Medino® Taq DNA Polymerase

Product nameMedino® Taq DNA Polymerase
Cat.No. LM203002
Size1,000 U/10,000 U
Storage and shippingThe product is shipped with ice pack and can be stored at -25℃ ~ -15 ℃ for  2 year.

Description

Product description

Medino® Taq DNA Polymerase is a thermostable recombinant DNA polymerase expressed from the thermophilic bacterium Thermus aquaticus, with a molecular weight of 94 kDa. It possesses 5’→3′ polymerase activity and 5’→3′ exonuclease activity, but lacks 3’→5′ exonuclease activity. The amplification products have a 3′-dA overhang, making them suitable for direct use in TA cloning.

 

Product Components:

Components No.Components
10101-A10×Taq Buffer (Mg2+ Free)
10101-B25 mM MgCl2
10101-CMedino® Taq DNA Polymerase (5 U/μL)

 

 Product Applications:

  • Genotyping
  • Colony PCR
  • Other routine PCR applications

 

Definition of Activity:

One unit (U) of activity is defined as the amount of enzyme that incorporates 10 nmol of total nucleotides into acid-insoluble material within 30 minutes at 74°C, using activated salmon sperm DNA as the template/primer.

 

Quality Control:

  • Nuclease Residual Testing (Exonuclease): Incubate 10 U of this product with 0.5 μg of λDNA-Hind III at 37°C for 4 hours. The DNA electrophoresis pattern should show no change.
  • Nicking Activity Testing: Incubate 10 U of this product with 0.5 μg of IL23R plasmid at 37°C for 4 hours. The DNA electrophoresis pattern should show no change.
  • RNase Residual Testing: Incubate 10 U of this product with 0.5 μg of total RNA from 293T cells at 37°C for 1 hour. The RNA electrophoresis pattern should show no change.

 

Instructions

  1. Recommended PCR Reaction System(Preparation on ice)
Components Volume μLFinal concentration
ddH2Oto 50
10×Taq Buffer (Mg2+ Free)5
25 mM MgCl231.5 mM
dNTP Mix (10 mM each)1 μL0.2 mM
Template DNA:optional
Primer 1(10 μM)2 μL0.4 μM
Primer 2(10 μM)2 μL0.4 μM
Medino® Taq DNA Polymerase (5 U/μL)0.4 μL 0.04 U/μL

[Note]:

  • Final Concentration of Mg²⁺:

The optimal Mg²⁺ concentration is between 1.5-2 mM. If necessary, you can adjust the concentration in increments of 0.2-0.5 mM to find the optimal concentration.

  • Polymerase Addition:

Taq polymerase has some 5’→3′ polymerase activity at room temperature. To prevent nonspecific amplification, it is recommended to add the polymerase as the final step in the reaction setup.

  • Polymerase Concentration:

The recommended concentration is 0.04 U/μL. Optimization can be performed within the range of 0.025-0.04 U/μL.

  • Recommended Template Amounts for a 50 μL Reaction:
Template TypesAmplification Fragments
Genomic DNA50 ng-100 ng
Plasmid DNA10 pg-20 ng
cDNA1-5 μL (not exceeding 1/10 of the reaction volume)

 

2.Reaction program

Cycle stepTemp.TimeCycles
Initial denaturation94 °C30 sec -5 min1
Denaturation94 °C30 sec35
Annealing50 – 60 °C30 sec
Extension72 °C60 sec/kb
Final extension72 °C10 min1

[Note]:

  1. Denaturation Temperature and Time: It is recommended to use 94°C for denaturation. Recommended denaturation times are:
  • 30 seconds for simple templates like plasmid DNA.
  • 3 minutes for complex templates like cDNA or genomic DNA.
  • 5-10 minutes for templates with high GC content.
  1. Annealing Temperature and Time: The recommended annealing temperature is 60°C, but you can establish a temperature gradient if needed to find the optimal annealing temperature for primers. The recommended annealing time is 20 seconds, adjustable within 10-30 seconds. Prolonged annealing times may cause smear on the gel for PCR products.
  2. PCR Products: Store PCR products at -20°C to prevent DNA degradation.

These guidelines ensure optimal PCR conditions for efficient amplification of target DNA fragments, considering template complexity and GC content.

 

Notes

  1. This product is for research use only.
  2. Please operate with lab coats and disposable gloves for your safety.
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