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.
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.
|Storage Temperature||-25⁰C to -15⁰C|
|Specific Activity||n/a||42,000 U/mg|
|SS Exonuclease||500||<1.0% Released|
|DS Exonuclease||500||<1.0% Released|
|DS Endonuclease||500||No Conversion|
|E.coli DNA Contamination||500||<10 copies|
* For a detailed summary of assay conditions and data, refer to the Quality Controls Analysis section.
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.
0.1 mM EDTA
0.1% Brij 58
pH 8.6 @ 25°C
10X TaqIT Reaction Buffer (B7620):
160 mM (NH4)2SO4
0.25% Brij 58
pH 9.2 @ 25⁰C
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.