Team:Tuebingen/Experiments

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Protocols

Prep

Colony-PCR

3A-Assembly

Restriction

Transformation

PCR

Ligation

Testrestriktion

Gel extraction

DNA Purification using Promega Wizard SV Gel and PCR Clean-Up System Kit

This is a modified protocol for the Wizard SV Gel and PCR Clean-Up System Kit from Promega

  • Following electrophoresis, excise DNA band from gel and place gel slice in a 1,5ml microcentrifuge tube.
  • Add 10μl Membrane Binding Solution per 10mg of gel slice. Vortex and incubate at 50-60°C until gel slice is completely dissolved.
  • Insert SV Minicolumn into Collection Tube.
  • Transfer dissolved gel mixture to the Minicolumn assembly. Incubate at room temp. for 1 minute.
  • Centrifuge at 16,000xg for 1 min. Discard flowthrough and reinsert Minicolumn into Collection tube.
  • Add 700μl Membrane Wash Solution (ethanol added). Centrifuge at 16,000xg for 1 min. Discard flowthrough and reinsert Minicolumn into Collection tube.
  • Repeat this step with 500μl Membrane Wash Solution. Centrifuge at 16,000xg for 5 min.
  • Empty the Collection Tube and recentrifuge the column assembly for 1,5 min.
  • Leave the tubes open for 10 min (to let any rest of ethanol evaporate).
  • Carefully transfer Minicolumn to a clean 1.5ml microcentrifuge tube.
  • Add 30μl of Nuclease-Free-Water (65°C) to the Minicolumn. Incubate at 65°C for 5 min. Centrifuge at 16,000xg for 1 min.
  • Discard Minicolumn and store DNA at 4°C or -20°C.

Experiments & Protocols

Describe the experiments, research and protocols you used in your iGEM project.

What should this page contain?
  • Protocols
  • Experiments
  • Documentation of the development of your project

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