DNA Polymerase I
|Cloning||Storage: -20°CShipping: Blue Ice|
DNA Polymerase I catalyzes the polymerization of nucleotides into duplex DNA in the 5’=>3’ direction. The enzyme exhibits 3’-5’ proofreading exonuclease activity and 5’=>3’ degrading activity. Purified from an E.coli strain carrying a DNA polymerase I overproducing plasmid.
• Second strand cDNA synthesis
• Nick translation
D3935-05: DNA Polymerase I Reaction Buffer, 10X
Storage and Stability:
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for 6 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
E. coli, strain carrying a DNA polymerase I overproducing plasmid.
Supplied as a liquid in PBS, pH 7.5, 1mM DTT, 50% glycerol.
D3935-05, Reaction Buffer (10X): Supplied as a liquid in 500mM Tris-HCl, pH 7.5 (25°C), 100mM MgCl2, 10mM DTT
One unit of enzyme will catalyze the exchange of 10nmole of total DNA to a DE-81 adsorbable form in 30 minutes at 37°C using poly[dA-dT)] as a template primer.
No detectable conversion of covalently closed circular DNA to nicked DNA was observed after incubation of 20u of DNA Ploymerase with 1ug of pBR322 DNA in 50ul of reaction buffer for 4 hours at 37°C.
67mM potassium phosphate (pH 7.4), 6.7mM MgCl2, 1mM 2-mercaptoethanol, 0.033mM dATP, 0.033mM dTTP, 0.4MBq/ml [3H]-dTTP and 62.5ug/ml poly(dA-dT)•poly(dA-dT).
Important Note: This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.
1. W. S. Kelly and K. H. Stump, J. Biol. Chem., 254: 3206 (1968). 2. T. Maniatis, et al., Proc. Natl. Acad. Sci. USA, 72: 1184 (1975). 3. N. E. Murray and W. S. Kelly, Molec. Gen Genet., 175: 77 (1979). 4. Gubler, U., Hoffmann, B.J., Gene 25: 263-2369 (1983). 5. Current Protocols in Molecular Biology, vol1 (Ausubel, F.M., et al., ed.), John Wiley and Sons, Inc., Brooklyn, NY, 3.5.3-3.5.6, (1994-2001).