http://2010.igem.org/wiki/index.php?title=Construction_usu.html&feed=atom&action=historyConstruction usu.html - Revision history2024-03-29T15:09:51ZRevision history for this page on the wikiMediaWiki 1.16.5http://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=201605&oldid=prevCatramp at 00:00, 28 October 20102010-10-28T00:00:31Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=E. coli Protein Secretion System Construction=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=E. coli Protein Secretion System Construction=</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of <del class="diffchange diffchange-inline">our 2009 project </del>[https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of [https://2009.igem.org/Team:Utah_State <ins class="diffchange diffchange-inline">our 2009 project</ins>] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Overview==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Overview==</div></td></tr>
</table>Catramphttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=198510&oldid=prevAhatch at 22:10, 27 October 20102010-10-27T22:10:11Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==New Contributions==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==New Contributions==</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Two <del class="diffchange diffchange-inline">additional </del>secretion composite parts have been completed and submitted to the parts registry. These parts allowed us to test the functionality of the HlyA signal peptide. Most significantly, the proof of concept of the phasin/PHA complex secretion was verified using the HlyA targeted phasin.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Two <ins class="diffchange diffchange-inline">new </ins>secretion composite parts have been completed and submitted to the parts registry. These <ins class="diffchange diffchange-inline">composite </ins>parts allowed us to test the functionality of the HlyA signal peptide<ins class="diffchange diffchange-inline">. The newly created parts build on two parts submitted to the parts registry in 2009 (parts BBa_K208029, BBa_K208030)</ins>. Most significantly, the proof of concept of the phasin/PHA<ins class="diffchange diffchange-inline">-</ins>complex secretion was verified using the HlyA targeted phasin.</div></td></tr>
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</table>Ahatchhttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=193888&oldid=prevAhatch: /* Overview */2010-10-27T19:42:02Z<p><span class="autocomment">Overview</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Overview==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Overview==</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Recovery costs of cellular products can account for as much as 80% of production expenses (Hearn and Acosta, 2001). In an attempt to reduce recovery costs, we developed a library of Silver fusion compatible gene constructs coding for secretion signal peptides. Five signal peptides commonly utilized in E. coli were converted to BioBrick format. Testing was performed by fusing signal peptides to the cycle 3 mutant of green fluorescent protein and the protein phasin. GFP was targeted as a model protein because of its ease of detection. Phasin is a protein coexpressed in organisms that produce polyhydroxyalkanoates (PHAs), a class of bioplastics. Phasin has been shown to interact and bind to PHA granules and to modify granule size (York, 2001; Maehara, 1999). Phasin targeted for secretion could produce a phasin-PHA complex, effectively transporting PHAs into the extracellular media. Transport of PHA into the media could reduce costs of bioplastic production and make this alternative plastic a more viable competitor to traditional petrochemically derived plastics. At the time of last year’s jamboree, SDS-PAGE indicated that GFP and Phasin were both successfully released into the extracellular media when fused to the GeneIII signal peptide. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Recovery costs of cellular products can account for as much as 80% of production expenses (Hearn and Acosta, 2001). In an attempt to reduce recovery costs, we developed a library of Silver fusion compatible gene constructs coding for secretion signal peptides. Five signal peptides commonly utilized in E. coli were converted to BioBrick format. Testing was performed by fusing signal peptides to the cycle 3 mutant of green fluorescent protein and the protein phasin. GFP was targeted as a model protein because of its ease of detection. Phasin is a protein coexpressed in organisms that produce polyhydroxyalkanoates (PHAs), a class of bioplastics. Phasin has been shown to interact and bind to PHA granules and to modify granule size (York, 2001; Maehara, 1999). Phasin targeted for secretion could produce a phasin-PHA complex, effectively transporting PHAs into the extracellular media. Transport of PHA into the media could reduce costs of bioplastic production and make this alternative plastic a more viable competitor to traditional petrochemically<ins class="diffchange diffchange-inline">-</ins>derived plastics. At the time of last year’s jamboree, SDS-PAGE indicated that GFP and Phasin were both successfully released into the extracellular media when fused to the GeneIII signal peptide.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==New Contributions==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==New Contributions==</div></td></tr>
