• MANUAL
• SDS：BTQ-101
• SDS：BTQ-201

DESCRIPTION

• Blend Taq™ and Blend Taq™ -Plus- are highly efficient Taq-based DNA polymerases developed based on the Barns' method(1). This method uses a DNA polymerase which lacked 3'→5' exonuclease (proofreading) activity (e.g., Taq DNA polymerase) and a small amount of an archaeal DNA polymerase with proofreading activity. Because the proofreading activity repairs misincorporated nucleotide bases causing the termination of the polymerase reaction, PCR with a 'mixed' enzyme solution enables highly efficient amplification.

  The enzyme solution of Blend Taq™ -Plus- contains anti-Taq DNA polymerase antibodies that inhibit polymerase activity, allowing for Hot Start PCR.

  Blend Taq™ and Blend Taq™ -Plus- generate dA overhang-ended PCR products. Therefore, the PCR products can be cloned using a standard TA-cloning method.

• Features
• Details
• Application Data

Features Details Application Data

Features

• ・Effective for the amplification of various targets from small starting template amounts.
• ・The use of Hot Start technology with anti-Taq DNA polymerase antibodies results in highly efficient amplification. <Blend Taq™ (Code No. BTQ-101) does not use hot start>
• ・The PCR error ratio of this enzyme is approximately 3-4 times less than that of Taq DNA polymerase.

Details

APPLICATIONS

• PCR

SOURCE

E. coli strain carrying the cloned Taq DNA polymerase gene

UNIT DEFINITION

One unit is defined as the amount of enzyme that will incorporate 10 nmoles of dNTP into an acid insoluble material in 30 min at 75ºC.

STORAGE CONDITION

20 mM Tris-HCl (pH8.0), 0.1 mM EDTA, 100 mM KCl, 1 mM DTT, 0.5 % Tween 20, 0.5% Nonidet P-40, 50% Glycerol Store at -20ºC

COMPONENTS

The provided reagents include the following components for 200 reactions:

<Blend Taq™>
Blend Taq™ (2.5U/µL)	100 µL
10× PCR Buffer for Blend Taq™	1.0 mL
2 mM dNTPs	1.0 mL

<Blend Taq™ -Plus->
Blend Taq™ -Plus-(2.5U/µL)*	100 µL
10× PCR Buffer for Blend Taq™	1.0 mL
2 mM dNTPs	1.0 mL

*This enzyme solution contains anti-Taq DNA polymerase antibodies.

PRINCIPLE

TYPICAL PCR REACTION SETUP

Component	Volume	Final Concentration
10x Buffer	5 µL	1×
2 mM dNTPs*	5 µL	0.2 µM each
10 pmol/mL Primer #1	1 µL	0.2 µM
10 pmol/mL Primer #2	1 µL	0.2 µM
Template DNA	X µL	Genomic DNA 10-1000 ng/50 µL Plasmid DNA 1-50 ng/50 µL cDNA ~200 ng （RNA equiv.)/50 µL
PCR grade water	Y µL
Blend Taq™ (2.5 U/mL) or Blend Taq™ -Plus (2.5 U/mL)	1 µL	1.25 U / 50 µL
Total reaction volume	50 µL

* Do not use dNTPs from other kits or companies.

PCR CYCLE CONDITIONS

*For colony-direct PCR, the pre-denaturation step should be set for 4 min.
**Extension time should be set at 1min per 1 kb of target length.

Application Data

Example 1.Amplification of long targets

The PCR performances of Blend Taq™ and Blend Taq™ -Plus- were evaluated by amplification of long targets. Blend Taq™ and Blend Taq™ -Plus- showed distinct bands whereas PCR enzymes from other companies showed poor bands or smears. No non-specific band was detected by Blend Taq™ -Plus-.

Template

Human genomic DNA

Target

β-globin gene

PCR cycling condition

M: 1 kb Ladder Marker
1: β-globin 1.3 kb
2: β-globin 2.7 kb
3: β-globin 3.6 kb
4: β-globin 17.5 kb
5: β-globin 23.0 kb

Example 2.Colony-direct PCR using E. coli cells

The sensitivities of Blend Taq™ and Blend Taq™ -Plus- were compared with those of other companies' high efficient Taq DNA polymerases. Blend Taq™ and Blend Taq™ -Plus- showed greater sensitivities than enzymes from other companies. The sensitivity of Blend Taq™ -Plus- was slightly greater than that of Blend Taq™.

Template

Human genomic DNA

Target

β-globin gene

PCR cycling condition

1: Negative control
2: Human genomic DNA 5 ng
3: Human genomic DNA 10 ng
4: Human genomic DNA 20 ng
5: Human genomic DNA 40 ng
M: 1 kb Ladder Marker

REFERENCES

• W.M. Barns, PCR amplification of up to 35-kb DNA with high fidelity and high yield from lambda bacteriophage
  templates.Proc. Natl. Acad. Sci. USA, 91: 2216-2220 (1994)