User:Karmella Haynes/Notebook/BioBrick cloning/2013/03/19

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03/19/13

  • Oligo annealing for new mammalian vector



Oligo annealing

  • Note: made fresh 10x annealing buffer
  1. KAH3/MV2(X) + K-XbaI-NLS-his-stop
  • K-XbaI-NLS-6his-st 1
  • K-XbaI-NLS-6his-st 2
  • K-XbaI-NLS-6his-st 3
  • K-XbaI-NLS-6his-st 4
DNA (oligos, 100 μM) 4x 3 μL ea. = 12.0
10x annealing buffer 2.0
dH2O 6.0
  20 μL

Thermal Cycling (try new thermal cycling approach)
Multigene Optimax Thermal Cycler

  • 99.9 °C, 10 min.
  • [99.9°C, temp. increment -2.0°C, 1 min.; 90.0°C, temp. increment -2.0°C, 1 min.] x35
  • 25°C, ∞



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