Difference between revisions of "Team:CPU CHINA/Improve"

 
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<h1>Improve</h1>
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<p>For teams seeking to improve upon a previous part or project, you should document all of your work on this page. Please remember to include all part measurement and characterization data on the part page on the Registry. Please include a link to your improved part on this page.</p>
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<h3>Gold Medal Criterion #2</h3>
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<p><b>Standard Tracks:</b> Create a new part that has a functional improvement upon an existing BioBrick part. The sequences of the new and existing parts must be different. You must perform experiments with both parts to demonstrate this improvement.  Document the experimental characterization on the Part's Main Page on the Registry for both the existing and new parts. Both the new and existing Main Page of each Part’s Registry entry must reference each other. Submit a sample of the new part to the Registry.
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The existing part must NOT be from your 2018 part number range and must be different from the part documented in bronze #4.
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<b>Special Tracks:</b> Improve the function of an existing iGEM project (that your current team did not originally create) and display your achievement on your wiki.</p>
 
  
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<h1>Part improvement
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<h4>Hepatitis C virus nonstructural protein 5B (NS5B) is a RNA-dependent RNA polymerase (RdRp) which initiates de novo RNA synthesis with the help of specific RNA promoters. In our project, the RNA interference is conditional because the pri-miRNA analogue which partially pairs with an inhibitory strand is unable to be processed by DROSHA. Once NS5B is introduced to catalyze a substitutional hybridation to release the pri-miRNA in the nucleus, the analogue gets to evolve into miRNA and interferes with the target gene mRNA.
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<h4>Since the existent NS5B from Standard Parts collection which can not translocate into the nucleus does not suit our needs, we tend to design our own part that is able to do its job in the nucleus. After deep consideration, we decided to improve the function of the part submitted by iGEM14<u> </u>Warwick (<a href="http://parts.igem.org/Part:BBa_K1442028">BBa_K1442028</a>). We load a nuclear location sequence (NLS) to the N terminal of NS5B (NS5B<sup>NLS</sup>) to transport the RdRP into the nucleus (<a herf="http://parts.igem.org/Part:BBa_K2624002">BBa_K2624002</a>). We demonstrate this improvement by indirect immunofluorescence. As the following figure shows, NS5B with NLS (NLS+) have a higher ability of nuclear translocation than NS5B without NLS (NLS-).
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Latest revision as of 14:11, 6 December 2018

Part improvement

Hepatitis C virus nonstructural protein 5B (NS5B) is a RNA-dependent RNA polymerase (RdRp) which initiates de novo RNA synthesis with the help of specific RNA promoters. In our project, the RNA interference is conditional because the pri-miRNA analogue which partially pairs with an inhibitory strand is unable to be processed by DROSHA. Once NS5B is introduced to catalyze a substitutional hybridation to release the pri-miRNA in the nucleus, the analogue gets to evolve into miRNA and interferes with the target gene mRNA.

Since the existent NS5B from Standard Parts collection which can not translocate into the nucleus does not suit our needs, we tend to design our own part that is able to do its job in the nucleus. After deep consideration, we decided to improve the function of the part submitted by iGEM14 Warwick (BBa_K1442028). We load a nuclear location sequence (NLS) to the N terminal of NS5B (NS5BNLS) to transport the RdRP into the nucleus (BBa_K2624002). We demonstrate this improvement by indirect immunofluorescence. As the following figure shows, NS5B with NLS (NLS+) have a higher ability of nuclear translocation than NS5B without NLS (NLS-).