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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 |
|
| 2 | EZchange™ Site-directed Mutagenesis Kit | EZ004M | - | 40 reactions | ￦ 270,000 |
|
▶ 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 DNAa (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.
● Enzynomics_EZ004_EZchange™ Site-directed Mutagenesis Kit_manual_kor
● Enzynomics_EZ004_EZchange™ Site-directed Mutagenesis Kit_manual_simple(eng)
● Material Safety Data Sheet
- Enzynomics_MSDS_EZ004_EZchange™ Site-directed Mutagenesis Kit_E
● Related paper
1. S-M Ahn, et al(2012). Oncogene 31, 3051-3059
2. Hayeong Kwon, et al(2009). Biochim Biophys Acta 1793, 1325-1333
3. Huong T. T. Ngo, et al(2013). J. Virol. 87, 5718-5731