</table>Ahatchhttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=192068&oldid=prevCdmiller: /* New Contributions */2010-10-27T18:44:29Z<p><span class="autocomment">New Contributions</span></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Cells containing the genes necessary for PHA production, HlyBD membrane proteins, and the HlyA targeted phasin (BBa_K390501) were grown in M9 minimal media. Cells containing the genes for PHA production and the HlyBD membrane proteins were grown as a negative control. <del class="diffchange diffchange-inline"> </del>After 30 hours of growth, <del class="diffchange diffchange-inline">a nucleating agent </del>was <del class="diffchange diffchange-inline">added to both cultures and centrifuged. The resulting pellet was purified </del>and analyzed using quantitative 1H NMR. Differences in intensity of the 1H NMR spectra indicate that PHA is released into the extracellular media when HlyA targeted phasin is expressed. We have reason to believe that this platform can be optimized to significantly reduce costs associated with commercial PHA production. This additional work further validates the parts created by USU’s 2009 team and the potential for their implementation in other cellular product recovery.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Cells containing the genes necessary for PHA production, HlyBD membrane proteins, and the HlyA targeted phasin (BBa_K390501) were grown in M9 minimal media. Cells containing the genes for PHA production and the HlyBD membrane proteins were grown as a negative control. After 30 hours of growth, <ins class="diffchange diffchange-inline">secreted PHA </ins>was <ins class="diffchange diffchange-inline">isolated </ins>and analyzed using quantitative 1H NMR. Differences in intensity of the 1H NMR spectra indicate that PHA is released into the extracellular media when HlyA targeted phasin is expressed. We have reason to believe that this platform can be optimized to significantly reduce costs associated with commercial PHA production. This additional work further validates the parts created by USU’s 2009 team and the potential for their implementation in other cellular product recovery.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:USU_NMR_spectra.png|600px|center]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:USU_NMR_spectra.png|600px|center]]</div></td></tr>
</table>Cdmillerhttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=191999&oldid=prevCdmiller: /* New Contributions */2010-10-27T18:42:18Z<p><span class="autocomment">New Contributions</span></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Fig 2: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control. Notice the differences in <del class="diffchange diffchange-inline">intensity. Due to the similarity in density of PHA granules and E. coli, PHA in the negative control is expected as cells containing intracellular PHA can be present in the resulting pellet</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Fig 2: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control <ins class="diffchange diffchange-inline">that is producing PHA but not secreting phasin</ins>. Notice the differences in <ins class="diffchange diffchange-inline">peak intensities (y-axis)</ins>.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td></tr>
</table>Cdmillerhttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=187633&oldid=prevCatramp at 16:23, 27 October 20102010-10-27T16:23:48Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Construction</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Construction</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></div></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div></div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">=Cyanobacterial Toolkit Construction=</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">==Integration Plasmid Construction==</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">The integration plasmid was constructed starting with pSB1A3 (Figure 1A). Four additional restriction sites (BamHI, XhoI, NdeI, and NheI) were added outside of the BioBrick docking sequence to allow recombination regions and Synechocystis-specific antibiotic resistance genes to be cloned in (Figure 1B). The two recombination regions (RS1 and RS2) where then cloned in flanking the