Team:Bilkent UNAM Turkey

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<p><b style=3D'mso-bidi-font-weight:normal'><span lang=3DTR>Biodegradation of TNT and TNT derivatives by <i>nfsI</i>-transfected <i>Chlamydomonas_reinhardtii</i><o:p></o:p></span></b></p>
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Our aim is to modify the unicellular microalga <i>Chlamydomonas reinhardtii</i> for effective trinitrotoluene (TNT) biodegradation by expression of Enterobacter cloacae nitroreductase gene nfsI.  <i>C. reinhardtii</i> is a ubiquitous species capable of thriving in soil, freshwater and marine environments, making this alga a suitable choice for removal of TNT from a wide variety of biomes. We have opted for the use of nfsI as the gene in question is a well-characterized nitroreductase with a known sequence. Our experimental procedure involves (a) development of a synthetic nfsI gene with flanking prefix and suffixes of standard Biobricks, (b) ligation of this sequence to the C. heinhardtii expression vector pRbcBRL, (c) transfection of <i>C. reinhardtii</i> with the recombinant plasmid and (d) TNT tolerance and biodegradation capacity assessment of the modified alga.
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        <li>Bilkent UNAM Turkey team established in January 2011 and member list has grown and changed in time. We shared a lot by that time and are extremely thankful to our former members. We did many brainstorming meetings, some of them was inconclusive. Team went through several problems that cause slightly negative impact on team. For a team to join from Turkey, money always becomes a problem. That’s why we are just able to send just two of our members to the regional jamboree. Moreover, laboratory equipment shipping takes very much time, even a restriction enzyme received one month after the ordering. Our team survive those restrictions and did its best. The force that always give impetus to ourselves is the friendship that we have.
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<h2>SAFETY AND SECURITY QUESTIONS</h2>
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In this summer; we learned about iGEM, how it works, what the biggest part of competition is and how we can win a medallion.<p>
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<b>a. Risks to the safety and health of team members or others in the lab?</b>
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When we were newbie, we organized a lot of meeting to find out our main project than we got idea for improving human welfare so we choose an idea from brainstorm results. We searched to get a gene to work on. As a result; lab works began.
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We added our protocols which are helpful to transform <i>Chlamydomonas reinhardtii</i> and <i>Escherichia coli</i>. Basic cloning protocols are available on our protocol page. Experiments and safety questions also available from link at the top section.
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<br>Using hazardous materials always has risk to cause trouble for team and lab members. We always <br>use hoods and wearing lab coat, gloves, goggles are essential for an experiment.
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<br>We are working on two species one of them is Chlamydomonas reinhardtii which has no dangerous <br>effect on human health. Other one is Bacillus subtilis which also is not human pathogen but it <br>could cause food poisoning.
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<br><i>Chlamydomonas reinhardtii is a strain of algae and it could cause algal bloom that means it <br>overgrown in a water ecosystem and become poisonous for other species which share same places.  
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<br>According to a Toxic Substances Control Act report from the Environmental Protection Agency, <br>Bacillus subtilis “is considered a benign organism as it does not possess traits that cause <br>disease. It is not considered pathogenic or toxigenic to humans, animals, or plants. The potential <br>risk associated with the use of this bacterium in fermentation facilities is low.” Its degree of <br>toxicity is III and IV the lowest toxicity effect on other species.
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<br><i>Our modified Chlamydomonas reinhardtii just degrade TNT and it could not use for terrorism <br>activity. Those two modified organism could not have ability to damage human health. Our facility <br>always has an active security system and it requires an identity card for enterance.
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<i>Natural strain cause algal bloom and threat environmental quality so we used biological <br>hazardous waste for them. And after our work is finished, we applies bleach (%5) and then it goes <br>to waste.</i>
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<br><i>Our parts have not any safety, security and health issues. However, adding NfsI gene into algae <br>could make it more competitive than normal strains. It requires checking before releasing to <br>nature.</i>
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<br><i>Our institution have its own biosafety rules and it takes 19 page so we added a link below which <br>leads to institution’s safety committee page and it contains two link; one of them orientation <br>of our lab and other one is chemical disposal form.
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<br></i><a href="http://unam.bilkent.edu.tr/UNAM%20Laboratory%20Safety.html">http://unam.bilkent.edu.tr/UNAM%20Laboratory%20Safety.html</a>
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<br><i>We have a Laboratory Safety Committee; however, we did not discuss about our project because our <br>experiment contains standard cloning procedure, which has always done in the lab.  
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<br><i>Our institution have an exam about biosafety, we passed it. It consists of orientation which <br>mentioned above.</i>
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IGEM Project is our number one priority. As we worked really hard for our project it is important for us to make our dreams come true which is taking a gold medal. Questioning different items like experiment time, materials and methods and the efficiency of the experiments improved our skills and we started to look as a scientist to problems we faced. There were lots of challenging parts and sometimes we thought to quit but we listened to “Eye of The Tiger” and we did not give up from our experiments we delayed them yes, but we never gave up. <p>
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IGEM team- meet-up was a great opportunity for us to communicate with the other groups, understood their projects and share our ideas. As we had a chance for an interview with Anatolia Agency, we also used media power to get attention to our project, iGEM and the importance of synthetic biology. <p>
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Thanks and get great time.<p>
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<h2>SAFETY AND SECURITY QUESTIONS</h2>
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<a href="http://partsregistry.org/cgi/partsdb/pgroup.cgi?pgroup=iGEM2011&group=Bilkent_UNAM_Turkey"><img src="https://static.igem.org/mediawiki/2011/e/ea/Our_parts.png" width="700"> </img></a>
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<p>
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<h3>1. Would the materials used in your project and/or your final product pose:</h3>
+
<p><b style=3D'mso-bidi-font-weight:normal'><span lang=3DTR>Favorite Parts<p></b>
-
<b>a. Risks to the safety and health of team members or others in the lab?</b>
+
 
