Team:NUS Singapore/Safety
overview
While engineering organisms to fulfil varying functions is getting simpler and more effective by the day, one major hurdle of using engineered organisms, in particular bacteria, is the need for effective biocontainment. Thus far, many biocontainment strategies have been proposed but little progress has been made to effectively prevent the undesired release of engineered bacteria into the natural environment.
To ensure the safety of our project to the environment, we have designed a plasmid retention system which effectively locks our system in our engineered cells. We adhered to strict lab safety practices throughout the project and actively sought the advice of safety experts such as Prof Prakash Kumar to shape our project.
Safe Project Design
As our project involves the use of toxin-antitoxin system to lengthen the functional lifespan of bacteria, our project has an inherent risk of extending the lifespan of undesirable bacteria if they acquired our designed system. To ensure that the myriad of cell-based applications benefiting from our technology is safe for use, we incorporated a dual plasmid retention system along with our longevity module. The dual plasmid retention system comprises of two opposing toxin and antitoxin systems, and requires both plasmids to be transferred simultaneously. This therefore reduces the chances of successful genetic transfer from our engineered cells to the environment.
Furthermore, this system also allows us to remove the usage of antibiotics to act as a selection pressure on the bacteria, which allows us to design plasmids lacking antibiotic resistance. This will thus prevent bacteria from the external environment acquiring undesired antibiotic resistance from our engineered bacteria.
We are cognizant of the fact that the system we have developed seems inherently dangerous due to the presence of 'toxins'. We have thus carefully selected and characterized our toxin-antitoxin systems to ensure that their effects occur solely in the cell and are unlikely to cause significant harm to the external environment.
To characterize the effect of our systems, we spent significant time designing our experiments to ensure that it would both provide us with important insights as well as ensure our safety while conducting them. To understand how the toxins and antitoxins interact with protein production, we intentionally utilized the commonly used and proven safe reporter genes such as GFP and LuxCDABE cassettes. Importantly, we avoided the use of dangerous chemicals and materials and made lab-safe reagents and strains the priority in all of our experiments.
Safe Lab Work
We have taken significant effort in all our experiments to utilize safe lab practices. During all lab work, PPE is worn and care is taken to ensure that a supervisor with the required safety knowledge is present to keep a lookout for us at work.
As lab equipment can also be a source of danger with untrained personnel, we ensured that all lab personnel undergo lab safety training before using any lab equipment. Our completed, detailed iGEM safety form demonstrates our commitment to safety, and we are pleased to invite you to find it here.
Human Practices on Safety
While we took significant steps in our project design and lab practices to ensure the overall safety of our project, we also sought experts’ opinions of any possible dangers that our project might pose and what measures would be needed to counter them. As such, we decided to speak with members of the Genetic Modification Advisory Committee of Singapore (GMAC), which is the leading body advising the biosafety of engineered organisms.
We first spoke to the Chairman of GMAC, Prof Prakash Kumar. He gave us an overview of the various activities the GMAC conducts, as well as their regulatory and administrative power in asserting safety rulings. He provided us with the understanding that the GMAC is solely a supervisory and advisory committee, with the on-the-ground safety decisions and approvals made by differing bodies, such as the Institutional Laboratory Safety Committee and/or the Institutional Biosafety Committee (IBC). He also gave his opinion on the general ethical and safety concerns of genetically engineered organisms, stipulating that these concerns are largely governing modifications affecting higher level organisms such as mammals. He opined that our modification, being in E. coli, would not pose an ethical risk, although further consideration and approval would be required on the safety of our engineered organisms.
Subsequently, we also sought advice from members of the research subcommittee to understand how we could conduct our research safely and effectively. To this end, we managed to get the advice of Prof Raymond Lin, the Chair of the research subcommittee. From speaking to him, we understood that the IBC, established under the National Institute of Health (NIH) guidelines, reviews research experimentations across multiple institutions in Singapore. Our laboratory, the BioMarkerSpace at E6 is approved by the IBC and is certified for BSC1 and BSC2. Most importantly, we ensured that the experimentation protocols to increase functional lifespan of E. coli strictly comply to the NIH guidelines.
These various interviews helped us understand the safety and regulatory concepts of our project. They enabled us to make better decisions on the experiments to conduct, and helped to shape our project towards safer and easier execution.