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Klenow Fragment, Exonuclease Minus

by Promega
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  • DNA Polymerase I Large (Klenow) Fragment, Exonuclease Minus, is a DNA-dependent DNA polymerase that lacks both the 5´?3´ and the 3´?5´ exonuclease activities present in intact E. coli DNA Polymerase I. Klenow Fragment, Exo Minus, is used for random primer labeling, DNA sequencing by the dideoxy method, and in strand displacement amplification (SDA). This enzyme will leave a single-base 3´ overhang on a significant proportion of DNA fragments during fill-in of 5´-overhangs. Therefore, Klenow Fragment, Exonuclease Minus, is not recommended for preparation of blunt-ended fragments for ligation.

    References

    1. Derbyshire, V. et al. (1988) Science 240, 199–201.
    2. Sanger, F. et al. (1977) Proc. Natl. Acad. Sci. USA 74, 5463–7.
    3. Feinberg, A.P. and Vogelstein, B. (1984) Anal. Biochem. 137, 266–7.
    4. Walker, G.T. (1993) PCR Meth. Appl. 3, 1–6.
    5. Clark, J.M. et al. (1987) J. Mol. Biol. 198, 123–7.

    Storage Buffer: 50mM Tris-HCl (pH 7.5 at 25°C), 1mM DTT, 0.1mM EDTA and 50% (v/v) glycerol.

    Source: Recombinant E. coli strain.

    QC Tests: Activity, SDS-PAGE/purity, endonuclease, DNase, RNase.

    Unit Definition: One unit is defined as the amount of enzyme that incorporates 10nmol of total deoxyribonucleotides into TCA-insoluble material in 30 minutes at 37°C. The reaction conditions are: 67mM potassium phosphate (pH 7.4 at 25°C), 6.7mM MgCl2, 1mM DTT, 50ug/ml activated calf thymus DNA, and 33uM dATP, dCTP, dGTP and dTTP (a mix of unlabeled and [3H]dTTP).

  • Protocols

    Complete Protocol

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    DNA Polymerase I Large (Klenow) Fragment, Exonuclease Minus Protocol

    PDF (112 KB)

  • Certificate of Analysis

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    Storage Conditions

    -30C TO -10C

    Use Restrictions

    For Research Use Only. Not for Use in Diagnostic Procedures.