Affymetrix™ Exonuclease I

Useful in preparing PCR products for applications involving sequencing or labeling methods

Overview
Brand: Affymetrix USB

Manufacturer Part Number: 70073Z 2500 UN

2500UNIT Exonuclease I, Standard Conc.: 10 units/ul

Code: NEW

Additional Details:
Additional Details: Weight: 0.47500kg



Product Code. 15513677

Quantity Price
1 £ 63.9 / 2500 units
Estimated Shipment date
from Supplier 15-12-2016
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Description and Specification

Specification

Product Name Exonuclease I
Concentration 10U/μL
Quantity 2,500U

Exonuclease I is particularly useful in preparing the products of PCR for applications involving sequencing or labeling methods1. Typically, the excess primers and any other extraneous single-stranded DNA present in PCR products will interfere with subsequent enzymatic reactions involving DNA synthesis. The hydrolytic properties of Exonuclease I degrade all single-stranded DNA present in the PCR mixture allowing the product to be used more efficiently in other applications.

  • Hydrolyzes single-stranded DNA in the 3'-5' direction, releasing 5'-mononucleotides and leaving the terminal 5'-dinucleotide intact; hydrolysis is processive and cannot proceed if the 3' terminus is phosphorylated
  • Use to measure the endonucleolytic cleavage of covalently closed circular single-stranded DNA reacted with an endonuclease of interest2
  • Also suitable for measuring DNA helicase activity3
  • When combined with Shrimp Alkaline Phosphatase (Mfr. No. 78390) for dNTP dephosphorylation, the use of alternative purification methods, such as columns, gels or magnetic separations, are completely eliminated

Source: E. coli strain containing an overproducing clone of E. coli Exonuclease I
Properties:

  • Molecular Weight: 55 kDa
  • Heat Inactivation: 80° for 15 min.
  • Degrades to terminal dinucleotides
  • Degrades glycosylated DNA
  • Optimum Temperature: 37°C
Purity:
  • Greater than 95% pure as determined by SDS-PAGE
  • Tested for contaminating endonucleases, double-stranded exonucleases and ribonucleases
Storage Buffer:
20mM Tris-HCI (pH 7.5), 5mM 2-mercaptoethanol, 0.5mM EDTA, 50% glycerol
Assay Conditions:
  • Reaction mixture (100μL) contains 67mM glycine buffer (pH 9.5), 10mM
    2-mercaptoethanol, 6.7mM MgCl2, 0.5mM denatured DNA and enzyme
  • Incubation is at 37°C for 30 min.
  • Unit Definition: One unit is the amount of enzyme which catalyzes the release of 10nmol of acid-soluble nucleotide from denatured DNA in 30 min at 37°C under standard conditions
    Concentration:
    • Standard Conc.: 10 units/μL, Mfr. No. 70073
    • High Conc.: 20 units/μL, Mfr. No. 72073
    Functional Assay: Treated PCR product with Exonuclease I to degrade unincorporated primers before performing sequencing reaction with Sequenase™ Version 2.0 DNA Polymerase Sequencing Kit (Mfr. No. 70770)

    Elimination of residual single-stranded DNA containing a 3' terminus; Measuring endonucleolytic cleavage of covalently closed circular ssDNA; Measuring DNA helicase activity