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Polymerases

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nTaq-multi HOT

No Product Name Cat.# Conc Size Price(₩) Add
1 nTaq-multi HOT P725HC 5 units/μl 250 units ₩ 100,000 추가
2 nTaq-multi HOT P750HC 5 units/μl 500 units ₩ 180,000 추가
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Product description

nTaq-multi HOT is a hot start PCR polymerase which remains inactive at temperatures lower than 75. Therefore, a heat activation step is required to activate nTaq-multi HOT. Activation is accomplished at denaturation steps of PCR cycles and, thus, nTaq-multi HOT is active only after initiation PCR cycles. nTaq-multi HOT produces up to 20 different amplified products with a single tube reaction. This product can be used in general multiplex PCR and genotyping experiment related to genetic diagnostics.

 

Characteristics

- Molecular weight: 94 kDa

- Error rate: 2.4 X 10-5

- Thermal stability: half life of 40 min at 95

- A-tail formation at 3’ ends of amplified duplex DNA.

- No activity at temperatures lower than 75. Heating up to 95 results in activation of enzyme.

 

Applications

- High specific amplification of DNA fragments shorter than 3kb.

- Amplification of cDNA and genomic DNA.

- Amplification of template DNA with secondary or higher ordered structure that is resistant to PCR amplification

- Well-suited for an automated PCR machines, for which PCR reaction mixtures are prepared at room 

  temperature

- Primer extension

- Multiplex PCR

 

Supplied with

- 10X nTaq-multi HOT buffer 

- dNTP Mixture (2 mM each)

- GC Melt I

- GC Melt II

- Sterile water

 

Quality control

- Purity: >99% on SDS-PAGE

- Endonuclease-free

- Exonuclease-free

- RNase free

- Inhibitor-free

 

Unit definition

One unit is defined as the amount of enzyme required to incorporate 10 nmol of dNTP into acid insoluble materials in 30 min at 74 in a 50-μl reaction mixture (20 mM Tris-HCl/pH 8.8, 50 mM KCl, 2.5 mM MgCl2, 10 mM β-mercaptoethanol, 12.5 μg of calf thymus DNA).

 

Storage buffer

20 mM Tris-HCl (pH 7.9), 100 mM KCl, 0.1 mM EDTA, 1 mM DTT, 0.5% NP-40, 0.5% Tween-20, 50% glycerol.

 

10X nTaq-multi HOT buffer

Containing 15 mM MgCl2

 

GC Melt l and ll

- This product is useful for amplification of DNA with GC rich sequences or to avoid amplification of non-specific

  bands (Use of GC Melt may reduce PCR efficiency).

- In general, use 1X strength in PCR reaction by diluting the 10X solution, but the amount should be adjusted 

  for optimal results.

 

Cautions

nTaq-multi HOT requires 10 minutes of initial denaturation to ensure effective activation of the enzyme.

 

Standard reaction conditions

- PCR mixturea

10X nTaq-multi HOT buffer

2 μl

nTaq-multi HOTb (5 units/μl)

0.2 μl

dNTP Mixture (2 mM each)

2 μl

Template DNAc (0.1~500 ng/μl)

1 μl

Primer 1 (5 pmole/μl)

1 μl

Primer 2 (5 pmole/μl)

1 μl

Sterile water

up to 20 μl

aAssemble the reaction mixture on ice

bAdd the PCR polymerase at the final step

cPlasmid DNA, 0.1 ng-30 ng; genomic DNA, 50 ng-500 ng

 

- PCR cycle

Step

Temperature

Time

Cycle

Initial denaturationa

95

10 min

1

Denaturation

95

30 sec

25~35

Annealingb

55~65

30~60 sec

Elongation

72

1 min/kb

Final elongation

72

5 min

1

When cycles are over, keep the reaction at 4; may add 10 mM EDTA until use to prevent DNA degradation

aAt least 10 min of initial denaturation time is required to fully activate the chemically modified PCR DNA polymerase

bRecommend annealing temperature is 5 to 10 below the lower Tm of the two primers used.

 


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