Difference between revisions of "Team:Sorbonne U Paris/Safety"

 
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<div class="container"> <article class="col-md-10"><h1> Safety </h1>
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<p> Biology is a domain that require extensive cautions especially when it comes to the use of Genetically Modified Organisms (GMO). For this reason we focused extensively on the safety part of our project. </p></article></div>
  
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<h1> Safety </h1>
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<h2 class="title-h2">I- Project Safety </h2>
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<h4>General</h4>
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<p> Throughout the whole project, we used <i>Chlamydomonas reinhardtii </i> as host organism for all the laboratory work. This single cell algae are widely distributed around the world, in soil and freshwater. We used <i>Chlamydomonas reinhardtii D66</i> : a non-pathogenic strain for humans and the environment.
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To assess the safety of <i>Chlamydomonas reinhardtii</i>, researchers estimated the genotoxic potential of the dried <i>C. reinhardtii</i> algal biomass and found no evidence of mutagenicity or genotoxic activity. (1)
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This strain is characterized by normal photosynthetic properties, but requires ammonia as a source of nitrogen. Intrestingly, this ability could be a key factor in the future success of algal-based biofuels.
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Therefore, <i>Chlamydomonas reinhardtii</i> is easy to use in conventional laboratory without any risk.
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</p>
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<h4>Goal</h4>
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<p>The aim of our project is to cultivate Chlamydomonas reinhardtii in marine environment, to 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 microalguae, we noticed several challenge in our project. For example, the release of our microalgae into the marine environment can lead to horizontal gene transfer between marine microorganisms. </p>
 +
 +
<p>Despite the genetic modifications made to our microalgae, which could lead to its death in case of strong environmental disturbances, our project requires a real average of bio-containment. A solution 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 only when it's confined in the photobioreactor. Transcription and translation should thus take place normally.</p>
 +
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<p> However when the microalgae escapes from the compartment it will have no more access to the non-natural amino acid. This will interrupt the traduction process which would ultimately lead to the death of the algae. Furthermore, in the case of horizontal genes transfer, the recipient organism does not associate transfer RNA with the codon of the mRNA, which causes the translation to stop. So the proteins will be truncated because the ribosomes will read the stop codon as it is.
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</p>
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<p>Further details about our Human practices  available on the Human practices pages </p>
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<div class="row text-center text-xs-center text-sm-enter text-md-center  text-lg-center  ">
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        <div class="col-xs-4 col-sm-4 col-md-12 col-lg-12">
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        <a class="btn-sm" href="https://2018.igem.org/Team:Sorbonne_U_Paris/Human_Practices" role="button" style=" color:white; border-radius: 10px; background-color: #22D66C; text-decoration:none; "><i class="fas fa-hands-helping fa-1x"></i> human practices</a> 
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          </div>  <br> <br>
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<h2 class="title-h2">II. Laboratory safety</h2>
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<h4>Laboratories</h4>
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<p>Every experiments was made in the “Institut de biologie physico-chimique” of Institut Curie, Sorbonne Université, Paris (France). Before any lab work, our team members were train about the good laboratory practices (GLP).</p>
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<p>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 risks through standard symbols and phrases. In the EU, the classification and labelling of hazardous chemicals is governed by Regulation N. 1272/2008 on classification, labelling and packaging of substances and mixtures (the 'CLP Regulation'). (3)
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<figure  style=" width: 50%;">
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<img  src="https://static.igem.org/mediawiki/2018/f/fe/T--Sorbonne_U_Paris--Safety_pictogramm.jpg"alt="Hazard pictograms" style="width: 100%">
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    <figcaption style="padding: 10px;">Hazard pictogramms (CLP regulation) </figcaption>
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<h4>Safety considering chemicals</h4>
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<p>Besides microorganisms, we also used several chemical compounds, one being Ethidium Bromide to visualize DNA in gel electrophoresis. Ethidium bromide belong to substances suspected of causing genetic defects.</p>
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<p>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.
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</p>
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<h4>Personnal safety</h4>
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<p>
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The experiment were done with several precautions such as:
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<ul>
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<li>We always wore lab coat, protectives gloves and clothing</li>
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<li>autoclaving or disinfecting </li>
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<li>Disinfecting work areas before and after use</li>
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<li>labeling with name and date every cultures, chemicals clearly</li>
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<li>Long pants and hair tied back</li>
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<li>We washed our hands with soap before leaving the lab</li>
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</ul>
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</p>
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<h4>Collective safety </h4>
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<p>The laboratory had common equipment for safety as level 1 organism such as laminar flow cabinet, eye wash stations and showers.  </p>
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 +
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<h4>Safety in transport</h4>
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<p>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.  </p>
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<br> <br>
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<div class="container" id="references">
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<h5 class ="text-center">References </h5>
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<ul class="list-unstyled text-left">
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  <li> <i class="fas fa-caret-right" aria-hidden="true"></i> [1] A toxicological evaluation of chlamydomonas reinhardtii, a green algae. Int. J. Toxicol. Murbach Ts et al. 2018.</li>
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  <li><i class="fas fa-caret-right" aria-hidden="true"></i> [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.
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</li>
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  <li> <i class="fas fa-caret-right" aria-hidden="true"></i>  [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)
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Latest revision as of 22:30, 17 October 2018

Safety

Biology is a domain that require extensive cautions especially when it comes to the use of Genetically Modified Organisms (GMO). For this reason we focused extensively on the safety part of our project.

I- Project Safety

General

Throughout the whole project, we used Chlamydomonas reinhardtii as host organism for all the laboratory work. This single cell algae are widely distributed around the world, in soil and freshwater. We used Chlamydomonas reinhardtii D66 : a non-pathogenic strain for humans and the environment. To assess the safety of Chlamydomonas reinhardtii, researchers estimated the genotoxic potential of the dried C. reinhardtii algal biomass and found no evidence of mutagenicity or genotoxic activity. (1) This strain is characterized by normal photosynthetic properties, but requires ammonia as a source of nitrogen. Intrestingly, this ability 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, to 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 microalguae, we noticed several challenge in our project. For example, the release 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, our project requires a real average of bio-containment. A solution 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 only when it's confined in the photobioreactor. Transcription and translation should thus take place normally.

However when the microalgae escapes from the compartment it will have no more access to the non-natural amino acid. This will interrupt the traduction process which would ultimately lead to the death of the algae. Furthermore, in the case of horizontal genes transfer, the recipient organism does not associate transfer RNA with the codon of the mRNA, which causes the translation to stop. So the proteins will be truncated because the ribosomes 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 experiments was made in the “Institut de biologie physico-chimique” of Institut Curie, Sorbonne Université, Paris (France). Before any lab work, our team members were train about the 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 risks through standard symbols and phrases. In the EU, the classification and labelling of hazardous chemicals is governed by Regulation N. 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 several chemical compounds, one being Ethidium Bromide to visualize DNA in 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 several precautions such as:

  • We always 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 organism such as laminar flow cabinet, eye wash stations and showers.

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)


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