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TaqIT

Product Description

TaqIT is an exonuclease deficient derivative of Taq DNA polymerase. TaqIT lacks the first 280 amino acids of native Taq polymerase that contain the 5′-3′ exonuclease domain. This deletion makes TaqIT slightly more thermostable and has slightly greater fidelity than full length Taq. Like Taq polymerase, TaqIT has no inherent 3′-5′ exonuclease activity.

Source of Protein
A recombinant E. coli strain carrying the TaqIT gene from the thermophilic organism Thermus Aquaticus YT-1.

Unit Definition
1 unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid-insoluble material in 30 minutes at 75°C.

Molecular weight
62,407 Daltons

Quality Control Analysis

Unit Activity is measured using a 2-fold serial dilution method. Dilutions of enzyme were made in a reduced-glycerol (5%) containing Taq-B DNA Polymerase storage solution and added to 50 µL reactions containing Calf Thymus DNA, 25 mM TAPS (pH 9.3), 50 mM KCl, 1 mM DTT, 3H-dTTP and 100 µM dNTPs. Reactions were incubated 10 minutes at 75°C, plunged on ice, and analyzed using the method of Sambrook and Russell (Molecular Cloning, v3, 2001, pp. A8.25-A8.26).

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 protein of interest band 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 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 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 16S rDNA Contamination is evaluated using 5 µL replicate samples of enzyme solution 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.

Supplied in:
20mM Tris-HCl
222mM (NH4)2SO4
10mM 2-mercaptoethanol
0.1 mM EDTA
0.1% Brij 58
50% glycerol
pH 8.6 @ 25°C

Supplied with:
10X TaqIT Reaction Buffer (B7620):
500mM Tris-HCl
160 mM (NH4)2SO4
35mM MgCl2
0.25% Brij 58
pH 9.2 @ 25⁰C

Product Information

TaqIT
Part NumberP7620L
Price$514
Concentration50,000 U/mL
Unit Size5,000 U
SDS Available on request

Product Specification*

Storage Temperature-25⁰C to -15⁰C
Test Units Tested Specification
SDS Purityn/a>99%
Specific Activityn/a42,000 U/mg
SS Exonuclease500<1.0% Released
DS Exonuclease500<1.0% Released
DS Endonuclease500No Conversion
E.coli DNA Contamination500<10 copies

* For a detailed summary of assay conditions and data, refer to the Quality Controls Analysis section.

References

  1. The fidelity of Taq polymerase catalyzing PCR is improved by an N-terminal deletion. Barnes WM. Gene. 1992. 112(1):29-35.
  2. Comparative thermal denaturation of Thermus aquaticus and Escherichia coli type 1 DNA polymerases. Karantzeni I, Ruiz C, Liu CC, Licata VJ. Biochem J. 2003 374(Pt 3):785-92.

Limitations of Use
This product was developed, manufactured, and sold for in vitro use only. The product is not suitable for administration to humans or animals. SDS sheets relevant to this product are available upon request.

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