Quantitative determination of a DNA polymerase and a test kit useful in same

A convenient method for the quantitative determination of a DNA polymerase includes contacting an aqueous test specimen suspected of containing the enzyme with the following: a single-stranded DNA template present in a concentration of at least 10 molar bases, a DNA primer complementary to the templ...

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Hauptverfasser: SHERIDAN, PATRICK JAMES, MEZEI, LOUIS MICHAEL, SUTHERLAND, JOHN W. H
Format: Patent
Sprache:eng ; fre ; ger
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Zusammenfassung:A convenient method for the quantitative determination of a DNA polymerase includes contacting an aqueous test specimen suspected of containing the enzyme with the following: a single-stranded DNA template present in a concentration of at least 10 molar bases, a DNA primer complementary to the template, a source of a metal polymerase cofactor, sufficient deoxyribonucleoside triphosphates to synthesize double-stranded DNA in the presence of the polymerase, and a colorimetric or fluorescent dye which is capable of providing a detectable signal when a primed single-stranded nucleic acid is converted to double-stranded DNA by the polymerase. The rate of signal generation is then measured and can be correlated with the level of DNA polymerase in the specimen using graphical or mathematical means. The results of this method are precise, having a covariance of less than 10%. A test kit includes the reagents needed for carrying out this method.