컨텐츠 바로가기 영역
주메뉴로 바로가기
본문으로 바로가기

Polymerases

Home > PRODUCTS > Polymerases

2X TOPsimple™ DyeMIX–nTaq

No Product Name Cat.# Conc Size Price(₩) Add
1 2X TOPsimple™ DyeMIX–nTaq P510T 2X 1 ml ₩ 45,000 추가
2 2X TOPsimple™ DyeMIX–nTaq P525T 2X 2.5 ml ₩ 90,000 추가
Product Name Cat.# Size Price(₩) EA Total(₩) 삭제
Total : 장바구니에 담기 바로 구매 하기
plus minus 삭제

Product description

2X TOPsimple™ DyeMIX-nTaq is similarly formulated to the 2X TOPsimple™ PreMIX-nTaq 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 includes nTaq DNA polymerase (Cat. P025, P050), and is optimized for high efficiency PCR amplification with a high success rate.

 

Characteristics

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

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

 

Applications

- Amplification of DNA fragments shorter than 3 kb (Suitable for general PCR analysis)

- Amplification of cDNA and genomic DNA.

- Primer extension

- Colony PCR

 

Components (2X)

- nTaq: 0.2 unit/μl

- nTaq buffer (containing 3 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

TOPsimple™ DyeMIX–nTaq

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

aAssemble 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 denaturation

95

2 min

1

Denaturation

95

30 sec

25~35

Annealinga

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

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



Figure 1. Amplification of target DNA using E. coli cells directly as template DNA source with 2X TOPsimple™ DyeMIX

Colonies to be tested were first suspended in distilled water (30 μl), followed by serially dilution. The diluted cells were used directly as template DNA source for PCR reactions containing 2X TOPsimple DyeMIX (Cat. P510). Target DNAs were amplified by using the following PCR cycles; 95 2 min, (95 30 sec,55 30 sec, 72 30 sec) x 30, 72 5 min. The numbers of E.coli cells used were as indicated on top of the gel. In lane 2, 1 μl of the original cell suspension was used for PCR. The PCR reaction mixtures were prepared by adding serially diluted cells (1 μl) and primers to 10 μl of 2X TOPsimple™ DyeMIX in distilled water. The final volume was then adjusted to 20 μl with water. 


List