Team:UNITN-Trento/Protocols

From 2013.igem.org

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</table>
</table>
</html>|Phusion-PCR}}
</html>|Phusion-PCR}}
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{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|OneTaq + Phu PCR|<html>
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1) Add the following components in a sterile microtube on ice:
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<table>
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<tr>
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<th colspan="2">
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Reaction Mix
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</th>
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</tr>
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<tr>
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<th>5x One Taq Buffer</th>
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<td>10 &micro;l</td>
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</tr>
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<tr>
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<th>Fwd Primer</th>
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<td>1 &micro;l</td>
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</tr>
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<tr>
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<th>Rev Primer</th>
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<td>1 &micro;l</td>
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</tr>
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<tr>
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<th>10 mM dNTP's</th>
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<td>1 &micro;l</td>
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</tr>
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<tr>
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<th>One Taq</th>
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<td>0.25 &micro;l</td>
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</tr>
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<tr>
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<th>Phusion</th>
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<td>0.3 &micro;l</td>
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</tr>
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<tr>
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<th>Template DNA</th>
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<td>50-100 ng</td>
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</tr>
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<tr>
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<th>H20</th>
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<td>up to 50 &micro;l</td>
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</tr>
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</table>
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2) Suggested reaction parameters:
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<table>
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<tr>
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<th>Cycle Step</th>
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<th>Cycles</th>
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<th>Temp [&deg;C]</th>
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<th>Time [s]</th>
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</tr>
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<tr>
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<td>Inital denaturation</td>
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<td>1</td>
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<td>94</td>
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<td>120</td>
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</tr>
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<tr>
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<td>Denaturation</td>
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<td>30</td>
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<td>94</td>
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<td>30</td>
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</tr>
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<tr>
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<td>Annealing</td>
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<td></td>
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<td>60 *</td>
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<td>60</td>
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</tr>
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<tr>
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<td>Extension</td>
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<td></td>
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<td>68</td>
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<td>60 s/Kbase</td>
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</tr>
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<tr>
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<td>Final Extension</td>
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<td>1</td>
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<td>68</td>
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<td>300</td>
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</tr>
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<tr>
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<td>Hold</td>
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<td>1</td>
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<td>4</td>
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<td>&#8734;</td>
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</tr>
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</table>
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</html>|OneTaq-Phu-PCR}}
{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Illustra GFX PCR DNA and Gel Band Purification Kit|<html>
{{:Team:UNITN-Trento/Templates/Styles/ProtocolSpoiler|Illustra GFX PCR DNA and Gel Band Purification Kit|<html>

Revision as of 09:23, 13 August 2013