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Gibson Assembly® Site-Directed Mutagenesis Kit
The Gibson Assembly® Method is a well-established assembly reaction that can be leveraged to join multiple, mutagenized DNA fragments with overlapping ends. Following mutagenesis, DNA fragments of various lengths are uniformly assembled using complementary overlaps between fragments. The inherent flexibility of this approach lends itself to small and large constructs and encompasses both single and multiple insert assemblies.
Summary
- Accurate, seamless, and efficient
- Multiple mutation sites (up to 5) incorporated in a single round
- Accommodates large insertions and deletions
- 4-6 hours process with less than 40 minutes hands-on time
- Optimal for 1–5 inserts
- Suitable for fragments ranging from 500 bp – 5.5 kb
- Use 40-mer primers for < 4 changes within a site
- Use 60-mer primers for more complex mutations (e.g. 40 base insertion)
- Expected efficiency: 95% for single site and 55% for five site mutations
Linear Template
Mutagenesis
- Generate primers for insertion, substitution, or deletion mutagenesis
- Perform PCR amplification to introduce mutation(s) and overlapping ends using GA SDM PCR Mix (2X)
Gibson Assembly® Method
- Combine DNA fragments and vector for assembly
- GA SDM Assembly Mix A (2X) mediates 3’ Chew Back
- Fragments anneal
- GA SDM Assembly Mix B (2X) mediates extension and repair
Circular Template
Mutagenesis
- Generate primers for insertion, substitution, or deletion mutagenesis
- Perform PCR amplification to introduce mutation(s) and overlapping ends using GA SDM PCR Mix (2X)
Gibson Assembly® Method
- Combine DNA fragments and vector for assembly
- GA SDM Assembly Mix A (2X) mediates 3’ Chew Back
- Fragments anneal
- GA SDM Assembly Mix B (2X) mediates extension and repair
Applications
- Protein Engineering
- Error-free Cloning
- Gene Editing
- Mutational Analysis
5-site mutations with Gibson Assembly® Site-Directed Mutagenesis Kit
5-site mutations were generated in a 11.7 kb plasmid and the PCR product was gel purified or Dpnl treated (and column purified)
Comparison of Various Site-Directed Mutagenesis Kits
1-site mutation was generated in a 2.7 kb pUC plasmid using Synthetic Genomic, Inc.'s kit and 2 competitors' kits
Product Ordering Information
| Products | Catalog # | Price |
|---|---|---|
| Gibson Assembly® Site-Directed Mutagenesis Starter Kit, 5 rxn | GA2100-S | $95 |
| Gibson Assembly® Site-Directed Mutagenesis Kit, 10 rxn | GA2100-10 | $179 |
Above mentioned pricing is valid only in the U.S. For international pricing please contact customerservice@sgidna.com.
Prices do not include shipping and handling. Submit by 2pm (PST) to receive order by the next day. For bulk packaged size, lead time is ~3 weeks.
Gibson Assembly method is also available under commercial license, please inquire at info@sgidna.com
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