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| | + | <img src="https://static.igem.org/mediawiki/2019/d/df/T--HK_SSC--HomePic.jpg" height="235"><img src="https://static.igem.org/mediawiki/2019/6/6b/T--HK_SSC--HomePic2.jpeg" height="235"><img src="https://static.igem.org/mediawiki/2019/1/12/T--HK_SSC--HomeCyano..jpeg" height="235"> |
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| − | [[File:T--HK_SSC--labphotos2.jpg]]
| + | <h2>Expression of dCas9-sgRNA Complex in Microcystis Aeruginosa Resulting in the Repression of its Toxin-producing Gene</h2><br> |
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| − | <div class="column two_thirds_size" > | + | <h3> Abstract: </h3> |
| − | <h1>Project description</h1>
| + | <p>Microcystis aeruginosa is one of the most common cyanobacteria responsible for harmful algal blooms. This cyanobacterium produces microcystin, a hepatotoxin that damages the liver. However, direct lysis of Microcystis aeruginosa may not best for the environment as it holds ecological values of heavy metal sorption and oxygen synthesis. We hope to silence the microcystin biosynthesis cluster(mcy) using a catalytically dead Cas9 (dCas9) enzyme lacking endonuclease activity. When the dCas9 enzyme is co-expressed with a single guide RNA(sgRNA), the dCas9-sgRNA complex specifically binds to the McyB gene and blocks transcript elongation, leading to the repression of the McyB gene without altering the chromosome of the Microcystis. Here we provide the design of a dCas9-sgRNA expression gene in a shuttle vector that can replicate in both E.coli and cyanobacteria. We will also be conducting downstream analysis to see how our dCas9-sgRNA expression plasmid affects the microcystin-production rate and oxygen synthesis rate of Microcystis.</p> |
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| − | </p> | + | |
| − | <h1>Abstract / Overview</h1>
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| | + | <h3>Our Team</h3> |
| | + | <p>We are a team of high school students aged 14-17 from St. Stephen’s College Hong Kong. We are all passionate about synthetic biology. Through synthetic biology, we hope to make a difference to the current world, and to share the joy of biology. |
| | + | Throughout this year, we have fought hard for this project, conducting numerous laboratory sessions and paperwork. We look forward to presenting our project to the world in the iGEM Giant Jamboree. </p> |
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| − | <p> | + | <div class="button_link"> <a href="https://2019.igem.org/Team:HK_SSC/Team"> Our Team Page </a> </div> |
| − | Microcystis aeruginosa is one of the most common cyanobacteria responsible for harmful algal blooms. This cyanobacterium produces microcystin, a hepatotoxin that damages the liver. However, direct lysis of Microcystis aeruginosa may not best for the environment as it holds ecological values of heavy metal sorption and oxygen synthesis. In this project, we hope to silence the microcystin biosynthesis cluster(mcy) using a catalytically dead Cas9 (dCas9) enzyme lacking endonuclease activity. When the dCas9 enzyme is co-expressed with a guide RNA(sgRNA), the dCas9-sgRNA complex specifically binds to the McyB gene and blocks transcript elongation, leading to the repression of the McyB gene without altering the chromosome of the Microcystis. Here we provide the design of a dCas9-sgRNA expression gene in a shuttle vector that can replicate in both E.coli and cyanobacteria. We will also be conducting downstream analysis to see how our dCas9-sgRNA expression plasmid affects the microcystin-production rate and oxygen synthesis rate of Microcystis.
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| − | </p> | + | |
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| − | <h2>Project Inspiration</h2>
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| − | <p>
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| − | Our project is inspired from various aspects, including school lessons, books, news articles hiking trips, and even previous iGEM teams.
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| − | </p>
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| − | <ol>
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| − | <li><h4>School lessons: </h4>
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| − | <h5>Chemistry:</h5>
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| − | <p>
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| − | We’ve learnt that certain species of cyanobacteria is capable of nitrogen fixation. We understand the ecological importance of nitrogen fixation. Microcystis aeruginosa can carry out metal sorption. However, they also produce neurotoxins and hepatotoxins, microcystin. These cyclic heptapeptides can cause lethal consequences.
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| − | </p>
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| − | [[File:T--HK SSC--microcystin-LR.png]]
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| − | </ol>
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| − | <h5>Biology:<h5/>
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| − | We have learnt the cellular structure of cyanobacteria. From teachers and textbooks, we have had a brief understanding in how photosynthesis is carried out in cyanobacteria. More than half of the world’s oxygen is produced via phytoplankton (including cyanobacteria). On the contrary, we are aware that the bloom of cyanobacteria will block sunlight and slow down the photosynthesis rate of other water plants. Certain cyanobacteria (including Microcystis) may even produce toxins that inhibit certain enzymes in animals.
