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Team:TU Darmstadt/protocols/Dephosphorylation
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Revision as of 12:35, 4 October 2013
Dephosphorylation
Short explanation
Antarctic Phosphatase catalyzes the removal of 5' phosphate groups of DNA and RNA and thus prevents religation of cut vectors. It is used before ligation.
Procedure
The reaction mix contains:
up to 5 µg of digested DNA (no purification needed)
1/10 of reaction volume of 10x Antarctic Phosphatase Reaction Buffer
1 µL of Antarctic Phosphatase
Incubate at 37° C for 30 minutes (3' ends) or 60 minutes (5' ends)
Heat inactivate at 65° C for 15 minutes
Continue with ligation
