Description

Calf intestine alkaline phosphatase catalyzes the removal of 5'-phosphate groups from DNA, RNA, and ribo- and deoxyribonucleoside triphosphates. It can be used to prevent self-ligation of vectors because alkaline phosphate-treated DNA fragments lack the 5'-phosphoryl termini required by DNA ligases.

Features

  • Can be inactivated by heating at 65ºC for 30 min.

Applications

  • Removes 5' and 3' phosphoryl groups from nucleic acids
  • Prepares templates for 5' end-labeling
  • Prevents DNA fragments from self-ligating
  • Dephosphorylates proteins

Source

Calf intestine

Unit definition

One unit is defined as the amount of enzyme that will dephosphorylate 1µmole of p-Nitrophenyl phosphate with the appropriate buffer (pH 10.2) in 1 min at 37ºC.

Storage conditionn

30 mM Triethanolamine (pH7.8), 1 mM MgCl2, 50% Glycerol
Store at -20ºC

Components

Calf Intestine Alkaline Phosphatase (1-3 U/µl)
10 x Buffer

Principle

Note

This enzyme can be inactivated by heating at 65ºC for 30 min. However, the reacted DNA fragments should be purified by phenol-chloroform extraction or DNA fragment purification kits (e.g., MagExtractor –PCR & Clean up–, Code No. NPK-601) prior to ligation with caution.

References

  • H.N.Ferunley, The Enzymes (3rd Ed.), 4: 417-447(1971)
  • R.K. Morton, The substrate specificity and inhibition of alkaline phosphatases of cow's milk and calf intestinal mucosa. Biochem J. 61: 232- 40 (1955)
  • F.Dray, E.Dith and C.Rougeot, Methods of Enzymatic Analysis, 9, 348 (1986)
  • P.Rathman and B.B.Saxena, Methods of Enzymatic Analysis, 9, 396 (1986)