Team:Lambert GA/Project

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<li class="first"><a href="https://2013hs.igem.org/Team:Lambert_GA" accesskey="1" title="">Home</a></li>
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                <li><a href="https://2013hs.igem.org/Team:Lambert_GA/team" accesskey="3" title="">Team</a></li>
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<li><a href="https://2013hs.igem.org/Team:Lambert_GA/project" accesskey="2" title="">Project</a></li>
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<li><a href="https://2013hs.igem.org/Team:Lambert_GA/labnotebook" accesskey="4" title="">Lab Notebook</a></li>
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<li><a href="https://2013hs.igem.org/Team:Lambert_GA/resultsconclusions" accesskey="5" title="">Conclusions</a></li>
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                <li><a href="https://2013hs.igem.org/Team:Lambert_GA/futureaspirations" accesskey="8" title="">Future</a></li>
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<li><a href="https://2013hs.igem.org/Team:Lambert_GA/safety" accesskey="5" title="">Safety</a></li>
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                <li><a href="https://2013hs.igem.org/Team:Lambert_GA/attributions" accesskey="6" title="">Attributions</a></li>
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                <li><a href="https://2013hs.igem.org/Team:Lambert_GA/humanpractices" accesskey="7" title="">Human Practices</a></li>           
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                <li><a href="https://2013hs.igem.org/Team:Lambert_GA/activities" accesskey="6" title="">Activities</a></li>             
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<font size=4><center><b><u><h1>Project</h1></u></b></center></font>
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      <h1>Our Project</h1>
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<center><img src="https://fbcdn-sphotos-a-a.akamaihd.net/hphotos-ak-prn1/p526x296/1016042_213819975433663_2086842284_n.jpg" width="450"/></center>
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      <h2>Project Calendar:</h2>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/1/15/Calendar.png" style="display:block; margin-left:auto; margin-right:auto;">
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    <h2>How we got our idea...</h2>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/1/10/LambertSlide1.PNG" style="display:block; margin-left:auto; margin-right:auto;">
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<font size=3><br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;In 2010, Georgia Institute of Technology’s iGem team submitted the part K410000, a periplasmic heat generator made with HybB and OmpA and AOX. The following year several teams used HybB in their projects with mixed results. Our project is built off of these projects  and it's purpose is to characterize and Biobrick HybB, a cold shock promoter.A Biobrick is a sequence of DNA that has standard prefixes and suffixes. Our goal is to test the reliability of HypB, the cold shock promoter, in order to use temperature to regulate protein expression. Our test will be to put RFP under HybB promotion and give it a cold shock treatment, which should enable HybB to act as a switch and turn on the red fluorescent protein in the <i> E. Coli</i> cells, if the HybB cells are reliable. We have decided to use HybB, even though it yeilded mix results in the past, because this cold shock promoter showed potential.<br><br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;HybB was synthesized for the first time by the 2005 UCSF iGEM team. The 2006 MIT iGEM team standardized HybB and submitted it to the parts registry. HybB has the part number J45503 and is not available from the Parts Registry. Georgia Tech’s 2010 iGEM team engineered a chassis to include HybB, OmpA, and AOX.  