Team:UCL London

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<h1>Our Project</h1>
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<strong>Making supercoiled pDNA technology viable</strong>
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<p>We are developing a toolkit for the industrial manufacturing of supercoiled plasmid DNA. This will enable a widespread adoption of pDNA based technologies, most relevantly DNA vaccines - the future of immunization. Therefore we are a manufacturing project with a strong impact on the global health sector. <br />
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<div id="header"><div id="topleftcorner"></div><a href="https://2011.igem.org/Team:UCL_London/Research" id="researchlink"></a><a href="https://2011.igem.org/Team:UCL_London/Manufacturing" id="manufacturinglink"></a><a href="https://2011.igem.org/Team:UCL_London/Medicine" id="medicinelink"></a><a href="https://2011.igem.org/Team:UCL_London/HumanPractices" id="humanpraclink"></a><a href="https://2011.igem.org/Team:UCL_London" id="indexlink"></a><a href="https://2011.igem.org/Team:UCL_London/Safety" id="safetylink"></a><a href="https://2011.igem.org/Team:UCL_London/Contributions" id="contributionslink"></a><a href="https://2011.igem.org/Team:UCL_London/SyntheticBiology" id="synbiolink"></a><a href="https://2011.igem.org" id="igemlink" target="_blank"></a><a href="https://2011.igem.org/Team:UCL_London/Team" id="teamlink"></a><a href="http://www.ucl.ac.uk" id="ucllink" target="_blank"></a></div></html>
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Our toolkit consists of five modules tackling different bottlenecks in the manufacturing process from production over lysis to filtration as well as providing tools for monitoring and improving the overall quantity and quality of the product.<br />
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<div id="footer">Sitemap | Contact | [[Team:UCL_London/Safety|Safety]] | Attributions | [[Team:UCL_London/Bibliography|Bibliography]] | Protocols | <html><a href="http://partsregistry.org/cgi/partsdb/pgroup.cgi?pgroup=iGEM2011&group=UCL_London" target="_blank">Registry of Parts</a> | <a href="http://www.ucl.ac.uk/biochemeng/index.html" target="_blank">UCL Biochemical Engineering</a></html></div>
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Recognizing that the adoption of vaccines and other biological technologies also depends strongly on their social acceptance, we're investing heavily into public engagement with synthetic biology and are exploring the process by which new scientific disciplines arrive in the public sphere.</p>
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<strong>More Details:</strong><p> Plasmid DNA can be used as a therapeutic product as a source of recombinant DNA for DNA vaccines or as a non-viral vector for gene therapy.
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DNA vaccination is a novel technique involving delivery of genetic material into a living host and elicitation of immune response (Choudhary, 2011). It has been shown that DNA vaccine technology has great potential in helping against a wide range of diseases.<br />  
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The success of gene therapy is largely dependent on the usage of vectors that can efficiently deliver genes with minimal toxicity. Viral vectors have high transduction, but their toxicity and immunogenicity raise safety concerns. Non-viral vectors, such as recombinant plasmids, are particularly suitable with respect to simplicity of use, ease of large-scale production and minimal immune response (Li and Huang, 2000).<br />  
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Guidance documents produced by the Food and Drug Administration (FDA) and the European Medicines Agency (EMEA), require plasmids in bulk production of plasmid DNA for therapeutic use to be greater than 80% supercoiled. Current production methods depend on multiple and costly purification steps to enable this quality.<br />  
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We propose a synthetic biology approach to bypass this production burden by making use of the enzyme, gyrase, involved in the supercoiling process. Gyrase, an ATP-dependent enzyme belonging to the topoisomerase family, makes double-strand breaks and introduces negative supercoils into DNA (Dominguez et al., 2003).<br />  
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Our project aim is to increase the yield of supercoiled plasmid DNA in large scale manufacture of plasmid DNA.<br />  
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To achieve our goal, we suggest a combination of two approaches:<br />
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1.     To increase the number of gyrase enzymes per cell by incorporating a gyrase expression cassette into plasmids.<br />
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2.     To improve the interaction that facilitates supercoiling by introducing new gyrase binding sites to the plasmid.<br />  
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The project outcome aims at improving industrial manufacturing of plasmid DNA for therapeutics by<br />
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1.     Increasing the stability of the product and making the product more compatible with downstream recovery, purification processes and final delivery process (Yau et al., 2008),<br />
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2.     Reducing the final cost of producing the product and thus increasing the availability to customers.
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<strong>References</strong>
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Choudhary, R. (2011) DNA Vaccine: A Promise For Cure [online] Available from:
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http://www.biotecharticles.com/Biotechnology-products-Article/DNA-Vaccine-A-
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Promise-For-Cure-557.html [Accessed 5 July 2011]
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Dominguez, Y. R., et al. (2003) Plasmid DNA Supercoiling and Gyrase Activity in
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Escherichia coli Wild-Type and rpsS Stationary-Phase. Journal of Bacteriology.
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185(3):1097-1100.
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Li, S. and Huang, L. (2000) Nonviral Gene Therapy: Promises and Challenges. Gene
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Therapy. 7:31-34.
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Yau, S. Y., et al. (2008) Host Strain Influences on Supercoiled Plasmid DNA
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Production in Escherichia coli: Implications for Efficient Design of Large-Scale
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Processes. Biotechnology and Bioengineering. 101(3):529-544
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The European Agency for the Evaluation of Medicinal Products. (2001) Note for
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Guidance on the Quality, Preclinical and Clinical Aspects of Gene Transfer Medicinal
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Products.
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U.S. Department of Health and Human Services, Food and Drug Administration.
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(2007) Guidance for Industry: Considerations for Plasmid DNA Vaccines for
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infectious Disease Indications.
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