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EZchange™ Site-directed Mutagenesis Kit

No Product Name Cat.# Conc Size Price(₩) Add
1 EZchange™ Site-directed Mutagenesis Kit EZ004S - 20 reactions ₩ 150,000 추가
2 EZchange™ Site-directed Mutagenesis Kit EZ004M - 40 reactions ₩ 270,000 추가
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▶ EZchange™ Site-directed Mutagenesis kit primer design program


Product description

EZchange™ Site-directed Mutagenesis kit allows site- specific mutations with virtually any double-stranded plasmid as template, and thus eliminates a time-consuming subcloning of target DNA into single stranded DNA-producing plasmids. In addition, no specialized vectors or unique restriction sites are required. Therefore, mutations can be produced in a simple and convenient way.

 

Characteristics

- No subcloning

- High efficiency

- Site-direted mutagenesis in a single day

 

Applications

- Substitution

- Single or multiple deletions

- Insertion mutation

 

Kit contents

- nPfu-Frote

- 10X Reaction Buffer

- dNTP mixture (2 mM each)

- EZ-MIX

- 2X EZ-MIX Buffer

- Control primers (10 pmol/μl)

- Control Plasmid (5ng/μl)

- Sterile water

 

Standard reaction conditions

- Mutant Strand Synthesis Reaction

10X Reaction buffer

5 μl

nPfu-Forte (2.5 units/μl)

1 μl

dNTP mixture (2 mM each)

4 μl

Plasmid DNA(1~10 ng)

X μl

Primer 1 (10 pmol/μl)

1 μl

Primer 2 (10 pmol/μl)

1 μl

Distilled water

up to 50 μl

aTemplate plasmids should be purified from dam+ E. coli strains. Mutation efficiency would decrease markedly if plasmids isolated from dam- strains are used, since they are not digested by Dpn l. Most E. coli strains used in laboratories are dam+ except JM110 and SCS110.

 

Initial denaturation

95

2 min

Denaturation

95

30 sec

Annealing

55

30~60 sec

Elongation

72

1 min/kb

Number of cycles

25 times

Final elongation

72

10 min

Cautions: Adjust the temperature of the reaction tube to 94 prior to the addition of nPfu-Forte. The high temperature prevents nonspecific amplification or unwanted primer degradation. As a result, the mutation efficiency is significantly increased.

 

- EZ-MIX reactions

2X EZ-MIX Buffer

10 μl

PCR products (100~200 ng)

5~9 μl

EZ-MIX

1 μl

Distilled water

up to 20 μl

→ Incubate the reaction mixture for 30 min at 37

 

- Transformation

 Use 1-2 μl of DNA treated with EZ-MIX for transformation and spread 1/100 to 1/10 of transformed  

cells on appropriate plates.

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