DNAポリメラーゼI, クレノウフラグメント 大腸菌由来

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.

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

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.

活性アッセイ:50mMリン酸カリウム, pH 7.5, 3mM MgCl₂, 1mM 2-メルカプトエタノ-ル, 32.5μM³²P-dATP, 32.5μM dTTP, 62.5μg/mLポリ(dA-dT), 0.01-1unit酵素。

1 unitは、37°C、30分間に、10 nmolのデオキシリボヌクレオシド三リン酸を酸性不溶性物質に変換する酵素量です。

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) .