety and sustainability of research.
3. Flexibility allows different research teams to customise it to meet the requirements of specific experiments and projects.

For other teams in iGEM, the method is helpfulThe low cost of the microbial cement means that more research teams and projects can take up the technology, thus boosting scientific research.
At the same time the microbial cement limits the spread of the strain in a way that is not only efficient but also cost-effective, without the need to introduce expensive hardware equipment and without putting too much extra burden on the strain. This innovative approach is expected to provide future research teams with a powerful tool to find solutions to prevent gene leakage that suit their needs.

Application potential

The need to limit genetic leakage: This technology can be used to prevent genetic leakage. It does so by designing microorganisms or biological systems with the ability to precipitate and sequester them under specific conditions. The core idea of this technology is to direct microorganisms to synthesise or secrete precipitable substances, such as carbonates, and encapsulate the genetic material in these precipitated substances, thus holding it in place and making it less prone to leakage or spread.
Designing new biosafety switches: Microbe-induced carbonate precipitation technology can be combined with biosafety switches. These switches can control the growth and metabolic activity of microorganisms as needed. When needed, the switch can be turned on, allowing the microorganism to perform its intended function. When not needed, the switches can be turned off, triggering carbonate precipitation and sequestering the microorganisms to prevent them from leaking or spreading.
Customised safety controls: The flexibility of this technology allows research teams to tailor safety controls to the needs of the project. Biological systems can be designed according to the specific conditions and risks of the experiment to ensure that genetic leakage is minimised

A vision for the future

In conclusion, our innovative approach represents a great advancement in science and engineering that will not only help push the frontiers of research, but also promises to improve the safety of biotechnology and genetic engineering for the benefit of human society. Through our efforts, in the future, more teams will be able to find their own ways to prevent genetic leakage and promote sustainable development of science and technology!

When selecting a project topic, we firstly assess the safety of the gene chassis and gene fragments used, and then ensure that our project will not cause pollution or damage to humans/animals/plants/environment through the process of prediction and implementation. Only then do we start to implement our projects

1.Selection of non-pathogenic chassis

The choice of E. coli as a chassis microbiology was based on its wide range of non-pathogenic properties. This microorganism is commonly found in natural environments and does not usually pose a hazard to the environment or humans. The wide range of applications for E. coli covers environmental monitoring, ecological research, and biotechnology and pharmaceuticals. Its relatively safe nature makes it an ideal tool to help us better understand ecosystems, monitor water quality, study microbial diversity, and even produce drugs and biochemicals. This choice reflects our commitment to pursuing knowledge and innovation in science and engineering, while ensuring minimal environmental and health risks. The choice of E. coli as a chassis microbiology is therefore practically and ethically justified in a number of areas

2.Safety of genetic engineering

a. Selection of genes that do not harm humans/animals/plants

Considering the protection of biosafety and the environment, we use genes that are not contaminated and do not harm any organisms.
The main genes include:
Urease gene: This gene encodes urease, one of the key enzymes in the MICP process. Urease catalyses the breakdown of urea to produce ammonia and carbon dioxide, which combine with calcium ions to form calcium carbonate.

3.Safety of other product ingredients in end use

Urea breaks down into ammonia and carbon dioxide, providing an alkaline environment for Ca2+ and carbonate to form calcium carbonate precipitate.
Carbon dioxide and calcium carbonate precipitate are not harmful to themselves or the environment.
The amount of ammonia is relatively small and does not spread, so ammonia is relatively safe.
Extracellular polysaccharides: This plant-derived biomolecule does not have a negative impact on the surrounding ecosystem and can also be used to improve the soil

4.Determining and assessing the risk of bioleakage in practical applications

 Since we could not complete a finished product in a short time, we made a hypothesis and evaluation of the risk of bioleakage based on the problems we might encounter at the practical application level. In order to ensure that the microorganisms in the coating do not leak, we considered Extracellular Polymeric Substances (EPS) to increase the adhesion between the soil and the microbial coating. They can provide adhesion of microorganisms to soil particles and promote microbial settlement and colonisation on the soil surface. The cells produce calcium carbonate along with extracellular polysaccharides (EPS). The extracellular polysaccharide has adhesive force and will stick to the engineered bacteria together with bacterial cellulose. And then the microorganism's self-generated calcium carbonate can trap itself in the artefact's tight crevices, ensuring that the genetically engineered microorganism in the coating, and its genes, do not leak out, thus meeting the need for biosafety. The second consideration is that there is a very small probability of "horizontal transfer" of our genes, but even if it does occur, the genes are not harmful to the environment. In particular, our urease is Ni ion dependent, and Ni ions are rarely exist in the environment

We have taken the assumptions that we made before the experiment regarding the practical application and added more specificity to them. Since we do not have a real product, we can prove that our design will not have an impact on the environment through the following aspects. The feasibility of our experiment is discussed in the following process

Before fixing:

When designing the project, we took into account the possibility that the liquid would be too fluid and flow to other places, which could have a negative impact on other parts of the heritage or the environment, so we added bacterial cellulose to the liquid, which enhances the viscosity of the product, so that the liquid does not flow as quickly out of the cracks that need to be filled, and so that the restoration is more efficient.

Fixed：

After Fixed:

We have considered that the liquid may have the probability to flow to other places, which may cause harm to the environment or other places of the artefacts, but according to the verification, it does not cause any other harm to the artefacts. Secondly, we have considered the impact of the harsh environment on the restoration of our artefacts. However, after verification, it was found that the formation time of calcium carbonate is very considerable, and it can form a solid quickly and fix in the cracks. In case of heavy rain, because of the addition of bacterial cellulose, it will not be easily washed away by the rain.
So in conclusion, our experiment will not cause any big problem in the application level.

1.We gave a talk on safety education

Our team held a seminar on lab safety at Hailiang Foreign Language School for students and teachers who have lab exams or who will enter the lab to conduct research. The seminar was aimed at bringing lab safety to the forefront of everyone's mind and reducing the risk of lab hazards

2.Preparation of laboratory manuals

We have compiled a safety manual of accidents that can occur in the lab and how to deal with them, which is distributed to all campuses, also to be able to reduce the number of accidents in the lab

1. Safety training before entering the laboratory

Safety training before entering the laboratory Before entering the laboratory, all members who need to enter the laboratory are trained on laboratory safety by the laboratory staff in charge. Basic operation of the lab equipment, what to do in case of an emergency, and basic preparations for entering the lab were given. These questions were given in the form of questionnaires, and only after all of us had passed these tests did we start to operate in the lab

2.Laboratory Safety Layout Options

3.Safety checks before and after entering and leaving the laboratory

Safety during the experimental process

Reagent placement

In the safety zone of iGEM, our team conducts in-depth research, adhering to the highest safety standards to ensure that our scientific experiments are not only innovative but also safe. We strictly comply with biosafety guidelines and take all necessary measures to prevent and control potential risks. Our goal is to advance cutting-edge research in the field of biological science while ensuring the safety of society and the environment. We are committed to continuing our efforts, maintaining our passion for biotechnology, and always taking human and environmental safety as our responsibility, striving for a better future.