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| − | </p>
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| − | [[File:T--HK_SSC--microcystis.png]]
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| − | <h5>Geography:</h5>
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| − | <p>
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| − | Through studies in soil and water sources, we learnt how excessive use of fertilizers create eutrophic waters, which gives rise to cyanobacterial blooms. Teachers also taught us how global warming raises the temperature, which favours the rapid growth of cyanobacteria (especially Microcystis Aeruginosa).
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| − | </p>
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| − | <ol>
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| − | <li><h4>News Articles</h4>
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| − | Cyanobacterial bloom has always been a global concern. Countries from all around the world, especially countries with agricultural industries. New articles from around the world have shown economic losses and environmental damages caused by algal blooms. Severe cases of blue-green algal blooms resulted in the contamination of fresh water sources. Recreational activities in nearby areas have to be banned. Inspired by these articles, we hope to solve the problem of the accumulation of toxic substances produced by harmful algal blooms.
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| − | </p>
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| − | <p>
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| − | <a href="https://www.bbc.com/news/uk-49344232">United Kingdom</a>
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| − | <a href="https://chinadialogueocean.net/2959-algal-blooms-are-starving-chinas-seas-of-oxygen/">China</a>
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| − | </p>
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| − | <p>
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| − | <a href="https://www.kwtx.com/content/news/Blue-green-algae-lethal-to-dogs-also-can-affect-humans-547547641.html">Texas, US</a>
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| − | </p>
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| − | <p>
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| − | <a href="https://www.iowapublicradio.org/post/toxic-bacteria-blooms-impacting-water-systems-across-iowa-dnr-survey-shows#stream/0">Laowa, US</a>
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| − | </p>
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| − | <p>
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| − | <a href="https://www.accuweather.com/en/weather-news/vibrant-green-algae-overwhelming-lake-erie-visible-from-outer-space/70008969">Lake Erie</a>
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| − | </p>
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| − | <p>
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| − | <a href="https://globalnews.ca/news/5660170/saint-john-river-blue-green-algae/">Canada</a>
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| − | </p>
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| − | <li>
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| − | Trips
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| − | </li>
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| − | <p>
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| − | T-PARK
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| − | </p>
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| − | [[File:T--HK_SSC--GeogFieldTrip.jpeg|500px|thumb|center|T-PARK]
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| − | <p>
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| − | We visited T-PARK, at Tuen Mun Hong Kong, near Deep Bay. This picture is taken facing Shen Zhen. Oyster fisheries are common there. As seen from the photo, certain areas were found to be green. We were unsure whether the green patches are algae or not, and decided to conduct further investigation.
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| − | </p>
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| − | <h4>iv: Hiking</h4>
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| − | [[File:T--HK_SSC--geog3.jpeg|500px|thumb|center|Photo taken at Yuk Kwai Shan, Hong Kong]
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| − | <p>
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| − | Some team members discovered unusual green patches, which were thought to be algae. We were interested in knowing how these organisms impact our daily lives. As we dwell into this topic, we realized that cyanobacteria can bring both benefits and disadvantages to the ecological system. Therefore, we decided to design a project that minimizes the harms that cyanobacteria may bring.
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| − | </p>
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Expression of dCas9-sgRNA Complex in Microcystis Aeruginosa Resulting in the Repression of its Toxin-producing Gene
Abstract:
Microcystis aeruginosa is one of the most common cyanobacteria responsible for harmful algal blooms. This cyanobacterium produces microcystin, a hepatotoxin that damages the liver. However, direct lysis of Microcystis aeruginosa may not best for the environment as it holds ecological values of heavy metal sorption and oxygen synthesis. We hope to silence the microcystin biosynthesis cluster(mcy) using a catalytically dead Cas9 (dCas9) enzyme lacking endonuclease activity. When the dCas9 enzyme is co-expressed with a single guide RNA(sgRNA), the dCas9-sgRNA complex specifically binds to the McyB gene and blocks transcript elongation, leading to the repression of the McyB gene without altering the chromosome of the Microcystis. Here we provide the design of a dCas9-sgRNA expression gene in a shuttle vector that can replicate in both E.coli and cyanobacteria. We will also be conducting downstream analysis to see how our dCas9-sgRNA expression plasmid affects the microcystin-production rate and oxygen synthesis rate of Microcystis.
Our Team
We are a team of high school students aged 14-17 from St. Stephen’s College Hong Kong. We are all passionate about synthetic biology. Through synthetic biology, we hope to make a difference to the current world, and to share the joy of biology.
Throughout this year, we have fought hard for this project, conducting numerous laboratory sessions and paperwork. We look forward to presenting our project to the world in the iGEM Giant Jamboree.
