IGEM:Harvard/2007/Protocols/Vector Dephosphorylation Protocol

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  1. Add 1/10 volume of 10X Antarctic Phosphatase Reaction Buffer to 1 µg of DNA cut with any restriction endonuclease in any buffer.
  2. Add 1 µl of Antarctic Phosphatase (5 units) and mix.
  3. Incubate for 15 minutes at 37°C for 5´ extensions or blunt-ends, 60 minutes for 3´ extensions.
  4. Heat inactivate (or as required to inactivate the restriction enzyme) for 5 minutes at 65°C.
  5. Proceed with ligation.
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