-
<i>
+
 
-
<br>Using hazardous materials always has risk to cause trouble for team and lab members. We always <br>use hoods and wearing lab coat, gloves, goggles are essential for an experiment.
+
<p><b style=3D'mso-bidi-font-weight:normal'><span lang=3DTR>BBa_K596001<p></b>
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<br>We are working on two species one of them is Chlamydomonas reinhardtii which has no dangerous <br>effect on human health. Other one is Bacillus subtilis which also is not human pathogen but it <br>could cause food poisoning.
+
 
-
<br></i>
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<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K596001"><img src="https://static.igem.org/mediawiki/2011/f/f3/BBa_K596001.JPG" width="700" height="300"> </img></a>
-
<br><b>b. Risks to the safety and health of the general public if released by design or accident?</b>
+
 
-
<br><i>Chlamydomonas reinhardtii is a strain of algae and it could cause algal bloom that means it <br>overgrown in a water ecosystem and become poisonous for other species which share same places.
+
<p><b style=3D'mso-bidi-font-weight:normal'><span lang=3DTR>BBa_K596004<p></b>
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<br>According to a Toxic Substances Control Act report from the Environmental Protection Agency, <br>Bacillus subtilis “is considered a benign organism as it does not possess traits that cause <br>disease. It is not considered pathogenic or toxigenic to humans, animals, or plants. The potential <br>risk associated with the use of this bacterium in fermentation facilities is low.” Its degree of <br>toxicity is III and IV the lowest toxicity effect on other species.
+
 
-
<br></i>
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<br><b>c. Risks to environmental quality if released by design or accident?</b>
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<a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K596004"><img src="https://static.igem.org/mediawiki/2011/0/05/Algae_protein_expression_vector_pAPEV.png" width="450" height="450"> </img></a>
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<i>
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<br>We haven’t checked modified organism is more competitive than natural strains. However; Bacillus <br>subtilis is well-known and commonly used model organism like Chlamydomonas reinhardtii.
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<p><b style=3D'mso-bidi-font-weight:normal'><span lang=3DTR>DNA Submission to Registry<p></b>
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<br></i>
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-
<br><b>d. Risks to security through malicious misuse by individuals, groups or states?</b>
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<img src="https://static.igem.org/mediawiki/2011/9/9f/DNA_submission_to_registry.JPG" width="700"><p>
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<br><i>Our modified Chlamydomonas reinhardtii just degrade TNT and it could not use for terrorism <br>activity. Those two modified organism could not have ability to damage human health. Our facility <br>always has an active security system and it requires an identity card for enterance.
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<br>Please explain your responses (whether yes or no) to these questions.
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<br><b>Specifically, are any parts or devices in your project associated with (or known to cause):</i></b>
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<br>- pathogenicity, infectivity, or toxicity?
+
 
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<br><i>No</i>
+
 
-
<br>- threats to environmental quality?
+
 
-
<br><i>No</i>
+
 
-
<br>- security concerns?
+
 
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<br><i>No</i>
+
 
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<br><h3>2. If your response to any of the questions above is yes: </h3>
+
 
