Team:UNIPV-Pavia/Protocols/AntarcticPhosphatase

From 2008.igem.org

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*'''Cut and gel-extracted vector'''
*'''Cut and gel-extracted vector'''
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*Add the proper amount of 10X buffer to a final concentration of 1X (e.g. 2 )
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*Add the proper amount of 10X buffer to a final concentration of 1X (e.g. 2 µl of 10X buffer in a final volume of 20 µl).
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*Cut paper spots using scalpel and tweezers (or punch tool, following provided instructions); if you use scalpel and tweezers, try to cut pieces of about the same dimension of the punch tool.
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*Add 1 µl of Antarctic Phosphatase (up to 5 µg of cut vector).
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*Put the cut paper into the 0.5 ml tube.
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*Incubate at 37°C for 1 hour (Antarctic Phosphatase works).
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*Clean scalpel and tweezers (or punch tool) with water and ethanol every time you cut a spot; be careful to dry your tools correctly, especially if you use punch tool, which needs much more time to dry than scalpel/tweezers.
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*Incubate at 65°C for 15 min (Antarctic Phosphatase inactivation).
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*Incubate at 42°C for 20 min.
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*Vortex and spin down.
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Revision as of 09:29, 11 June 2008

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The protocols we used


Antarctic Phosphatase

(estimated time: 1 hour and 30 min)

Materials needed:

  • NEB Antarctic Phosphatase
  • 10X NEB Antarctic Phosphatase buffer
  • Cut and gel-extracted vector


  • Add the proper amount of 10X buffer to a final concentration of 1X (e.g. 2 µl of 10X buffer in a final volume of 20 µl).
  • Add 1 µl of Antarctic Phosphatase (up to 5 µg of cut vector).
  • Incubate at 37°C for 1 hour (Antarctic Phosphatase works).
  • Incubate at 65°C for 15 min (Antarctic Phosphatase inactivation).


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