User:Jorge E. Buendia Buendia/Notebook/iGEM UNAM-Genomics-Mexico/2010/08/19

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August 19th, 2010

1. Make PCR to amplify BluePromoter to re-do ligation L4: Backbone pSB3K3 + Blue Promoter BBa_K238013 + GFP BBa_K145015 (74 min). As control I used primers to amplify OGR gene.

  • Reactive (ul x sample)
Taq Polymerase -> 1
Taq Reaction Buffer -> 5
MgCl 50mM (can be used up to 3ul) -> 2.5
dNTP’s 0.4ug/ul -> 2.5
Primer Forward (can be used up to 3ul) -> 2.5
Primer Reverse (can be used up to 3ul) -> 2.5
HPLC -> 32
DNA -> 2
Total volume -> 50
  • Thermocycler program:
1. 95ºC 5 min
2. 35 cycles
  • 95ºC 45 seg
  • 55ºC 45 seg
  • 72ºC 1 min
3. 72ºC 10 min
4. Hold 4ºC