-
<b>a. Explain how you addressed these issues in project design and while conducting laboratory work.</b><br>
+
 
-
<i>Natural strain cause algal bloom and threat environmental quality so we used biological <br>hazardous waste for them. And after our work is finished, we applies bleach (%5) and then it goes <br>to waste.</i>
+
 
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<br><b>b. Describe and document safety, security, health and/or environmental issues as you submit your <br>parts to the Registry.</b>
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<br><i>Our parts have not any safety, security and health issues. However, adding NfsI gene into algae <br>could make it more competitive than normal strains. It requires checking before releasing to <br>nature.</i>
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<br><h3>3. Under what biosafety provisions will / do you operate? </h3>
+
-
<b>a. Does your institution have its own biosafety rules and if so what are they? Provide a link to <br>them online if possible. </b>
+
-
<br><i>Our institution have its own biosafety rules and it takes 19 page so we added a link below which <br>leads to institution’s safety committee page and it contains two link; one of them orientation <br>of our lab and other one is chemical disposal form.
+
-
<br></i><a href="http://unam.bilkent.edu.tr/UNAM%20Laboratory%20Safety.html">http://unam.bilkent.edu.tr/UNAM%20Laboratory%20Safety.html</a>
+
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<br>
+
-
<br><b>b. Does your institution have an Institutional Biosafety Committee or equivalent group? If yes, <br>have you discussed your project with them? Describe any concerns or changes that were made based <br>on this review. </b>
+
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<br><i>We have a Laboratory Safety Committee; however, we did not discuss about our project because our <br>experiment contains standard cloning procedure, which has always done in the lab.
+
-
<br></i>
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<br><b>c. Will / did you receive any biosafety and/or lab training before beginning your project? If <br>so, describe this training.</b>
+
-
<br><i>Our institution have an exam about biosafety, we passed it. It consists of orientation which <br>mentioned above.</i>
+
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<br><br><b>d. Does your country have national biosafety regulations or guidelines? If so, provide a link to <br>them online if possible.</b>
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<br><a href="http://www.unep.org/biosafety/files/TRNBFrep.pdf">http://www.unep.org/biosafety/files/TRNBFrep.pdf</a>
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</ul>
</ul>
 +
<ul>
 +
<li>
 +
We find it our duty to thank Gulce Itir Percin, Aydan Torun and Ömer Faruk
 +
Sarıoglu for their valuable assistance in conducting the experiments
 +
within our project’s scope, without which we could scarcely have expected
 +
to complete our project in time. We must also extend our thanks to
 +
Sustainable Technologies Laboratory Manager Zeynep E. Ülger for providing
 +
the necessary instructions and training for the use of a wide variety of
 +
laboratory equipment. We would like to acknowledge Prof. Dr. Tayfun Özçelik and Prof. Dr. Engin Umut Akkaya for their sincere support on our team and efforts for getting finanical support from Bilkent University. We thank the Turkish National Nanotechnology
 +
Institute (UNAM) for the administration of routine training sessions regarding
 +
laboratory safety, which allowed all our team members to operate without
 +
any personal danger throughout the experiments so far. We are also
 +
grateful to Assistant Professor Ayse Begum Tekinay and her laboratory for
 +
supplying much-needed reagents and assistance, both of which allowed our
 +
small team to work much faster than it otherwise could. And lastly we must
 +
thank our supervisor Turgay Tekinay for making this project possible and
 +
granting us a summer that was as entertaining as it was informative.
 +
<h1>Sponsors</h1>
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<a href="http://www.bilkent.edu.tr"><img src="http://www.bilkent.edu.tr/~yayinbir/amblem/ing-amblem.jpg" height="100"></img></a>
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<a href="http://www.nano.org.tr/index1.html"><img src="http://www.nano.org.tr/files/Unam_logo.jpg" height="100"></img></a>
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</div>
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<a href="http://www.sentegen.com/"><img src="https://static.igem.org/mediawiki/2011/8/8d/Sentegen_logo.jpg" width="150" height="100"></img></a>
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</div>
+
<h2>Share</h2>
 +
<span  class='st_email_large' ></span><span  class='st_facebook_large' ></span><span  class='st_twitter_large' ></span></span><span  class='st_yahoo_large' ></span><span  class='st_friendfeed_large' ></span><span  class='st_blogger_large' ></span><span  class='st_delicious_large' ></span><span  class='st_linkedin_large' ></span>
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Latest revision as of 03:44, 22 September 2011

