DNA POLYMERASE I: FAQS

DNA Polymerase I is a mesophilic E. coli polymerase that catalyzes 5’→3′ template-directed DNA synthesis.

One unit of DNA Polymerase I is defined as the amount of enzyme that will incorporate 300 nmol of dNTPs into a DNA template in 60 minutes at 37°C.

• Second-strand synthesis
• Nick translation

• 25 mM Tris-HCl, pH 7.4
• 0.1 mM EDTA
• 50% Glycerol
• 1 mM DTT

DNA Polymerase I performs well in a range of buffers. The recommended 10X buffer composition is a standard reaction buffer. This buffer should be optimized for specific applications.

Industry standard 10X DNA Polymerase I reaction buffer (not supplied with kit):

• 500 mM Tris-HCl, pH 7.9 at 25°C
• 100 mM MgCl₂
• 10 mM DTT

Please reference our technical guide for instructions for use.

• Depending on the application, add DNA Polymerase I at 0.25 – 1.0 U/μL and dNTPs at 25 – 100 μM final concentration in 1X DNA Polymerase I reaction buffer
• Incubate at 37°C for 30 minutes to 20 hours
• Stop the reaction by heating at 75°C for 20 minutes to inactivate the enzyme, or alternatively stop the reaction by adding an equal volume of 0.25M EDTA pH 8.0

No, reaction buffer and dNTPs are not supplied with the kit.

The optimal temperature for DNA Polymerase I activity is 37°C.

DNA Polymerase I is inactivated by heating at 75°C for 20 minutes, or through the addition of an equal volume of 0.25M EDTA pH 8.0.

Yes, DNA Polymerase I can be used to generate blunt ends. Its 3’→5′ activity can remove 3′ overhangs and the polymerase activity will fill in 5′ overhangs.

The extinction coefficient for DNA Polymerase is 0.837. If measuring protein concentration (A280), we recommend you use the protein’s extinction coefficient to ensure an accurate reading.

Yes.

Product 7K0113-250UL is our drop-in replacement for NEB DNA Polymerase I.
Product 3K0084-250UL is our drop-in replacement for QIAGEN DNA Polymerase I.

For ordering, please Request a Quote or contact sales@watchmakergenomics.com.
Please reach out to support@watchmakergenomics.com for any other drop-in replacement inquiries.

Yes! Watchmaker offers customer fills, packaging, concentrations, and labeling – including private label – designed to meet your unique needs. We offer flexible terms to serve organizations of any size, and our right-sized processes enable rapid turnaround time on customization. Please contact sales@watchmakergenomics.com to learn more about our capabilities.

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