Product Description

Hieff™ Taq DNA polymerase is a thermostable recombinant DNA polymerase expressed by Thermus aquaticus with a molecular weight of 94 kDa. It has 5'→3' polymerase activity and 5'→3' exonuclease activity but without 3'→5' exonuclease activity. The amplified product has 3'- dA and can be directly used for TA cloning.

Product Components

Component Number	Name	Cat#/Size
		10101ES80(1,000 U)	10101ES92(10,000 U)
10101-A	10×Taq Buffer (Mg2+ Free)	4×1 mL	4×10 mL
10101-B	25 mM MgCl2	2×1 mL	2×10 mL
10101-C	Hieff™ Taq DNA Polymerase (5 U/μL)	200 μL	2×1 mL

Applications

Genotyping; Colony PCR and other conventional PCR.

Unit Definition

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

Shipping and Storage

The product is shipped with ice packs and can be stored at -20°C for 2 years.

Cautions

1) For your safety and health, please wear lab coats and disposable gloves for operation.

2) This product is for research use ONLY！

PCR Reaction System composition (preparation on ice)

Components	Volume (μL)	Final Concentration
ddH2O	to 50	-
10×Taq Buffer (Mg2+ Free)	5	1×
25 mmol/L MgCl2	3	1.5 mmol/L
dNTP Mix (10 mmol/L each)	1	0.2 mmol/L
DNA template	optional	-
Forward primer (10 μmol/L)	2	0.4 μmol/L
Reverse primer (10 μmol/L)	2	0.4 μmol/L
Hieff® Taq DNA Polymerase (5 U/μL)	0.4	0.04 U/μL

[Notes]:

1) Final concentration of Mg²⁺：The optimal concentration of Mg²⁺ is 1.5-2 mmol/L. If necessary, the optimal concentration of Mg²⁺ can be explored upward at intervals of 0.2-0.5 mmol/L.

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

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

4) 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

Cycle step	Temperature	Time	Cycles
Initial denaturation	94℃	30 secs-5 mins	1
Denaturation	94℃	30 secs	35
Annealing	50-60℃	30 secs
Extension	72℃	60 secs/kb
Final Extension	72℃	10 mins	1

[Notes]:

1) Initial denaturation temperature and time: the recommended temperature is 94°C. The recommended pre-denaturation time is 30 secs for plasmid DNA and other simple templates; 3 min for complex templates such as cDNA and genomic DNA; 5-10 min for the template with high GC.

2) Annealing temperature and time: the recommended temperature is 60°C. Temperature gradient can be set up to find the optimum temperature for primer annealing. The recommended annealing time is set to 20 secs and can be adjusted within 10-30 sec. Too long annealing time may cause the amplified product diffusion on the agarose gel.

3) Amplification products: Please store the PCR amplification products at - 20°C to prevent DNA degradation.