Team:Groningen/protocols/PCR

From 2013.igem.org

(Difference between revisions)
Line 108: Line 108:
   <tr>
   <tr>
     <td>98&deg;C</td>
     <td>98&deg;C</td>
-
     <td>2min</td>
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     <td ALIGN=CENTER>2min</td>
     <td>1td>
     <td>1td>
   </tr>
   </tr>
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   <tr>
   <tr>
     <td>98&deg;C</td>
     <td>98&deg;C</td>
-
     <td>10sec</td>
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     <td ALIGN=CENTER>10sec</td>
     <td rowspan="3">34</td>
     <td rowspan="3">34</td>
   </tr>
   </tr>
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   <tr>
   <tr>
     <td>T<sub>m</sub>-5&deg;C</td>
     <td>T<sub>m</sub>-5&deg;C</td>
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     <td>25sec</td>
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     <td ALIGN=CENTER>25sec</td>
   </tr>
   </tr>
   <tr>
   <tr>
     <td>72&deg;C</td>
     <td>72&deg;C</td>
-
     <td>30sec/kb</td>
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     <td ALIGN=CENTER>30sec/kb</td>
   </tr>
   </tr>
   <tr>
   <tr>
     <td>72&deg;C</td>
     <td>72&deg;C</td>
-
     <td>10min</td>
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     <td ALIGN=CENTER>10min</td>
     <td>1</td>
     <td>1</td>
   </tr>
   </tr>
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     <td>4&deg;C</td>
     <td>4&deg;C</td>
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     <td>&infin;</td>
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     <td ALIGN=CENTER>&infin;</td>
     <td>1</td>
     <td>1</td>
   </tr>
   </tr>

Revision as of 10:43, 28 July 2013

PCR

Materials:
  • PCR tubes
  • MQ water
  • DMSO 5%
  • Phusion buffer
  • DNTPs
  • F- and R-primer
  • Phusion Polymerase
  • DNA template


PCR reaction mixture:
Component 50µl Final concentration
MQ water up to 50µl
5x Phusion Buffer 10µl 1x
DMSO 5% 1.5µl
10mM dNTPs 1µl 200µM
primer F 2.5µl 0.5µM
primer R 2.5µl 0.5µM
template DNA 1µl ∼1ng
Phusion Polymerase
2U/µl		 0.3µl 0.01U/µl


Cycling instructions:
Temperature Time Number of Cycles
98°C 2min 1td>
98°C 10sec 34
Tm-5°C 25sec
72°C 30sec/kb
72°C 10min 1
4°C ∞ 1

Reference: http://www.thermoscientificbio.com/uploadedFiles/Resources/f-530s-product-information.pdf