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Exonuclease I
Exonuclease I (Exo I) specifically degrades single-stranded DNA in the 3′→5′ direction while leaving double-stranded DNA intact. It is ideal for removing residual primers from PCR reactions and eliminating ssDNA with a free 3′-OH terminus. The enzyme is compatible with a wide range of Mg²⁺-containing buffer systems and can be heat-inactivated at 80 °C for 15 minutes.
Custom formats, package sizes and bulk pricing are available upon request.
Price range: $110.00 through $325.00
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Intact Genomics IG®’ Exonuclease I (Exo I) progressively degrades single-stranded DNA (ssDNA) in the 3′→5′ direction, releasing deoxyribonucleoside 5′-monophosphates in a stepwise manner and leaving double-stranded DNA and the 5′-terminus intact. It does not cleave double-stranded DNA.
The cleavage of single-stranded DNA is blocked when the terminal 3′-OH group is phosphorylated or acetylated. Intact Genomics’ Exonuclease I can work in a wide range of buffers from different manufacturers , as long as there is sufficient magnesium (1 mM—10 mM).
Intact Genomics’ Exonuclease I can be heat-inactivated by incubation at 80 °C for 15 minutes.
Applications and Benefits:
- PCR Cleanup – Removes residual primers while preserving dsDNA.
- Sequencing & Genotyping – Ideal for Sanger sequencing, SNP analysis, and cloning workflows.
- Selective ssDNA Removal – Specifically degrades ssDNA with a free 3′-OH end.
- Broad Buffer Compatibility – Works in many Mg²⁺-containing buffer systems.
- Heat Inactivation – Inactivated at 80 °C for 15 minutes.
- High-Throughput & OEM Ready – Suitable for automated workflows, bulk supply, and OEM applications.
Product Source:
Recombinant Exo I expressed from a plasmid carrying E.coli sbcB gene.
Specifications:
- Purified recombinant Exo I with a molecular weight of 55 kDa
- Free of detectable endonuclease activity
- ≥95% pure from host proteins
Product Components and Storage Temperature:
The following reagents are supplied with this product:
Custom formats and package sizes are available upon request.
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Quality Control:
Protein Purity is evaluated by SDS-PAGE.
Double-stranded DNA endonuclease activity is measured in a 10 μL reaction containing 0.5 μg of plasmid DNA and 0.5 μL of enzyme solution incubated for 14 hours at 37°C.
ssDNase Activity: 20 units of Exonuclease I enzyme in a 10 µL reaction digests 2 µg of ssDNA in a 30 minute reaction at 37ºC.
Definition of Activity Unit:
One unit is defined as amount of Exonuclease I that catalyzes the release of 10 nmol of acid soluble nucleotides in 30 min at 37°C from 3H-labeled single-stranded DNA substrate.
The activity is assayed in a mixture containing 67 mM glycine-KOH (pH 9.5), 6.7 mM MgCl2, 10 mM 2-Mercaptoethanol, and 10 nmol 3H-labeled single-stranded DNA.
Protocol for the Digestion of Single-Stranded DNA:
The reaction will digest ssDNA including primers.
Use the table below to set up the reaction:
Incubate the mixture at for 10 to 20 minutes 37°C. Time may be extended if desired.
Incubate for 15 minutes at 80°C to inactivate enzyme/s.
PCR products can be used for direct sequencing or for other downstream applications.
References:
Lehman IR, Nussbaum AL. The Deoxyribonucleases of Escherichia Coli. V. On The Specificit of Exonuclease I (Phosphodiesterase). J Biol Chem. 1964 AUG;239:2628-36. PMID: 14235546.
Werle E, Schneider C, Renner M, Völker M, Fiehn W. Convenient single-step, one tube purification of PCR products for direct sequencing. Nucleic Acids Res. 1994 Oct 11;22(20):4354-5. doi: 10.1093/nar/22.20.4354. PMID: 7937169; PMCID: PMC331970.
Nabavi S, Nazar RN. Simplified one-tube “megaprimer” polymerase chain reaction mutagenesis. Anal Biochem. 2005 Oct 15;345(2):346-8. doi: 10.1016/j.ab.2005.06.033. PMID: 16061194.
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