Paris/October 12
From 2007.igem.org
(Difference between revisions)
| Line 4: | Line 4: | ||
<br> | <br> | ||
------ | ------ | ||
| + | Mi PCR :<br> | ||
- 27µL H2O<br> | - 27µL H2O<br> | ||
- 10µL Buffer10X GCRich (Finnzyme)<br> | - 10µL Buffer10X GCRich (Finnzyme)<br> | ||
| Line 12: | Line 13: | ||
- 0,5µL Phusion Taq<br> | - 0,5µL Phusion Taq<br> | ||
------ | ------ | ||
| + | Timing : <br> | ||
| + | First :<br> | ||
- 2' at 98°C<br> | - 2' at 98°C<br> | ||
| - | |||
* 30X :<br> | * 30X :<br> | ||
- 20" at 98°C<br> | - 20" at 98°C<br> | ||
Revision as of 23:37, 14 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.
Mi 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
