Safety

Biology is a domain that need many precautions especially when we use GMOs. Indeed, safety is an important factor that must be considered carefully.

I- Project Safety

General

Throughout the whole project, we used Chlamydomonas reinhardtii as host organism for all the laboratory work. It’s a single cell alga that are widely distributed worldwide in soil and freshwater. We used Chlamydomonas reinhardtii D66 : it is is considered non pathogenic for humans and environment. To assess the safety of Chlamydomonas reinhardtii, researchers estimated the genotoxic potential of the dried C. reinhardtii algal biomass was investigated and no evidence of mutagenicity or genotoxic activity was observed. (1) This strain is characterized by normal photosynthetic characteristic, but requires ammonia as a source of nitrogen for growth and it could be a key factor in the future success of algal-based biofuels. Therefore, Chlamydomonas reinhardtii is easy to use in conventional laboratory without any risk.

Goal

The aim of our project is to cultivate Chlamydomonas reinhardtii in marine environment, for avoid competition with arable land. However, many laws are governing the use of modified organisms in natural environments. Indeed, after our interview with Stephane Lemaire, an expert of microalgue, we noticed the difficulty or our project. The escape of our microalgae into the marine environment can lead to horizontal gene transfer between marine microorganisms.

Despite the genetic modifications made to our microalgae, which could lead to its death in case of strong environmental disturbances, it would require a real average of bio-containment. A solution we were then proposed. « In Chlamydomonas reinhardtii, none of the 69 protein‐coding genes in the plastome uses the stop codon UGA, therefore this spare codon can be exploited as a useful synthetic biology tool ». (2). The goal is to integrate these stop codon in the genome, then to introduce a tRNA to associate with a non-natural amino acid.The microalgae will receive this unnatural amino acid when it is confined in the photobioreactor. Transcription and translation will take place normally.

However when the microalgae escapes from the compartment it will have no more access to the non-natural amino acid, that is going to engender the stop the traduction of a protein which would be essential in the survival of chlamydomonas which will die.In the case of horizontal gene transfer, the recipient organism does not associate transfer RNA with the codon of the mRNA, which causes the translation to stop. So the protein will be truncated because the body will read the stop codon as it is.

Further details about our Human practices available on the Human practices pages

human practices

II. Laboratory safety

Laboratories

Every experiment was made in the “ Institut de biologie physico-chimique” of Institut Curie, Sorbonne université, Paris (France). Before any lab work started, our team members were alerted about good laboratory practices (GLP).

All components used in our experiments are classified as safe and were used under established protocols and with proper guidance. “Classification and labelling” regulation identify hazardous chemicals and inform users about their hazards through standard symbols and phrases. In the EU, the classification and labelling of hazardous chemicals is governed by Regulation No 1272/2008 on classification, labelling and packaging of substances and mixtures (the 'CLP Regulation'). (3)

Hazard pictograms — **Hazard pictogramms (CLP regulation)**

Safety considering chemicals

Besides microorganisms, we also used various chemical compounds such as Ethidium Bromide to visualize DNA for DNA gel electrophoresis. Ethidium bromide belong to substances suspected of causing genetic defects.

Ethidium Bromide is a sensitive fluorescent dye used to detect nucleic acids in agarose gels. It is a mutagen and probable carcinogen. It is toxic and we always wore gloves when working with ethidium bromide. We wiped the area with a damp cloth after the work with ethidium bromide. Also, while wearing gloves after handling ethidium bromide, we were careful to not touch and thereby contaminate other surfaces.

Personnal safety

The experiment were done with many precautions such as:

We wore lab coat, protectives gloves and clothing

autoclaving or disinfecting

Disinfecting work areas before and after use

labeling with name and date every cultures, chemicals clearly

Long pants and hair tied back

We washed our hands with soap before leaving the lab

Collective safety

The laboratory had common equipment for safety as level 1 pathogen laminar flow cabinet, eye wash stations and showers.

Ethical risks

People may be against the genetic modification of microorganisms

Safety in transport

We didn’t face any problems during the sending of our DNA parts, we sealed out our parcel and filled the custom declaration for shipment.

References

[1] A toxicological evaluation of chlamydomonas reinhardtii, a green algae. Int. J. Toxicol. Murbach Ts et al. 2018.

[2]Codon reassignment to facilitate genetic engineering and biocontainment in the chloroplast of Chlamydomonas reinhardtii. Plant biotechnology journal. Rosanna E. B. Young et al. 2016.

[3] Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008 on classification, labelling and packaging of substances and mixtures, amending and repealing Directives 67/548/EEC and 1999/45/EC, and amending Regulation (EC) No 1907/2006 (Text with EEA relevance)