Paris/October 12

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[[Paris/October 11|yesterday]] -- [[Paris/October 13|tomorrow]] <br>
== PCR ==
== PCR ==
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<br>
<br>
------
------
 +
MIX PCR :<br>
- 27µL H2O<br>
- 27µL H2O<br>
- 10µL Buffer10X GCRich (Finnzyme)<br>
- 10µL Buffer10X GCRich (Finnzyme)<br>
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- 0,5µL Phusion Taq<br>
- 0,5µL Phusion Taq<br>
------
------
-
- 2' 98°C<br>
+
Timing : <br><br>
-
<br>
+
First :<br>
-
30X
+
- 2' at 98°C<br>
-
- 20" 98°C<br>
+
* 30X :<br>
-
- 1'40" 72°C<br>
+
- 20" at 98°C<br>
-
<br>
+
- 1'40" at 72°C<br>
-
- 10' 72°C<br>
+
then :<br>
-
- infinitas 4°C
+
- 10' at 72°C<br>
 +
- infinitas at 4°C
 +
 
 +
== Digestion reactions ==

Latest revision as of 14:27, 15 October 2007

yesterday -- tomorrow

PCR

I made a PCR onto the L59 miniprep template having the [ptet>>lox66-GFP-terminator-lox71-mRFP] construction in order to add a forward restriction site for PacI and a backward restriction site for XmaI.


MIX PCR :
- 27µL H2O
- 10µL Buffer10X GCRich (Finnzyme)
- 1,5µL DMSO 100%
- 2µL Template L59
- 4µL O47
- 4µL O48
- 0,5µL Phusion Taq


Timing :

First :
- 2' at 98°C

  • 30X :

- 20" at 98°C
- 1'40" at 72°C
then :
- 10' at 72°C
- infinitas at 4°C

Digestion reactions