BioBrick site, and a resistance gene (either Kanamycin resistance or Chloramphenicol resistance) was cloned in next to the BioBrick site (Figure 1C). An additional HindIII restriction site was added between the resistance gene and the RS1 region. This plasmid design allows for selection of the integrated BioBrick using the inserted resistance gene. These recombination regions and resistance genes can be easily exchanged for any desired region or resistance using restriction digest and ligations. </ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">=Overview=</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">We decided to expand the current BioBrick plasmid naming convention for integration plasmids by adding the "_Int[Integrated Resistance][Species Abbreviation (include only if recombination regions are added)]" to the plasmid name. This provides the user with information on what resistance the cells with integrated BioBricks will have, and what species-specific recombination sites (if any) are part of the plasmid.</ins></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:USU_Plamid_1C.png|279px]]</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">Recovery costs of cellular products can account for as much as 80% of production expenses </del>(<del class="diffchange diffchange-inline">Hearn and Acosta, 2001</del>)<del class="diffchange diffchange-inline">. In an attempt to reduce recovery costs, we developed a library of Silver fusion compatible gene constructs coding for secretion signal peptides. Five signal peptides commonly utilized in E. coli were converted to BioBrick format. Testing was performed by fusing signal peptides to </del>the <del class="diffchange diffchange-inline">cycle 3 mutant </del>of <del class="diffchange diffchange-inline">green fluorescent protein and the protein phasin</del>. <del class="diffchange diffchange-inline"> GFP was targeted as a model protein because of its ease of detection. Phasin is a protein coexpressed in organisms that produce polyhydroxyalkanoates </del>(<del class="diffchange diffchange-inline">PHAs</del>)<del class="diffchange diffchange-inline">, a class of bioplastics</del>. <del class="diffchange diffchange-inline"> Phasin has been shown to interact and bind to PHA granules and to modify granule size </del>(<del class="diffchange diffchange-inline">York, 2001; Maehara, 1999</del>)<del class="diffchange diffchange-inline">. Phasin targeted for secretion could produce a phasin-PHA complex, effectively transporting PHAs into the extracellular media. Transport of PHA into the media could reduce costs of bioplastic production </del>and <del class="diffchange diffchange-inline">make this alternative plastic a more viable competitor to traditional petrochemically derived plastics. At the time of last year’s jamboree, SDS-PAGE indicated that GFP </del>and <del class="diffchange diffchange-inline">Phasin were both successfully released into the extracellular media when fused to the GeneIII signal peptide</del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">Figure 1. </ins>(<ins class="diffchange diffchange-inline">A</ins>) <ins class="diffchange diffchange-inline">The pSB1A3 plasmid which served as </ins>the <ins class="diffchange diffchange-inline">starting point </ins>of <ins class="diffchange diffchange-inline">construction</ins>. (<ins class="diffchange diffchange-inline">B</ins>) <ins class="diffchange diffchange-inline">Four restriction sites added surrounding the BioBrick site in pSB1A3</ins>. (<ins class="diffchange diffchange-inline">C</ins>) <ins class="diffchange diffchange-inline">Finished pSB1A3_IntC plasmid with recombination regions RS1 </ins>and <ins class="diffchange diffchange-inline">RS2 </ins>and <ins class="diffchange diffchange-inline">chloramphenicol resistance gene</ins>.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>=<del class="diffchange diffchange-inline">New Contributions</del>=</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>==<ins class="diffchange diffchange-inline">Promoter Construction==</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">Using PCR, we amplified 15 native Synechocystis promoters and their native 5’ UTR and RBS regions from genomic DNA. Using oligonucleotide pairs, we created the 15 promoters alone (without their native 5’UTR and RBS), a standard native 5’UTR and RBS from Synechocystis, a consensus 5’UTR and RBS, and two secretion tags that function in Synechocystis, one native to the species and one artificially created. </ins></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">Table 2. Primers used for PCR amplification from genomic DNA and oligonucleotide pairs. Four base pairs were added to the ends of the primers to increase digestion efficiency and equalize the Tm of the primer pairs. Green sequences are added BioBrick prefix and suffix sequences. Red bases are the bases mutated with site-directed mutagenesis.