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. Application: Second strand cDNA synthesis Nick translation Supplied with: D3935-05: DNA Polymerase I Reaction Buffer, 10X Source: E. coli, strain carrying a DNA polymerase I overproducing plasmid. Form: Supplied as a liquid in PBS, pH 7.5, 1mM DTT, 50% glycerol. Concentration: 10u/ul D3935-05, Reaction Buffer (10X): Supplied as a liquid in 500mM Tris-HCl, pH 7.5 (25C), 100mM MgCl2, 10mM DTT Unit Definition: One unit of enzyme will catalyze the exchange of 10nmole of total DNA to a DE-81 adsorbable form in 30 minutes at 37C using poly[dA-dT)] as a template primer. Quality Control: Endodeoyuribonuclease Assay 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 37C. Activity Assay: 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). Storage and Stability: May be stored at 4C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20C. Aliquots are stable for 6 months at -20C. 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.
Molecular Weight:
109
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