Enzymatic method for continuous monitoring of DNA polymerase activity.
A simple and rapid method for the assay of DNA polymerase activity has been developed. The PPi formation in the DNA polymerase reaction is continuously monitored by a coupled enzymatic method (P. Nyrén and A. Lundin, 1985, Anal. Biochem. 151, 504-509) utilizing the enzymes ATP-sulfurylase and firefly luciferase. The method has been used for continuous monitoring of DNA synthesis in vitro, and the effect of an inhibitor, adriamycin, on the polymerase activity was studied. The assay is very sensitive and yields linear responses between 1.5 and 30 micrograms/ml of DNA polymerase (Micrococcus luteus) (2-40 pmol of PPi generated per minute).
Nyrén
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Department of Biochemistry, Arrhenius Laboratory, University of Stockholm, Sweden.
D004247:DNA / Q000096:biosynthesis; D004259:DNA-Directed DNA Polymerase / Q000032:analysis*; D011756:Diphosphates / Q000096:biosynthesis; D004317:Doxorubicin / Q000494:pharmacology; D066298:In Vitro Techniques; D008156:Luciferases; D008722:Methods; D019384:Nucleic Acid Synthesis Inhibitors; D013430:Sulfate Adenylyltransferase
D011756:Diphosphates; D019384:Nucleic Acid Synthesis Inhibitors; D004317:Doxorubicin; D004247:DNA; D008156:Luciferases; D013430:Sulfate Adenylyltransferase; D004259:DNA-Directed DNA Polymerase