</ins></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></ins></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">=E. coli Protein Secretion System Construction=</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">==Overview==</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">Recovery costs of cellular products can account for as much as 80% of production expenses (Hearn and Acosta, 2001). In an attempt to reduce recovery costs, we developed a library of Silver fusion compatible gene constructs coding for secretion signal peptides. Five signal peptides commonly utilized in E. coli were converted to BioBrick format. Testing was performed by fusing signal peptides to the cycle 3 mutant of green fluorescent protein and the protein phasin. GFP was targeted as a model protein because of its ease of detection. Phasin is a protein coexpressed in organisms that produce polyhydroxyalkanoates (PHAs), a class of bioplastics. Phasin has been shown to interact and bind to PHA granules and to modify granule size (York, 2001; Maehara, 1999). Phasin targeted for secretion could produce a phasin-PHA complex, effectively transporting PHAs into the extracellular media. Transport of PHA into the media could reduce costs of bioplastic production and make this alternative plastic a more viable competitor to traditional petrochemically derived plastics. At the time of last year’s jamboree, SDS-PAGE indicated that GFP and Phasin were both successfully released into the extracellular media when fused to the GeneIII signal peptide. </ins></div></td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">==New Contributions==</ins></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Two additional secretion composite parts have been completed and submitted to the parts registry. These parts allowed us to test the functionality of the HlyA signal peptide. Most significantly, the proof of concept of the phasin/PHA complex secretion was verified using the HlyA targeted phasin.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Two additional secretion composite parts have been completed and submitted to the parts registry. These parts allowed us to test the functionality of the HlyA signal peptide. Most significantly, the proof of concept of the phasin/PHA complex secretion was verified using the HlyA targeted phasin.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Fig 2: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control. Notice the differences in intensity. Due to the similarity in density of PHA granules and E. coli, PHA in the negative control is expected as cells containing intracellular PHA can be present in the resulting pellet.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Fig 2: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control. Notice the differences in intensity. Due to the similarity in density of PHA granules and E. coli, PHA in the negative control is expected as cells containing intracellular PHA can be present in the resulting pellet.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td></tr>
</table>Catramphttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=183291&oldid=prevCatramp at 13:57, 27 October 20102010-10-27T13:57:24Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Fig 1: Parts BBa_K390501 and BBa_K390502 submitted to the parts registry.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Fig 1: Parts BBa_K390501 and BBa_K390502 submitted to the parts registry.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>York GM, Junker BH, Stubbe J, Sinskey AJ (2001) Accumulation of the PhaP Phasin of Ralstonia eutropha is dependent on production of polyhydroxybutyrate in cells. J Bacteriol 183:4217-4226</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>York GM, Junker BH, Stubbe J, Sinskey AJ (2001) Accumulation of the PhaP Phasin of Ralstonia eutropha is dependent on production of polyhydroxybutyrate in cells. J Bacteriol 183:4217-4226</div></td></tr>
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</table>Catramphttp://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=175828&oldid=prevAbiezer20 at 07:15, 27 October 20102010-10-27T07:15:13Z<p></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">Lorem ipsum dolor sit amet, consectetur adipiscing elit. Pellentesque vel sapien lectus, in dignissim leo. Duis diam tortor, facilisis id tincidunt in, fringilla a est. Ut elit libero, venenatis porttitor ultrices eget, rutrum sed diam. Duis semper libero posuere lorem pharetra pretium. Nulla a lacus eu ante elementum rutrum et a neque. Aenean volutpat tellus non est tempus rhoncus in nec lacus. Vivamus ac nibh at eros porta iaculis. Duis non magna ligula. Vivamus nec ipsum quis magna faucibus eleifend pretium sed lectus. Pellentesque sit amet augue non dolor tempor pharetra. Vivamus rhoncus turpis sed risus mollis sit amet ullamcorper tellus molestie. Vivamus eget lorem eget purus scelerisque sollicitudin vel ut ipsum. Nam aliquet urna odio. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos. Etiam semper condimentum felis, eget mollis neque pulvinar et. Cras a augue enim, a facilisis nisi. Suspendisse potenti. Sed sit amet est augue. Quisque non tortor enim, quis pretium ante.</del></div></td><td colspan="2"> </td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td></tr>
