Lab safety is essential for the well-being of the individuals working in the lab, and the environments that could be impacted by the work conducted in the laboratory. By following the safety guidelines implemented by the NIH, CDC, USDA, Michigan OSHA, iGEM, and Michigan Department of Public Health, we can take all the necessary precautions to ensure a safe working environment within our laboratory. With our work, we intend to create a safe and effective treatment for the P. destructans fungus without causing any environmental or personal safety concerns. To achieve this, the following safety practices have been implemented.
Since releasing modified bacteria into the environment can be hazardous, our team has incorporated an extra level of safety into our project by implementing a kill switch. The addition of a kill switch into our project derives from community concerns about previous projects using modified bacteria in the environment. To address these concerns, our team has created and added the switch to our overall circuit design as extra biosecurity. The kill switch is designed so that if any of the modified bacteria were to escape into the environment, they would die off before they could multiply and spread1,2.
In previous cycles, our team has worked on creating and modifying our kill switch to ensure effectiveness. The goal of our kill switch design is that the bacteria become toxic to themselves if it is not in a high enough concentration of bacteria. This means that if a small amount of bacteria escapes containment, the switch is activated, and they kill themselves off. The current version of the switch we use is the second modification detailed in our 2023 wiki safety page1,2.
Twice a week, members of the wet lab met to practice and perform our skills to reduce inaccuracies that may be caused by poor lab techniques. Some more basic techniques include: pipetting, plating, dilutions, and making gels. After repetition of performing basic laboratory procedures, we moved on to applying what we learned about safety to our project material, Miniprep procedures, running gel electrophoresis, etc. During these sessions, we were also taught how our project will work, and delve into the specifics of what happens at each stage. Some examples of these stages are how GG assembly will work, or why abnormal cell growth appeared on our agar plates. This allows us to be more prepared and understand these issues for the future.
The strain of E. coli we are using is named Top10. It is a non-pathogenic strain of E. coli and is regarded as safe by NIH.
Since P. destructans is a possible pathogenic spore, it can pose safety risks. Having P. destructans in the lab could lead to the risk of a possibly pathogenic spore being introduced into the environment, or members of our team inhaling the spores. However, we are using a safer alternative to growing P. destructans to alleviate these risks. We are able to replicate the same effects by using Top 10 E. coli that produce Destructin-1 as it is the main endoprotease of P. destructans that is used to infect other organisms5.
To analyze results after a 1% agarose gel is run through electrophoresis, dyes are essential, however, some dyes pose hazardous risks while being used. Since ethidium bromide is considered toxic, mutagenic, and carcinogenic, the Alma team has opted to use GelRed when we run gel electrophoresis. This allows us to reduce possible safety hazards since GelRed is considered a non-hazardous dye to observe DNA3,4.
We include a code for antibiotic resistance into the plasmids that our E.coli uptakes to ensure we are growing our intended modifications. Then before we streak plate our cells, we pour the agar containing the target antibiotics into a petri dish that we will streak plate onto the petri dishes. However, this means that we are handling antibiotics in the lab, which can raise safety risks. This means wearing the proper protection while handling solutions, such as gloves, and disinfecting work surfaces once finished is essential. To further reduce the chances of antibiotic resistance, we ensure that all of our materials that contain or could have come into contact with antibiotics are disposed of properly. This means using waste containers, and the proper disposal method for each type of antibiotic used.