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Hieff® Taq DNA Polymerase

Product SKU

Specifications

Price

10101ES80

1000 U

$23.93

10101ES90

5×1000 U

$113.48

Product Description

Hieff® Taq DNA polymerase is a thermostable recombinant DNA polymerase expressed by Thermus aquaticus with a molecular weight of 94 KD. It has 5'→3' polymerase activity and 5'→3' exonuclease activity, but no 3'→5' exonuclease activity. The amplified product has 3'- dA and can be directly used for TA cloning.

Package Information

Contents No.

Name

10101ES80(1000 U)

10101ES90(5×1000 U)

10101-A

10×Taq Buffer (Mg2+ Free)

1 mL×4

1 mL×20

10101-B

25 mM MgCl2

1 mL×2

1 mL×10

10101-C

5×1000 U

200 μL

200 μL×5

Application

Genotyping, colony PCR and other conventional PCR.

Unit Definition

Using the activated DNA of salmon sperm as template / primer, the activity was defined as one active unit (U) when 10 nmol of total nucleotide was ingested as acid insoluble substance at 74℃ for 30 min.

Quality Control

Exonuclease Activity: In 20 μL reaction system, 10 U of this product and 0.6 μg of λ - HindIII were incubated at 74℃ for 1 h, and the electrophoresis band of DNA had no change.

Endonuclease Activity: In 20 μL reaction system, 10 U of the product and 0.6 μg of supercoiled pBR322 DNA were incubated at 74 ℃ for 1 h, and the DNA electrophoretic bands did not change.

Residual E.coli gDNA Test: The E.coil 16S rDNA gene was amplified by adding 2 U of the product into 50 μL system and using sterile ddH2O as template. After 35 cycles, the amplified products were subjected to 1% agarose gel electrophoresis and EB staining without amplification bands.

Shipping and Storage

Transport in ice packs,it can be stored at -20℃ for 2 years.

Important points before starting

For your safety and health, please wear experimental clothes and disposable gloves.

Reaction composition(Preparation on ice)

Components

Volume(μL)

Final Concentration

ddH2O

to 50

-

10×Taq Buffer(Mg2+ Free)

5

25 mM MgCl2

3

1.5 mM

dNTP Mix (10 mM each)

1 μL

0.2 mM

DNA template

optional

-

Forward primer (10 μM)

2 μL

0.4 μM

Reverse primer (10 μM)

2 μL

0.4 μM

Hieff® Taq DNA Polymerase (5 U/μL)

0.4 μL

0.04 U/μL

【Note】

Final concentration of Mg2+:The best concentration of Mg2+ is 1.5-2 mm. If necessary, the best concentration of Mg2+ can be found at intervals of 0.2-0.5 mm.

Polymerase addition: at room temperature, the polymerase has a certain degree of 5'- 3' polymerase activity. In order to prevent non-specific amplification, it is suggested to add the polymerase to the reaction system in the last step.

Concentration of polymerase:0.04 U/μL is recommended. It can be optimized between 0.025-0.04 U/ μL.

Recommended use of different templates (50 μL reaction system):

Type of template

Template usage

Genomic DNA

50 ng-100 ng

Plasmid DNA

10 pg-20 ng

cDNA

1-5 μL (No more than 1/10 of the reaction system)

Thermal cycling protocol

Stage

Temperature(ºC)

Time

Cycles

Pre-denaturation

94

30 sec-5 min

1

Denaturation

94

30 sec

35

Annealing

50-60

30 sec

35

Extension

72

60 sec/kb

35

Final Extension

72

10 min

1

【Note】

Pre denaturation temperature and time:94℃ is recommended. The recommended pre denaturation time is 30 sec for plasmid DNA and other simple templates; 3 min for complex templates such as cDNA and genomic DNA; The template with high GC was 5-10 min.

Annealing temperature and time: 60℃ is recommended. Temperature gradient can be set up to find the optimum temperature of primer annealing. The recommended annealing time is set to 20 sec and can be adjusted in 10-30 sec. The annealing time is too long, which may lead to the diffusion of the product on the adhesive.

Amplification products: Please store the PCR amplification products at - 20℃ to prevent DNA degradation.

15221439726

15221439726