The 2011 Leuven iGEM team reported, “We see that promoter activity is induced when cells are transferred to 25°C and even when they are put in an ice bath (4°C). Unfortunately, however, cells that are kept at 37°C also display an increase in promoter activity, indicating leakiness in the system.” The 2011 Groningen team reported that the HybB Promoter did not work. Through sequencing, they proved that HybB was found in the cells, and they ran the reactions at the same temperatures.However, they could not get the same results as the Leuven team. Due to these more recent findings, we are not sure whether HybB works or not.<br><br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;We obtained HybB from Georgia Tech's 2010 iGEM Team, but upon sequencing and experimentation, we learned that the stock was not reliable.  We then designed primers to isolate the proposed HybB promoter sequence from the <i>E. Coli</i> DH10 genome. </font>
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      <h2>What we did in the lab...</h2>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/8/8a/LambertSlide2.PNG" style="display:block; margin-left:auto; margin-right:auto;">
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<center>http://ecocar.gatech.edu/wp-content/uploads/2010/11/buzz.jpg
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      <h2>CDA Methodology and Results</h2>
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      <h3>pMAL Protein Expression Kit</h3>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/4/42/LambertSlide3.PNG" width="722" height="542" style="display:block; margin-left:auto; margin-right:auto;">
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      <h3>Biobrick</h3>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/7/79/Biobrick_Slide_1.png" width="722" height="542" style="display:block; margin-left:auto; margin-right:auto;">
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      <td><img src="https://static.igem.org/mediawiki/2014hs/8/8f/Biobrick_Slide_2.png" width="722" height="542" style="display:block; margin-left:auto; margin-right:auto;">
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      <h2>Assay Methodology and Results</h2>
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      <p>
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      <td><img src="https://static.igem.org/mediawiki/2014hs/7/7c/LambertSlide4.PNG" style="display:block; margin-left:auto; margin-right:auto;">
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      <td><img src="https://static.igem.org/mediawiki/2014hs/a/ad/LambertSlide5.PNG" style="display:block; margin-left:auto; margin-right:auto;">
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      <h2> References</h2>
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      <p>Anitha, A., Sowmya, S., Kumar, P. T. S., Deepthi, S., Chennazhi, K. P., Ehrlich, H., . . . Jayakumar, R. Chitin and chitosan in selected biomedical applications. Progress in Polymer Science(0). doi: http://dx.doi.org/10.1016/j.progpolymsci.2014.02.008</p>
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      <p>Christodoulidou, A., Bouriotis, V., & Thireos, G. (1996). Two sporulation-specific chitin deacetylase-encoding genes are required for the ascospore wall rigidity of Saccharomyces cerevisiae. J Biol Chem, 271(49), 31420-31425.</p>
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      <p>Christodoulidou, A., Briza, P., Ellinger, A., & Bouriotis, V. (1999). Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett, 460(2), 275-279. </p>
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      <p>Christodoulidou, A., Briza, P., Ellinger, A., & Bouriotis, V. (1999). Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett, 460(2), 275-279. doi: http://dx.doi.org/10.1016/S0014-5793(99)01334-4</p>
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      <p>Jarmila, V., & Vavrikova, E. (2011). Chitosan derivatives with antimicrobial, antitumour and antioxidant activities--a review. Curr Pharm Des, 17(32), 3596-3607.</p>
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      <p>Liu, N., Chen, X.-G., Park, H.-J., Liu, C.-G., Liu, C.-S., Meng, X.-H., & Yu, L.-J. (2006). Effect of MW and concentration of chitosan on antibacterial activity of Escherichia coli. Carbohydrate Polymers, 64(1), 60-65. doi: http://dx.doi.org/10.1016/j.carbpol.2005.10.028</p>
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      <p>Martínez, J. P., Falomir, M. P., & Gozalbo, D. (2001). Chitin: A Structural Biopolysaccharide eLS: John Wiley & Sons, Ltd.</p>
