Paris/October 12
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< Paris(Difference between revisions)
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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> | ||
| Line 12: | Line 14: | ||
- 0,5µL Phusion Taq<br> | - 0,5µL Phusion Taq<br> | ||
------ | ------ | ||
| - | - 2' 98°C<br> | + | Timing : <br><br> |
| - | <br> | + | First :<br> |
| - | + | - 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
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
