Mako DNA Polymerase

QIAGEN T4 DNA Polymerase (exo–)

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Mako DNA Polymerase (3ʹ→ 5ʹ exo–) (3000 U)

Cat. No. / ID: P7090L

3000 U (evaluation pack) of Mako DNA Polymerase (3'→5' exo–) (0.1 mL at 30,000 U/mL) and 10x Blue Buffer (1 x 1.5 mL)
The Mako DNA Polymerase (3ʹ→ 5ʹ exo–) (3000 U) is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Need this product modified? Contact us to get started with tailored OEM solutions.

Features

  • OEM by QIAGEN offers bulk manufacturing of Mako DNA Polymerase in custom formulations
  • QIAGEN’s version of T4 DNA Polymerase (exo –)
  • Extends DNA in the 5′→3′ direction
  • Lacks 3′→5′ and 5′→3′ exonuclease activities
  • No strand displacement function

Product Details

Mako DNA Polymerase is exonuclease deficient and lacks strand displacement activity. The enzyme catalyzes the extension of a primed DNA template in the 5ʹ→3ʹ direction. Inherent 3ʹ→5ʹ and 5ʹ→3ʹ exonuclease activities are absent and the enzyme exhibits no measurable strand displacement function.

The enzyme is supplied in 100 mM KPO4, 1.0 mM DTT, 0.1 mM EDTA and 50% glycerol; pH 6.5 at 25°C.

10x Blue Buffer (B0110) contains the following: 500 mM NaCl, 100 mM Tris-HCl, 100 mM MgCl2 and 10 mM DTT; pH 7.9 at 25°C.

Performance

Properties

  • Storage temperature: –25°C to –15°C   
  • Molecular weight: 103,565 Daltons
Test Specification
Purity >99%
Specific activity 6100 U/mg
Single-stranded exonuclease 300 U; <10.0% released
Double-stranded exonuclease 300 U; <1.0% released
Double-stranded endonuclease 300 U; no conversion
E. coli DNA contamination 300 U; <10 copies

Principle

Source of recombinant enzyme protein
The protein is produced by a strain of E. coli that expresses the recombinant Mako DNA Polymerase (3ʹ→ 5ʹ exo–) gene.

Unit definition:
One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid-precipitable material in 30 minutes at 37°C.

 

Procedure

Quality control analysis

Protein concentration is determined by OD280 absorbance.

Physical purity is evaluated by SDS-PAGE of concentrated and diluted enzyme solutions followed by silver-stain detection. Purity is assessed by comparing the aggregate mass of contaminant bands in the concentrated sample to the mass of the band corresponding to the protein of interest in the diluted sample.

Single-stranded exonuclease is determined in a 50 µl reaction containing a radiolabeled single-stranded DNA substrate and 10 µl of enzyme solution incubated for 4 hours at 37°C.

Double-stranded exonuclease activity is determined in a 50 µl reaction containing a radiolabeled double-stranded DNA substrate and 10 µl of enzyme solution incubated for 4 hours at 37°C.

Double-stranded endonuclease activity is determined in a 50 µl reaction containing 0.5 µg of plasmid DNA and 10 µl of enzyme solution incubated for 4 hours at 37°C.

E. coli contamination is evaluated using 5 µl replicate samples of enzyme solution that are denatured and screened in a TaqMan qPCR assay for the presence of contaminating E. coli genomic DNA using oligonucleotide primers corresponding to the 16S rRNA locus.

Applications

This OEM by QIAGEN product is available for bulk purchase for the following commercial assay applications.

  • Labeling
  • Partial fill-in reactions

Resources