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      <p>Martinou, A., Koutsioulis, D., & Bouriotis, V. (2003). Cloning and expression of a chitin deacetylase gene (CDA2) from Saccharomyces cerevisiae in Escherichia coli: Purification and characterization of the cobalt-dependent recombinant enzyme. Enzyme and Microbial Technology, 32(6), 757-763. doi: http://dx.doi.org/10.1016/S0141-0229(03)00048-6</p>
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      <p>Moussa, S. H., Tayel, A. A., & Al-Turki, A. I. (2013). Evaluation of fungal chitosan as a biocontrol and antibacterial agent using fluorescence-labeling. International Journal of Biological Macromolecules, 54(0), 204-208. doi:http://dx.doi.org/10.1016/j.ijbiomac.2012.12.029</p>
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      <p>Tsigos, I., Martinou, A., Kafetzopoulos, D., & Bouriotis, V. (2000). Chitin deacetylases: new, versatile tools in biotechnology. Trends in Biotechnology, 18(7), 305-312. doi: http://dx.doi.org/10.1016/S0167-7799(00)01462-1</p>
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      <p>Zakrzewska, A., Boorsma, A., Brul, S., Hellingwerf, K. J., & Klis, F. M. (2005). Transcriptional response of Saccharomyces cerevisiae to the plasma membrane-perturbing compound chitosan. Eukaryot Cell, 4(4), 703-715. doi: 10.1128/ec.4.4.703-715.2005</p>
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      <p>Zhang, H., Li, R., & Liu, W. (2011). Effects of Chitin and Its Derivative Chitosan on Postharvest Decay of Fruits: A Review. International Journal of Molecular Sciences, 12(2), 917-934.</p>
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      <p> (Anitha et al.; Christodoulidou, Bouriotis, & Thireos, 1996; A. Christodoulidou, P. Briza, A. Ellinger, & V. Bouriotis, 1999; Anna Christodoulidou, Peter Briza, Adi Ellinger, & Vassilis Bouriotis, 1999; Jarmila & Vavrikova, 2011; Liu et al., 2006; Martínez, Falomir, & Gozalbo, 2001; Martinou, Koutsioulis, & Bouriotis, 2003; Moussa, Tayel, & Al-Turki, 2013; Tsigos, Martinou, Kafetzopoulos, & Bouriotis, 2000; Zakrzewska, Boorsma, Brul, Hellingwerf, & Klis, 2005; Zhang, Li, & Liu, 2011)</p>
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    <p>Lõoke M, Kristjuhan K, Kristjuhan A. Extraction of genomic DNA from yeasts for PCR-based applications..  doi:10.2144/000113672. PubMed PMID: 21548894.</p>
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    <p>El Hadrami, A.; Adam, L.R.; El Hadrami, I.; Daayf, F. Chitosan in Plant Protection. Mar. Drugs 2010, 8, 968-987.</p>
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        <LI class="column" id="HOME"><p><a href="https://2014hs.igem.org/Team:Lambert_GA"><img src="https://static.igem.org/mediawiki/2014hs/5/57/Lambertlogo.png" width="100px" height="100px"></a></p></LI>
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<LI class="column" id="PROJECT"><p><a href="https://2014hs.igem.org/Team:Lambert_GA/Project"><img src="https://static.igem.org/mediawiki/2014hs/e/e2/Peach.png" width="100px" height="100px"></a></p></LI>
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        <LI class="column" id="SPONSORS"><p><a href="https://2014hs.igem.org/Team:Lambert_GA/Sponsors"><img src="https://static.igem.org/mediawiki/2014hs/6/6d/Strawberry.png" width="100px" height="100px"></a></p></LI>
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        <LI class="column" id="TEAM"><p><a href="https://2014hs.igem.org/Team:Lambert_GA/Team"><img src="https://static.igem.org/mediawiki/2014hs/c/c3/Blackberries.png" width="100px" height="100px"></a></p></LI>
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        <LI class="column" id="SAFETY"><p><a href="https://2014hs.igem.org/Team:Lambert_GA/Safety"><img src="https://static.igem.org/mediawiki/2014hs/0/0d/Raspberries-transparent.jpg" width="100px" height="100px"></a></p></LI>
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        <LI class="column" id="HUMAN PRACTICES"><p><a href="https://2014hs.igem.org/Team:Lambert_GA/HumanPractices"><img src="https://static.igem.org/mediawiki/2014hs/3/39/Grapes.png" width="100px" height="100px"></a></p></LI>
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Latest revision as of 01:11, 21 June 2014

