Description: E. coli DNA topoisomerase I was purified from E. coli strain BL21(DE3) carrying a plasmid overexpressing wildtype E. coli DNA topoisomerase I according to the procedure as described in reference 1. A 10% SDS PAGE gel shows the purity of T5 exonuclease (Figure 1). E. coli DNA topoisomerase I is stored in the storage buffer. E. coli DNA topoisomerase I is free from contaminations by other E. coli DNA topoisomerases including E. coli topoisomerase III. E. coli DNA topoisomerase I is also free from RNase and other DNase contaminations.
Unit definition: One unit of E. coli DNA topoisomerase I was defined as the amount of enzyme that catalyzes the relaxation of 0.5 μg of supercoiled plasmid DNA template pAB1 in 30 μL of 1x E. coli DNA topoisomerase I assay buffer (50 mM Tris–HCl, pH 8.0, 50 mM KOAc, 6 mM Mg(OAc)2, 1 mM DTT, and 0.1 mg/mL BSA) at 37 °C in 15 min via agarose gel-based assays and fluorescence-based assays as well.
Concentrations: Two concentrations of E. coli DNA topoisomerase I are available: 2,000 units/mL (0.6 uM or 60 ug/mL) and 20,000 units/mL (6 uM or 600 ug/mL). It is recommended that the enzyme of 2,000 units/mL should be used as is and not be diluted. Please contact us for higher concentrations of E. coli DNA topoisomerase I and DNA topoisomerase I without BSA..
Shipping and storage: E. coli DNA topoisomerase I is shipped and should be stored in a -80°C freezer (-70-80°C) and is stable for at least 12 months in this condition. E. coli DNA topoisomerase I can be aliquoted on first thawing to prevent loss of activities from freeze-thaw cycles.
Dilution buffer: 50 mM Tris-HCl, pH 8, 100 mM KCl, 2 mM DTT, 0.1 mg/mL BSA, and 10% glycerol. Dilution buffer is supplied. Dilution buffer is supplied.
10xE. coli DNA topoisomerase I assay buffer: 300 mM Tris-HCl, pH 8.0, 500 mM KAC, 60 mM Mg(AC)2, 10 mM DTT, and 1 mg/mL BSA. 10x assay buffer is supplied.
Stop solution: 250 mM EDTA and 3% SDS.
Unit definition: One unit of E. coli DNA topoisomerase I was defined as the amount of enzyme that catalyzes the relaxation of 0.5 μg of supercoiled plasmid DNA template pAB1 in 30 μL of 1x E. coli DNA topoisomerase I assay buffer (50 mM Tris–HCl, pH 8.0, 50 mM KOAc, 6 mM Mg(OAc)2, 1 mM DTT, and 0.1 mg/mL BSA) at 37 °C in 15 min via agarose gel-based assays and fluorescence-based assays as well.
Concentrations: Two concentrations of E. coli DNA topoisomerase I are available: 2,000 units/mL (0.6 uM or 60 ug/mL) and 20,000 units/mL (6 uM or 600 ug/mL). It is recommended that the enzyme of 2,000 units/mL should be used as is and not be diluted. Please contact us for higher concentrations of E. coli DNA topoisomerase I and DNA topoisomerase I without BSA..
Shipping and storage: E. coli DNA topoisomerase I is shipped and should be stored in a -80°C freezer (-70-80°C) and is stable for at least 12 months in this condition. E. coli DNA topoisomerase I can be aliquoted on first thawing to prevent loss of activities from freeze-thaw cycles.
Dilution buffer: 50 mM Tris-HCl, pH 8, 100 mM KCl, 2 mM DTT, 0.1 mg/mL BSA, and 10% glycerol. Dilution buffer is supplied. Dilution buffer is supplied.
10xE. coli DNA topoisomerase I assay buffer: 300 mM Tris-HCl, pH 8.0, 500 mM KAC, 60 mM Mg(AC)2, 10 mM DTT, and 1 mg/mL BSA. 10x assay buffer is supplied.
Stop solution: 250 mM EDTA and 3% SDS.
This product is for research only.
DNA Relaxation Assay Instruction