Team:Toulouse INSA-UPS/Safety
Elixio Safety
In the present era marked by the desire to build a bioeconomy, attempts to reduce the dependence of our societies on oil do not concern only fuel but should also concern the development of strategies to design renewable and sustainable molecules and materials for our daily lives. In this context, our project ELIXIO aims to develop an innovative biosynthetic pathway to produce silent flower fragrances, in particular violet fragrance. This project consists in using microorganisms modified by a genomic introduction of heterologous genes. Containment of the resulting GMOs is essential since we cannot determine their impact outside of the lab. Consequences of our modifications has also been thoroughfully scrutinized. We will explain in this part how our project has been thought to be respectful of the legislation and safe for the environment.
Classification of the microorganism
The co-culture developed in the Elixio project is based on two microorganisms: the yeast Saccharomyces cerevisiae and the cyanobacterium Synechococcus elongatus. We have chosen these two microorganisms for their potential in the context of metabolic engineering, but also for the low risk they present for human safety and the environment. Indeed, microorganisms are classified according to the health risks they can present to humans and the environment:
S. cerevisiae and S. elongatus are classified in the security group 1 by the American Type Culture Collection (ATCC) and therefore require a laboratory with a mere security level 1 to be handled. They belong to the white list set up by iGEM for safety.
Parts safety
Our project is based on metabolic engineering of the two microorganisms to produce odorous molecules in order to recreate the native violet flagrance. To produce these molecules, a number of genes had to be cloned in these microorganisms. All these genes (table below) come from plants, and therefore, as explained in the iGEM risk group tools, the molecule produced should prevail in terms of safety over the donor organism.
In accordance with the Hazardous Substances Data Bank (HSDB), we have checked the toxicity of each molecule. As presented in the table below, we have listed the potential hazards of each molecule. For each molecule with a potential hazard, we have searched for the quantity presenting a danger in the litterature.
We have also estimated from the literature the maximal quantity of molecules we could produce in a 1 L culture. Even if the culture desorbs the whole quantity of produced molecules, and even if one person breathes this whole quantity, the amounts are way under the toxicity threshold. Only dihydro-b-ionone is over this threshold, but no toxicity has ever been mentioned for this molecule. We therefore concluded that the amounts of each molecule that would be produced are way under the concentration where they have been tested. The production of those molecules is thereby not harmful for the experimenters.
The violet effluvia from the petri dish was smelled without the addition of dodecane to the medium (the molecules produced in the project are not dangerous for humans as explained above). Indeed, it is considered an irritant and a health hazard for humans by the HSDB. We handled it under the chemical host in compliance with safety rules.
Sustainable process
Everything has been done to ensure a safe handling of our modified microorganisms and to guarantee a strict respect of the iGEM "Do not release policy". However, the project aims to be used in the industry at a larger scale. We have anticipated this perspective and thought of a solution compatible with the actual industrial process. The system found by the team is called pervaporation (see section Design) and consists in a technique of separation of molecules in solution by their capacicity to preferentially cross a selective membrane, evaporate and to be recovered at the downstream side of the process into a final solvent. The selectivity is obtained by the affinity of the membrane material for the component that passes through it. This system allows to recover only the molecules and not the GMOs in the culture medium, thus avoiding accidental release.
Lab Safety
Safety and security of the project are of primary importance for us. First, the use and creation of GMO could always be a risk for the environment, albeit unlikely. A second form of safety equally important to us concerns the experimenters in their workplace. iGEM Toulouse INSA-UPS rigorously follows the French legislation and the recommendations of the National Consultative Bioethics Committee for Health and Life Sciences (CCNE) to accomplish its task in the best conditions.
Work under french legislation
As we work in a French laboratory, we must proceed in compliance with the French
legislation, in particular the
Orders of July 16, 2007 and December 27, 2017 fixing the technical measures of
prevention, in particular
of
containment, to be implemented in the laboratories of research, teaching and
analyses where the workers
are
likely to be exposed to biological pathogenic agents (NOR: MTST0756429A and
MTRT1633568A). Moreover, we
must
respect the decree of March 7, 2008 on the prevention of biogenic risks (Articles
R4421-1 to R4427-5),
present in the French labor code. In addition, as we more specifically handle GMOs,
we had to
follow the order of March 28, 2012 relating to the technical file requested for
contained uses of
genetically modified organisms provided for in articles R. 532-6, R. 532-14 and R.
532-26 of the
environmental code.
In this cope, we filled out application files for authorization to handle GMOs and
our work and safety measures were officially approved by the french Scientific
Committee of the High Council for Biotechnology.
Ministry of Higher Education, Research and Administration website, click here to see the website
Formation
The team was trained to proper laboratory procedures for handling chemicals and machines, and in recognizing and preventing hazards inherent in our work environment. This table shows our main training:
Lab security
Due to the non pathogenicity of our microorganismes and the non toxicity of our molecules of interest, the Elixio project was classified as a class 1 work. Nevertheless, a certain number of dedicated rules apply even in a class 1 laboratory, as illustrated with the following pictures taken in our laboratory.
Work in compliance with CoVID19 restriction
Unfortunately, Covid-19 pandemic was still running and mask and social distancing were the rule in our lab. We also limited the risk of contamination by a full vaccination of the whole team and a regular usage of hydroalcoholic gel for both our hands or equipment. Fortunately, we did not suffer from any contamination during the course of the competition.
