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  PCR & DNA modifying enzymes

Taq DNA Polymerase

Enzyme is purified from an E. coli strain carrying a plasmid with Taq DNA polymerase gene from Thermus aquaticus YT-1. 

Reagents supplied: 10x Taq DNA polymerase Buffer with or w/o MgCl2 and 25mM MgCl2 

Unit definition: One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmoles of dNTPs into acid insoluble material in 30 minutes at 72ºC. 

(SKUs: PD 001S-W, PD 001S-W/O, PD 001M-W, PD 001M-W/O, PD 001L-W, PD 001L-W/O)

48.00185.00
  • 5u/μl
  • 500 U
  • 1000 U
  • 2500 U
  • with MgCl2
  • w/ο MgCl2

 

Product Details

Enzyme is purified from an E. coli strain carrying a plasmid with Taq DNA polymerase gene from Thermus aquaticus YT-1. 

Reagents supplied: 10x Taq DNA polymerase Buffer with or w/o MgCl2 and 25mM MgCl2 

Unit definition: One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmoles of dNTPs into acid insoluble material in 30 minutes at 72ºC. 

Taq DNA Polymerase is a thermostable enzyme that catalyzes 5’→3′ synthesis of DNA. The enzyme has no detectable 3’→5′ proofreading exonuclease activity, but possesses low 5’→3′ exonuclease activity.

PCR Guidelines:

PCR recommended reaction:

Components20μl assay50μl assayFinal Concentration/Quantity
10x Taq pol. buffer2μl5μl1x
dNTP mix/10mM each0.4μl1μl0.2mM each
25μΜ forward primer0.4μl1μl0.5μM
25μΜ reverse primer0.4μl1μl0.5μM
Template DNAVariableVariable1-500ng
Taq DNA pol. (5u/μl)0.1-0.2μl0.25-0.5μl1.25-2.5u / 50μl reaction
Sterile ultrapure waterUp to 20 μlUp to 50 μl 

PCR recommended conditions:

StepTemperatureTime
Initial denaturation94oC2 min
25-35 cycles  
Denaturation94oC45 sec
Annealing45-68oC*30 sec
Extension72oC1min/kb
  
Final extension72oC10 min
Hold4oCIndefinitely

*Annealing temperature depends on primer Tm

10x Taq pol. buffer: 500 mM KCl, 100 mM Tris-HCl pH 8.5 @ 25oC, 15 mM MgCl2, 1% Triton X-100

Storage conditions:

  • 100 mM NaCl
  • 50 mM Tris-HCl (pH 8.0 @ 25oC)
  • 1 mM DTT
  • 0.1 mM EDTA
  • 1% Triton X-100
  • 50% glycerol
  • Store at -20oC

Standard DNA Polymerase Assay Conditions (not PCR conditions): The polymerase activity is assayed in 10 mM Tris-HCl (pH 8.5), 50 mM KCl, 1.5 mM MgCl2, 0.1% Triton X-100, 200 μM each of dATP, dGTP, dCTP, dTTP (a mix of unlabeled and [3H] dTTP) and 12.5μg activated calf thymus DNA, in a final volume of 50 μl.

Functional Assay: Taq DNA Polymerase is tested for performance in the polymerase chain reaction (PCR) using 1.5 units of enzyme to amplify a 5000-bp region of Lambda DNA using 10ng as substrate. The resulting PCR product is visualized as a single band on an ethidium bromide-stained agarose gel.

Quality controls: Tested extensively for the absence of endo- and exodeoxyribonucleases. More details concerning quality controls could be found in Certificate of analysis.

Stability: Taq DNA polymerase is guaranteed to maintain stability for six months from the date of shipment when stored as directed.

 

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