Team:Freiburg/Notebook/Methoden

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Revision as of 17:13, 25 September 2012

First Extension PCR

This step is necessary to add the the necessary restriction sites to the Direpeats. Pipette the following volumes into a PCR- tube:

Component	Volume
Water, nuclease-free	15,75 µl
DMSO	0,75 µl
dNTPs	0,5 µl
Phusion Buffer	5,0 µl
Phusion Polymerase	0,25 µl
Water, nuclease-free	up to 20 µl
Total vloume	20 µl

Second Extension PCR

This step is necessary to add enzyme- binding –sites to the outer restriction sites. Use the product of the first extension- PCR as template. Pipette the following volumes into a PCR- tube:

@@ Line 38: / Line 38: @@
 </p><br>
-==<span style="color:#000000">bla ==
+==<span style="color:#000000">Second Extension PCR ==
 <p>
-bla
+This step is necessary to add enzyme- binding –sites to the outer restriction sites.
+Use the product of the first extension- PCR as template.
+Pipette the following volumes into a PCR- tube:
 </p><br>