Difference between revisions of "Team:Paris Bettencourt/Description"

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<h1 class=project>Project description</h1>
 
<h1 class=project>Project description</h1>
 
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Dry cleaning is a process globally used to clean delicate fabrics that cannot withstand the conventional detergents, physical forces and temperatures inside a washing machine. Despite being useful, dry cleaning can pose a threat to human health and the environment. Throughout the process, different hazardous solvents are used to remove the stains from the fabrics. Among the most widely used chemicals one is perchloroethylene (also called "PERC" by the industry). “PERC” is a volatile organic compound, hence it generates fumes that allow it to spread through the air from the clothes after the dry-cleaning process. Furthermore, “PERC” is listed as "reasonably anticipated to be a human carcinogen" in the Thirteenth Report on Carcinogens published by the National Toxicology Program, due to long-term exposure to perchloroethylene having been linked to different types of cancer. Luckily, the French government has begun the process of banning the use of “PERC”, and promoting eco-friendly alternatives in all establishments close to the inhabited areas. <br>
 
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Here is where our iGEM project starts. We interviewed all dry cleaning facilities in Paris to get a better insight on their needs, and the problems they might be facing. Not surprisingly, they told us that wine stains are one of the most difficult stains to remove from clothes, and thus we decided to have them as our main focus. <br>
 
  
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<br>Wine, in general, is a complex mixture of acids, sugars, minerals, water and many different phenolic metabolites, such as anthocyanins or tannins. These two last compounds play an important role in the red color of wine by being part of wine pigments. Our project is inspired by the ability of some organisms to degrade the pigment molecules from wine. Our main focus is to find and improve these enzymes to take advantage of their potential to degrade, and therefore clean, wine stains. We believe that finding an enzymatic alternative to PERC is of great value for the dry cleaning industry, specially given the changes that this industry is facing in the near future.<br>
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<br>In this project we produced synthetic enzymes to remove red wine stains from fabric. These enzymes are designed to replace perchloroethylene (PERC), a toxic solvent used in dry cleaning that will soon be banned in France.  
  
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<br>To achieve our goal, we divided our project in different subprojects which are intrinsically interconnected. Each subproject is carried out by different students from our iGEM team. We divided the team in the following subprojects: “Assay group”, “Microbes screening group”, “Enzyme testing group”, “Binding domain group” and “Indigo group”, in which the last subproject is a proof of concept of all the others. <br>
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<br>We listened to the people who will use our product, conducting face-to-face interviews with every single dry cleaner in Paris. From this came a plan for a realistic product, a stain fighting enzymatic pretreatment compatible with existing cleaning technologies and workflows.
  
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<br>With these subprojects we hope <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Assay">to design and implement an assay platform</a>  in order to <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Microbiology">identify the different microbes from environmental samples</a> that are able to degrade anthocyanins.<br>
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<br>We isolated microbes capable of degrading red wine from a library of 150 strains taken from vineyard soil samples collected around the world with the help of our fellow iGEMers. The most effective microbes were submitted for whole genome resequencing, then analyzed to produce a short list of candidate stain-fighting enzymes.
  
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<br> We will also follow a different approach by expressing exogenous <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Enzyme">enzymes with potential to degrade these pigments in E. coli, in order to purify them and carry out in vitro assays</a>. <br>
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<br>We modeled enzymatic activity at the fabric surface and determined that activity could be substantially improved if the enzymes had a moderate binding affinity for the fabric itself. This effectively increases the enzyme concentration at the fabric surface and reduces the quantity of enzymes lost through diffusion into the medium.
  
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<br>To improve the activity of the candidate enzymes, we will fuse them  with different <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Binding">Fabric Binding Domains (FBD), which have been developed by our team</a>. <br>
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<br>We identified short amino acid sequences with affinity to cotton, linen, wool, polyester and silk using the method of phage display. The resulting Fabric Binding Domains (FBDs) were quantitiatively characterized using ELISA, to determine peptides with optimal affinity as determined by our model.
  
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<br>All these techniques <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Indigo">will be carried out in parallel</a> by the Indigo group, which will work with different enzymes already described as able to degrade specific pigments. By doing so, we will have a proof of concept for the feasibility  of the whole pipeline of the project. In addition, as a consequence of the experimental processes followed by the Indigo group, we decided to look at  pigment removal through an artistic point of view, in what we called Mission Indigo. With Mission Indigo, we really want to <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Indigo">highlight how synthetic biology, and more generally science, can be fused with art</a>.<br>
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<br>We developed a high-throughput assay to quantify stain removal on real cloth. Laser-cut fabric samples are sealed to 96-well microplates and imaged on flat-bed scanners. We coded custom image analysis software to identify each circular fabric sample and measure the stain intensity.
  