Our Project

Project Calendar:

How we got our idea...

What we did in the lab...

CDA Methodology and Results

pMAL Protein Expression Kit

Biobrick

Assay Methodology and Results

References

Anitha, A., Sowmya, S., Kumar, P. T. S., Deepthi, S., Chennazhi, K. P., Ehrlich, H., . . . Jayakumar, R. Chitin and chitosan in selected biomedical applications. Progress in Polymer Science(0). doi: http://dx.doi.org/10.1016/j.progpolymsci.2014.02.008

Christodoulidou, A., Bouriotis, V., & Thireos, G. (1996). Two sporulation-specific chitin deacetylase-encoding genes are required for the ascospore wall rigidity of Saccharomyces cerevisiae. J Biol Chem, 271(49), 31420-31425.

Christodoulidou, A., Briza, P., Ellinger, A., & Bouriotis, V. (1999). Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett, 460(2), 275-279.

Christodoulidou, A., Briza, P., Ellinger, A., & Bouriotis, V. (1999). Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett, 460(2), 275-279. doi: http://dx.doi.org/10.1016/S0014-5793(99)01334-4

Jarmila, V., & Vavrikova, E. (2011). Chitosan derivatives with antimicrobial, antitumour and antioxidant activities--a review. Curr Pharm Des, 17(32), 3596-3607.

Liu, N., Chen, X.-G., Park, H.-J., Liu, C.-G., Liu, C.-S., Meng, X.-H., & Yu, L.-J. (2006). Effect of MW and concentration of chitosan on antibacterial activity of Escherichia coli. Carbohydrate Polymers, 64(1), 60-65. doi: http://dx.doi.org/10.1016/j.carbpol.2005.10.028

Martínez, J. P., Falomir, M. P., & Gozalbo, D. (2001). Chitin: A Structural Biopolysaccharide eLS: John Wiley & Sons, Ltd.

Martinou, A., Koutsioulis, D., & Bouriotis, V. (2003). Cloning and expression of a chitin deacetylase gene (CDA2) from Saccharomyces cerevisiae in Escherichia coli: Purification and characterization of the cobalt-dependent recombinant enzyme. Enzyme and Microbial Technology, 32(6), 757-763. doi: http://dx.doi.org/10.1016/S0141-0229(03)00048-6

Moussa, S. H., Tayel, A. A., & Al-Turki, A. I. (2013). Evaluation of fungal chitosan as a biocontrol and antibacterial agent using fluorescence-labeling. International Journal of Biological Macromolecules, 54(0), 204-208. doi:http://dx.doi.org/10.1016/j.ijbiomac.2012.12.029

Tsigos, I., Martinou, A., Kafetzopoulos, D., & Bouriotis, V. (2000). Chitin deacetylases: new, versatile tools in biotechnology. Trends in Biotechnology, 18(7), 305-312. doi: http://dx.doi.org/10.1016/S0167-7799(00)01462-1

Zakrzewska, A., Boorsma, A., Brul, S., Hellingwerf, K. J., & Klis, F. M. (2005). Transcriptional response of Saccharomyces cerevisiae to the plasma membrane-perturbing compound chitosan. Eukaryot Cell, 4(4), 703-715. doi: 10.1128/ec.4.4.703-715.2005

Zhang, H., Li, R., & Liu, W. (2011). Effects of Chitin and Its Derivative Chitosan on Postharvest Decay of Fruits: A Review. International Journal of Molecular Sciences, 12(2), 917-934.

(Anitha et al.; Christodoulidou, Bouriotis, & Thireos, 1996; A. Christodoulidou, P. Briza, A. Ellinger, & V. Bouriotis, 1999; Anna Christodoulidou, Peter Briza, Adi Ellinger, & Vassilis Bouriotis, 1999; Jarmila & Vavrikova, 2011; Liu et al., 2006; Martínez, Falomir, & Gozalbo, 2001; Martinou, Koutsioulis, & Bouriotis, 2003; Moussa, Tayel, & Al-Turki, 2013; Tsigos, Martinou, Kafetzopoulos, & Bouriotis, 2000; Zakrzewska, Boorsma, Brul, Hellingwerf, & Klis, 2005; Zhang, Li, & Liu, 2011)

Lõoke M, Kristjuhan K, Kristjuhan A. Extraction of genomic DNA from yeasts for PCR-based applications.. doi:10.2144/000113672. PubMed PMID: 21548894.

El Hadrami, A.; Adam, L.R.; El Hadrami, I.; Daayf, F. Chitosan in Plant Protection. Mar. Drugs 2010, 8, 968-987.

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