DNA Polymerase I, Klenow Fragment from Escherichia coli, buffered aqueous glycerol solution

DNA Polymerase I, Klenow Fragment from Escherichia coli, buffered aqueous glycerol solution

Stock Code: 3577272
Manufacturer Part No: D8276-100UN
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Analysis Note


The activity is assayed in a reaction mixture containing 50 mM potassium phosphate (pH 7.5), 3 mM MgCl2, 1 mM 2-mercaptoethanol, 32.5 µM 32P-dATP, 32.5 µM dTTP, 62.5 µg/ml poly(dA-dT) and 0.01-1 unit enzyme.


Application


Suitable for: DNA sequencing by the Sanger dideoxy method Synthesis of the complementary strand of cDNA Filling in 5′-overhangs in double stranded DNA to form blunt ends Mutagenesis of DNA with second strand synthesis using oligonucleotides Labeling DNA by the random primer method


Components


DNA Polymerase I is supplied as a solution in 50 mM Tris-HCl (pH 7.5), 1 mM EDTA, 5 mM dithiothretol, and 50% glycerol (v/v) .


General description


DNA polymerase I yields two fragments (small and large) upon protease digestion. The large fragment (Klenow fragment) loses the 5′ exonuclease activity that is present in the intact holoenzyme. However, it retains both the polymerase 5′→3′ activity and the 3′→5′ exonuclease activity of the native enzyme.


Reconstitution


The enzyme solution may be diluted with 50 mM Tris-HCl, pH 7.5, 100 mM ammonium sulfate, 10 mM 2-mercaptoethanol, and 1 mg/ml bovine serum albumin.


Unit Definition


One unit converts 10 nanomoles of deoxyribonucleoside triphosphates into acid insoluble material in 30 min. at 37?°C.

Quality Level200
ManufacturerSIGMA-ALDRICH
Formbuffered aqueous glycerol solution

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