From 2007.igem.org

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.

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

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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