Safe Lab Work

Our time in the lab started with a 1-week bootcamp led by one of our assistants, Emanuele Boni. During this week, we learned how to set up our working space and how to perform our main experiments. We also began writing the corresponding protocols.

Key points of the bootcamp:

• Setting up a contaminated space for the handling of special and dangerous chemicals: For that, we decided on a space in our lab that would be dedicated only to performing experiments involving these chemicals. We labelled the bench and the equipment involved to make sure that contaminated instruments would be handled with gloves only and would be kept in their defined area until the last day of lab, when we would finally get them to decontamination. This was especially useful in the case of agarose gels, for which we used SybR-safe as DNA staining.

Figure 1. Correct handling of SybR-Safe with labeled instruments and gloves.

• Setting up a clean space where to use laptops and write the lab notebook. Setting up a clean workspace for team members who work on software and hardware that should not be in contact with any contaminated lab material.
• Leaving dangerous and flammable chemicals under a chemical hood while they are not being used, to avoid accidental spills or unintended contact with a flame, especially in the case of ethanol, which is a standard disinfection chemical. We chose to store stock bottles under our chemical fume hood, apart from where we kept combustibles such as gas bottles. Storing dangerous chemicals under a chemical hood when not in use.
• Before leaving the lab, the last person was responsible of checking that:
• All devices are turned off (energy saving!)
• All windows are closed
• All gas bottles are correctly closed
• All benches have been disinfected with ethanol
• All samples and chemicals are correctly stored where they belong (shelf, bench, fridge, freezer, incubator, chemical fume hood)
• The lab is locked

During our bootcamp, we followed a lab safety course provided by Marco Zini, safety officer and practicals coordinator at our institution. During this course, we had the opportunity to practise and develop our knowledge on good lab practices and safety, by putting lab related or non-related objects in the context of lab work and by reviewing potential scenarios of accidents.

• No eating or drinking is allowed within the lab.
• Wear gloves when handling dangerous chemicals, do not wear gloves when using a flame.
• How to dispose of contaminated waste at our institution.
• First actions to take in case of an emergency.
• Whom to report accidents to, who to contact in case of a problem with the lab equipment.

Safe Project Design

Our project focuses on targeting an invasive species of hymenopterans, the Asian hornet. As it is the case for most conservation related projects, the final implementation of our idea would require the release of synthetically engineered organisms in a non-contained environment.

Although open release is strictly regulated in most countries and especially forbidden by iGEM rules, safety cannot be separated from a project design and is the easiest to implement when it is thought about from the very beginning. The following section summarise the key challenges that we faced regarding safety:

Dealing with invasive species in synthetic biology: What are the main challenges while designing a safe project?

• GMO regulations and open release in the environment Release of genetically engineered organisms being strictly forbidden by iGEM and closely regulated in our country, our team had to find a way to safely gain more knowledge on the effect of our engineered protein bait on an Asian hornet population. For that, we simulated the spread of our bacteria in a mathematical model. that allows us to plan a more precise implementation without the need for a live experiment.
  If we were given more time, we would also investigate the possibility of implementing a kill-switch or auxotrophy, and we would try to get more insight on how the RNAse deletion in our strain would help us limit its survival in an open environment, thus preventing its uncontrolled spread.
• Animal testing Testing an engineered bacteria in the context of a conservation project may often involve animal use, though most teams probably do not have enough experience with animal care to be able to carry out such experiments. This was the case for our team. Even though we would have been allowed to work on insects (Asian hornets) in our country, because there are no special regulations in Switzerland about experimentation on invertebrates, iGEM’s stricter rules on social hymenopterans encouraged us to rethink our strategy and to focus on animal replacing alternatives. From our first brainstorming sessions, we found an interesting project developed by the Estonia-TUIT 2023 iGEM team. After deciding on our project, it became clear to us that we wanted to base our first testing method on their idea of modelling an RNA interference system in yeast, because it would allow us to test our shRNAs in eukaryotes, while avoiding animal experimentation.
• Our idea: how to be specific? Our whole project is centred around the need to find a way to specifically target an invasive species, without impacting others. We found a potential solution in RNA interference, that is based on gene specificity. This promising method is currently attracting a lot of attention in conservation fields. As a result, we created an app to allow other iGEM teams to find potential shRNA targets that are species specific.
  We also developed a second layer of specificity by adapting and implementing an image recognition software (VespAI) to a trap box. This box only allows Asian hornets to access our RNA interference inducing bait, thus protecting other species from unintended RNA interference effects.

Special Mentions: What We Learned Along the Way

References

• « saferalternatives.pdf ». s. d. Consulté le 30 septembre 2024. https://www.uvic.ca/ohse/assets/docs/haz-waste/saferalternatives.pdf.