Team:UC Chile2/Protocols

From 2012.igem.org

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<li>Adjust pH to 7.5 with NaOH 1M</li>
<li>Adjust pH to 7.5 with NaOH 1M</li>
</ul>
</ul>
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<h3>BG-11</h3>
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<br>
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For 1L:
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<ul>
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<li>1.5gr NaNO3</li>
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<li>1L distilled H2O </li>
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<li>10mL Concentrated salts solution 100X (4°C)</li>
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<li>200µL H2PO4 1M (Room T°)</li>
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<li>10gr BactoAgar -optional-</li>
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</ul>
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<h3>BG-110</h3>
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<br>
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For 1L:
 +
<ul>
 +
<li>1L distilled H2O </li>
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<li>10mL Concentrated salts solution 100X (4°C)</li>
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<li>200µL H2PO4 1M (Room T°)</li>
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<li></li>
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<li>10gr BactoAgar -optional-</li>
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</ul>
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<h3>BG-11C</h3>
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<br>
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For 1L:
 +
<ul>
 +
<li>1.5gr NaNO3</li>
 +
<li>1L distilled H2O </li>
 +
<li>10mL Concentrated salts solution 100X (4°C)</li>
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<li>200µL H2PO4 1M (Room T°)</li>
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<li>0.84gr NaHCO3</li>
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<li>10gr BactoAgar -opcional-</li>
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</ul>
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<h3>Concentrated salts solution 100X</h3>
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For 1L:
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<ul>
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<li>7.5gr SO4Mg * 7H2O</li>
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<li>3.6gr CaCl2 * 2H2O</li>
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<li>0.6gr Citric Acid</li>
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<li>0.6gr Ammonium Ferric Citrate</li>
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<li>0.093gr EDTA-Na2 </li>
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<li>2.0gr CO3 Na2 </li>
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<li>0.286gr BO3H3</li>
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<li>0.181gr MnCl2 * 4H2O</li>
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<li>0.022gr ZnSO4 * 7H2O</li>
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<li>0.039gr NaMoO4 * 2H2O</li>
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<li>0.0079gr CuSO4 * 5H2O</li>
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<li>0.0049gr CoCl2 * 6H2O</li>
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<li>1L distilled H2O </li>
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</ul>
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<br>
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Autoclave and store at 4°C.
<div id="Buffers">
<div id="Buffers">
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<h3>Gibson Assembly</h3>
<h3>Gibson Assembly</h3>
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<p>Gibson Assembly is a DNA assembly method created by Daniel Gibson during the development of the first Synthetic Genome (Synthia) (reference YYY). Its adaptation to a cloning method allows fast and accurate production of increasingly complex constructions. The strategy behind the method relies on PCR to obtain different parts which share a 40bp homology region, and a 3 enzyme reaction which produces cohesive ends, fills the gaps between the parts and ligates the resulting construct into a scarless assembly of various (>2) parts.</p>
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<p>Gibson Assembly is a DNA assembly method created by Daniel Gibson during the development of the first Synthetic Genome (Synthia) [[#2|2]]. Its adaptation to a cloning method allows fast and accurate production of increasingly complex constructions. The strategy behind the method relies on PCR to obtain different parts which share a 40bp homology region, and a 3 enzyme reaction which produces cohesive ends, fills the gaps between the parts and ligates the resulting construct into a scarless assembly of various (>2) parts.</p>
<h4>Primer design</h4>
<h4>Primer design</h4>
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<h1>References</h1>
<h1>References</h1>
<div id="1">
<div id="1">
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[1] Julian J. Eaton-Rye , “Construction of Gene Interruptions and Gene Deletions in the Cyanobacterium Synechocystis sp. Strain PCC 6803 ”, Photosynthesis Research Protocols, Methods in Molecular Biology, vol. 684, DOI 10.1007/978-1-60761-925-3_22  
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[1]Julian J. Eaton-Rye , “Construction of Gene Interruptions and Gene Deletions in the Cyanobacterium Synechocystis sp. Strain PCC 6803 ”, Photosynthesis Research Protocols, Methods in Molecular Biology, vol. 684, DOI 10.1007/978-1-60761-925-3_22  
</div>
</div>
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<div id="2">
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[2]Gibson DG, Young L, Chuang RY, Venter JC, Hutchison CA 3rd, Smith HO. (2009). "Enzymatic assembly of DNA molecules up to several hundred kilobases". Nature Methods 6 (5): 343–345. doi:10.1038/nmeth.1318
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</div>
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Latest revision as of 11:06, 25 September 2012

Project: Luxilla - Pontificia Universidad Católica de Chile, iGEM 2012