 
PROJECT
Background Project_Description
TEAM
Gallery Team_Page
LABWORKS
Protocols Experiment Lab Safety
HUMAN PRACTICE
BBQ Party Team Meetup iGEM Everywhere Collaboration
BIOBRICK PARTS ATTRIBUTIONS

    Biodegradation of TNT and TNT derivatives by nfsI-transfected Chlamydomonas_reinhardtii

    Abstract

    Our aim is to modify the unicellular microalga Chlamydomonas reinhardtii for effective trinitrotoluene (TNT) biodegradation by expression of Enterobacter cloacae nitroreductase gene nfsI. C. reinhardtii is a ubiquitous species capable of thriving in soil, freshwater and marine environments, making this alga a suitable choice for removal of TNT from a wide variety of biomes. We have opted for the use of nfsI as the gene in question is a well-characterized nitroreductase with a known sequence. Our experimental procedure involves (a) development of a synthetic nfsI gene with flanking prefix and suffixes of standard Biobricks, (b) ligation of this sequence to the C. heinhardtii expression vector pRbcBRL, (c) transfection of C. reinhardtii with the recombinant plasmid and (d) TNT tolerance and biodegradation capacity assessment of the modified alga.
  • Bilkent UNAM Turkey team established in January 2011 and member list has grown and changed in time. We shared a lot by that time and are extremely thankful to our former members. We did many brainstorming meetings, some of them was inconclusive. Team went through several problems that cause slightly negative impact on team. For a team to join from Turkey, money always becomes a problem. That’s why we are just able to send just two of our members to the regional jamboree. Moreover, laboratory equipment shipping takes very much time, even a restriction enzyme received one month after the ordering. Our team survive those restrictions and did its best. The force that always give impetus to ourselves is the friendship that we have.

  • In this summer; we learned about iGEM, how it works, what the biggest part of competition is and how we can win a medallion.

    When we were newbie, we organized a lot of meeting to find out our main project than we got idea for improving human welfare so we choose an idea from brainstorm results. We searched to get a gene to work on. As a result; lab works began. We added our protocols which are helpful to transform Chlamydomonas reinhardtii and Escherichia coli. Basic cloning protocols are available on our protocol page. Experiments and safety questions also available from link at the top section.

  • IGEM Project is our number one priority. As we worked really hard for our project it is important for us to make our dreams come true which is taking a gold medal. Questioning different items like experiment time, materials and methods and the efficiency of the experiments improved our skills and we started to look as a scientist to problems we faced. There were lots of challenging parts and sometimes we thought to quit but we listened to “Eye of The Tiger” and we did not give up from our experiments we delayed them yes, but we never gave up.

    IGEM team- meet-up was a great opportunity for us to communicate with the other groups, understood their projects and share our ideas. As we had a chance for an interview with Anatolia Agency, we also used media power to get attention to our project, iGEM and the importance of synthetic biology.

    Apart from that we did not forget having fun apart from our lab. As we planned a BBQ party, it was also a great chance for us to relax and gain some energy for our experiments, I did not even mention about the delicious foods and how joyful was our activities. Also like in iGEM team meeting up, we had a chance to introduce our project and main aim to other scientists and researchers in different areas.

    Links at above lead you to specific event and its photos suppose to be at that link. If not, look at gallery link. Thanks and get great time.

    Favorite Parts

    BBa_K596001

    BBa_K596004

    DNA Submission to Registry

  • We find it our duty to thank Gulce Itir Percin, Aydan Torun and Ömer Faruk Sarıoglu for their valuable assistance in conducting the experiments within our project’s scope, without which we could scarcely have expected to complete our project in time. We must also extend our thanks to Sustainable Technologies Laboratory Manager Zeynep E. Ülger for providing the necessary instructions and training for the use of a wide variety of laboratory equipment. We would like to acknowledge Prof. Dr. Tayfun Özçelik and Prof. Dr. Engin Umut Akkaya for their sincere support on our team and efforts for getting finanical support from Bilkent University. We thank the Turkish National Nanotechnology Institute (UNAM) for the administration of routine training sessions regarding laboratory safety, which allowed all our team members to operate without any personal danger throughout the experiments so far. We are also grateful to Assistant Professor Ayse Begum Tekinay and her laboratory for supplying much-needed reagents and assistance, both of which allowed our small team to work much faster than it otherwise could. And lastly we must thank our supervisor Turgay Tekinay for making this project possible and granting us a summer that was as entertaining as it was informative.

    Sponsors

    Share

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