Team:Groningen/protocols/Colony PCR

From 2013.igem.org

(Difference between revisions)
Line 48: Line 48:
   <tr>
   <tr>
     <td>MQ water</td>
     <td>MQ water</td>
-
     <td ALIGN=CENTER>up to 50&micro;l</td>
+
     <td ALIGN=CENTER>up to 20&micro;l</td>
     <td></td>
     <td></td>
   </tr>
   </tr>
Line 140: Line 140:
   </tr>
   </tr>
</table>  
</table>  
-
 
-
<br>Reference: http://www.thermoscientificbio.com/uploadedFiles/Resources/f-530s-product-information.pdf
 
</div>
</div>

Revision as of 11:09, 28 July 2013

Colony PCR

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


PCR reaction mixture:
Component 20µl Final concentration
MQ water up to 20µl
5x Phusion Buffer 2µl 1x
DMSO 5% 1.5µl
10mM dNTPs 1µl 200µM
primer F 1µl 0.5µM
primer R 1µl 0.5µM
template DNA 1µl ∼1ng
Phusion Polymerase
2U/µl		 0.3µl 0.01U/µl


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