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<br> At the end of this nice experience we expect our project to be a nice contribution to the iGEM community, by having developed and sharing a <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Project/Microbiology">database of all the microbes</a> we will isolate from the <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Collaboration">samples we are receiving from all around the world</a>, and by having submitted several new biobricks in phytobrick format. These phytobricks will consist on the candidates enzymes we will test for degradation of anthocyanins and the library of the binding domains with affinity for different fabrics, such as cotton, wool, silk, nylon or polyester. Additionally, we will also share the results of our <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Practices">human practices project</a> created to produce data to answer the question of how our product can fit in the dry cleaning professionals necessities, and how well are they aware about the potential dangers of PERC and the inevitable end of its use in the following years. <br>
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<br>We constructed new BioBricks, fusions of our most promising fabric binding domains to our favorite wine-degrading enzymes. The resulting proteins were expressed, purified and characterized both in vitro (in solution) and in situ (on real stained fabric).
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<br>Finally, we succeeded in increasing the degradation of red wine stains by XX%. These results were obtained under real world conditions, on store-bought fabrics stained with our favorite red wines. We hope these pages convince you that our project is mature, thoroughly documented and effective.
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<br>This project was selected from a pool of ideas generated in a series of discussions, brainstorming sessions and votes. Our selection process has included constant deducting and adding of new ideas, up until a final voting which resulted in the Frank&Stain project. To achieve the best results on this year's iGEM competition, we have created a <a class=”bold” href="https://2016.igem.org/Team:Paris_Bettencourt/Team">team</a> based on our motivation, cultural variety, interdisciplinarity, creativity and scientific background. Together, with a little help from our advisors: Ariel Lindner, Jake Wintermute, Jason Bland and Nadine Bongaerts, we are going to work hard and rid the world of the nasty (although very French) wine stains! <br>
 
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Sources:<br>
 
Sources:<br>
 
   
 
   

Revision as of 09:24, 7 October 2016


Project description


In this project we produced synthetic enzymes to remove red wine stains from fabric. These enzymes are designed to replace perchloroethylene (PERC), a toxic solvent used in dry cleaning that will soon be banned in France.
We listened to the people who will use our product, conducting face-to-face interviews with every single dry cleaner in Paris. From this came a plan for a realistic product, a stain fighting enzymatic pretreatment compatible with existing cleaning technologies and workflows.
We isolated microbes capable of degrading red wine from a library of 150 strains taken from vineyard soil samples collected around the world with the help of our fellow iGEMers. The most effective microbes were submitted for whole genome resequencing, then analyzed to produce a short list of candidate stain-fighting enzymes.
We modeled enzymatic activity at the fabric surface and determined that activity could be substantially improved if the enzymes had a moderate binding affinity for the fabric itself. This effectively increases the enzyme concentration at the fabric surface and reduces the quantity of enzymes lost through diffusion into the medium.
We identified short amino acid sequences with affinity to cotton, linen, wool, polyester and silk using the method of phage display. The resulting Fabric Binding Domains (FBDs) were quantitiatively characterized using ELISA, to determine peptides with optimal affinity as determined by our model.
We developed a high-throughput assay to quantify stain removal on real cloth. Laser-cut fabric samples are sealed to 96-well microplates and imaged on flat-bed scanners. We coded custom image analysis software to identify each circular fabric sample and measure the stain intensity.
We constructed new BioBricks, fusions of our most promising fabric binding domains to our favorite wine-degrading enzymes. The resulting proteins were expressed, purified and characterized both in vitro (in solution) and in situ (on real stained fabric).
Finally, we succeeded in increasing the degradation of red wine stains by XX%. These results were obtained under real world conditions, on store-bought fabrics stained with our favorite red wines. We hope these pages convince you that our project is mature, thoroughly documented and effective.

Sources:
Perchloroethylene, Pubchem
US National Library of Medecine
Report on Carcinogens, Thirteenth edition
Ministère de l'Environnement, de l'Énergie, et de la Mer

Centre for Research and Interdisciplinarity (CRI)
Faculty of Medicine Cochin Port-Royal, South wing, 2nd floor
Paris Descartes University
24, rue du Faubourg Saint Jacques
75014 Paris, France
+33 1 44 41 25 22/25
igem2016parisbettencourt@gmail.com
2016.igem.org