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</table>Abiezer20http://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=175798&oldid=prevAbiezer20: /* USU TEAM */2010-10-27T07:13:26Z<p><span class="autocomment">USU TEAM</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Lorem ipsum dolor sit amet, consectetur adipiscing elit. Pellentesque vel sapien lectus, in dignissim leo. Duis diam tortor, facilisis id tincidunt in, fringilla a est. Ut elit libero, venenatis porttitor ultrices eget, rutrum sed diam. Duis semper libero posuere lorem pharetra pretium. Nulla a lacus eu ante elementum rutrum et a neque. Aenean volutpat tellus non est tempus rhoncus in nec lacus. Vivamus ac nibh at eros porta iaculis. Duis non magna ligula. Vivamus nec ipsum quis magna faucibus eleifend pretium sed lectus. Pellentesque sit amet augue non dolor tempor pharetra. Vivamus rhoncus turpis sed risus mollis sit amet ullamcorper tellus molestie. Vivamus eget lorem eget purus scelerisque sollicitudin vel ut ipsum. Nam aliquet urna odio. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos. Etiam semper condimentum felis, eget mollis neque pulvinar et. Cras a augue enim, a facilisis nisi. Suspendisse potenti. Sed sit amet est augue. Quisque non tortor enim, quis pretium ante.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Lorem ipsum dolor sit amet, consectetur adipiscing elit. Pellentesque vel sapien lectus, in dignissim leo. Duis diam tortor, facilisis id tincidunt in, fringilla a est. Ut elit libero, venenatis porttitor ultrices eget, rutrum sed diam. Duis semper libero posuere lorem pharetra pretium. Nulla a lacus eu ante elementum rutrum et a neque. Aenean volutpat tellus non est tempus rhoncus in nec lacus. Vivamus ac nibh at eros porta iaculis. Duis non magna ligula. Vivamus nec ipsum quis magna faucibus eleifend pretium sed lectus. Pellentesque sit amet augue non dolor tempor pharetra. Vivamus rhoncus turpis sed risus mollis sit amet ullamcorper tellus molestie. Vivamus eget lorem eget purus scelerisque sollicitudin vel ut ipsum. Nam aliquet urna odio. Class aptent taciti sociosqu ad litora torquent per conubia nostra, per inceptos himenaeos. Etiam semper condimentum felis, eget mollis neque pulvinar et. Cras a augue enim, a facilisis nisi. Suspendisse potenti. Sed sit amet est augue. Quisque non tortor enim, quis pretium ante.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>In addition to our work developing the cyanobacterial toolkit, an effort was made to complete and further characterize portions of our 2009 project [https://2009.igem.org/Team:Utah_State] during this iGEM season. USU’s 2009 iGEM team focused on simplifying cellular product recovery by developing a platform for protein secretion systems in the BioBricks format.</div></td></tr>
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</table>Abiezer20http://2010.igem.org/wiki/index.php?title=Construction_usu.html&diff=175709&oldid=prevAhatch: /* USU TEAM */2010-10-27T07:06:57Z<p><span class="autocomment">USU TEAM</span></p>
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<td colspan='2' style="background-color: white; color:black;">← Older revision</td>
<td colspan='2' style="background-color: white; color:black;">Revision as of 07:06, 27 October 2010</td>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Fig: Parts BBa_K390501 and BBa_K390502 submitted to the parts registry.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Fig <ins class="diffchange diffchange-inline">1</ins>: Parts BBa_K390501 and BBa_K390502 submitted to the parts registry.</div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Fig: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control. Notice the differences in intensity. Due to the similarity in density of PHA granules and E. coli, PHA in the negative control is expected as cells containing intracellular PHA can be present in the resulting pellet.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Fig <ins class="diffchange diffchange-inline">2</ins>: 1H NMR spectra with PHA characteristic peaks amplified. The top spectrum reports PHA isolated gravimetrically from a culture containing HlyA targeted phasin. The bottom spectrum reports PHA in the negative control. Notice the differences in intensity. Due to the similarity in density of PHA granules and E. coli, PHA in the negative control is expected as cells containing intracellular PHA can be present in the resulting pellet.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=References=</div></td></tr>
</table>Ahatch