Taq DNA Polymerase, 5 u/µl

Applications


  • Routine PCR up to 5 kb under common and fast conditions

  • RT-PCR, colony PCR,TA cloning

  • Library construction, genotyping, screening

Benefits


  • High yields in routine PCR and under fast cycling conditions

  • Robust performance under variety of conditions

  • Excellent performance on complex (GC rich) templates

Catalog. # Product Name Size Price Qty
PCE0201 Taq DNA Polymerase, 5 u/µl 1500 u
€249.50
PCE0202 Taq DNA Polymerase, 5 u/µl 3000 u
€449.50

Taq DNA Polymerase, 5 u/µl

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Description

highQu Taq DNA Polymerase is the classical enzyme for routine PCR applications providing high amplification yields of 3-5 kb targets under various conditions. Taq DNA Polymerase is purified from a recombinant E. coli strain carrying the Taq DNA polymerase gene.

Taq DNA polymerase is thermostable 5´ → 3´ DNA polymerase. It lacks 3´ → 5´ exonuclease (proofreading) activity and has low 5´ → 3´ exonuclease activity. Polymerase exhibits deoxynucleotidyl transferase activity resulting in A-overhang at the 3'-ends of PCR products and allowing for TA cloning. The PCR accuracy of Taq DNA Polymerase is 4.5 x 104 (a number of incorporated nucleotides before the first error occurs).

The supplied reaction buffer for most flexibility in optimization, contains no dNTPs and no magnesium. The 50 mM MgCl2 solution is provided separately what allows for magnesium optimization starting from 1 mM up to 4 mM final concentration upon the need. The dNTPs in sets or mixes can be purchased from highQu separately.

Protocols

Print version of the protocol: Product Insert Taq DNA Polymerase, 5 u/ul
Important Notes
  • Take typical measures to prevent PCR cross over contamination, keep your bench clean, wear gloves, use sterile tubes and filter pipet tips.
  • Include a no-template control and positive control in parallel.
  • Thaw and keep reagents on ice. Mix well before use.
  • The longer the amplicon, the longer the extension time. Taq DNA Polymerase speed is ~ 2000 nucleotides/min, so 15-90 seconds of extension can be provided per cycle, depending on amplicon size.
  • Start annealing with 55°C and perform gradient by increasing temperature in 2°C up to 65°C to choose the best. Calculate primer annealing temperature using software.
 
Use of magnesium and dNTPs
  • Typical concentration of each dNTP in the reaction is 0.2 – 0.25 mM. Higher concentration increase yields, however Mg2+-ions  bind to dNTPs, therefore, both components shall be present in coordinated concentrations. Too high dNTPs and magnesium concentrations reduce PCR fidelity.
  • Mix well each dNTP and magnesium solution.
  • Use final 3 mM MgCl2 and 0.25 mM each dNTP concentrations for routine PCR.

 

Starting dNTP Mix conc. Vol. of dNTP mix in 50 µl r.   Final Mg2+ conc. in r. Vol. of 50 mM MgCl2 in 50 µl rxn to achieve the desired conc.
10 mM 1.25 µl   2 mM 2 µl
25 mM 0.5 µl   3 mM 3 µl
 
Prepare a 50 µl PCR reaction

 

Rev. & For. Primers 0.1-0.5 µM final (1-2 µl of 10 µM each)
cDNA Template  or
gDNA Template
<100 ng  or
5 - 500 ng
10X PCR Buffer 5 µl
dNTPMix 0.25 mM final (1.25 µl of 10 mM dNTP Mix or 0.5 µl of 25 mM dNTP Mix)
50 mM MgCl2 3 μl
PCR Water to 49 μl
Taq DNA  Polymerase, 5 u/µl 0.25 - 1 µl

 

  • Mix gently, avoid bubbles.
  • Place into the instrument set like:

 

Initial denaturation 1 cycle: 95°C – 60 sec
Denaturation 30-40 cycles: 95°C - 15 sec
Annealing 30-40 cycles: 55-65°C – 15 sec
Extension 30-40 cycles: 72°C – 15-90 sec
Final extension 1 cycle: 72°C – 5 min (for TA cloning)

 

  • After the PCR store probes on ice shortly, store at -20°C for long term.

 

Specifications

Cat. Size Components Composition
PCE0201 1500 u 1500 u - Taq DNA Polymerase, 5 u/µl
4 x 2 ml - 10X PCR Buffer
2 x 2 ml - 50 mM MgCl2
Enzyme in storage buffer.
10X PCR Buffer contains enhancers and stabilizers, but no dNTPs and no Mg2+.
PCE0202 3000 u 2 x 1500 u - Taq DNA Polymerase, 5 u/µl
8 x 2 ml - 10X PCR Buffer
4 x 2 ml - 50 mM MgCl2
Enzyme  in storage buffer.
10X PCR Buffer contains enhancers and stabilizers, but no dNTPs and no Mg2+.

Storage

In the dark at -20°C.

Resources

Product Insert