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Polymerases

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2X TOPsimple™ DyeMIX-multi HOT

No Product Name Cat.# Conc Size Price(₩) Add
1 2X TOPsimple™ DyeMIX-multi HOT P510HC 2X 1 ml ₩ 80,000 추가
2 2X TOPsimple™ DyeMIX-multi HOT P525HC 2X 2.5 ml ₩ 160,000 추가
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Product description

2X TOPsimple™ DyeMIX-multi HOT is similarly formulated to 2X TOPsimple™ PreMIX-multi HOT except that it contains loading dyes for further convenience of use. Thus, the reaction mixtures after PCR cycles are ready for agarose gel electrophoresis. This product can be used in general multiplex PCR and genotyping experiment related to genetic diagnostics because it produces up to 20 different amplified products within a single tube reaction.

 

Characteristics

- 2-Dye system: Easy for gel electrophoresis (xylene cyanol and Orange G)

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

 

Applications

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

- Amplification of cDNA and genomic DNA.

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

- Primer extension

- Multiplex PCR

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

  temperature

- Colony PCR

 

Components (2X)

nTaq-multi HOT DNA Polymerase: 0.2 unit/μl

nTaq-multi HOT buffer (containing 4 mM MgCl2)

- dNTP mixture: 0.4 mM each

- Stabilizer

- Dyes: Xylene cyanol and Orange G

 

Migration of dyes in agarose gel

In an ordinary agarose gel, xylene cyanol co-migrate with 4-kb DNA fragments, and Orange G with 50-bp DNA fragments.

 

Storage

Stable up to 18 months at -20 or 3 months at 4 (Storage at -20 is recommended).

 

Standard reaction conditions

- PCR mixturea

2X TOPsimple™ DyeMIX-multiHOT

10 μl

Template DNAb (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

aAssmeble the reaction mixture on ice

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

  

- PCR cycles

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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