GenieClone DNA Assembly Site-Directed Mutagenesis Kit v1

GenieClone Site-Directed Mutagenesis Kit v1 is designed for rapid site-directed mutagenesis of up to 2 separate mutations in the same reaction in less than 3 hours. Based on GenieClone rapid cloning technology, this kit uses homologous recombination to replace the conventional annealing ring-forming reactions meaning much less template and a more flexible primer design strategy. With seamless splicing of the two PCR products by GenieClone technology, the kit can complete up to two separate mutations with a single amplification reaction. DpnI digested products of the specific amplicons can be directly added to the recombination reaction without purification. The highly optimized reaction buffer, fast protocol and superior site-directed mutagenesis efficiency make GenieClone Site-Directed Mutagenesis Kit v1 make an excellent choice for introducing 1 or 2 mutations at discontinuous sites simultaneously on the same plasmid.

Figure 1: Single base site-directed mutation using GenieClone Site-Directed Mutagenesis Kit v1

• Introduce up to 2 separate mutations in the same reaction.
• Site-directed mutagenesis can be performed on up to 2 discontinuous sites (more than 50 bp apart) simultaneously on the target plasmid.
• Single kit for cloning and mutagenesis.
• Includes Genie Fusion Ultra High-Fidelity DNA Polymerase providing high-fidelity PCR with extremely low mutation rates & excellent long fragment amplification for any plasmid amplification up to 20 kb as well as GC-rich templates.
• Amplification is carried out exponentially and the template usage is extremely low, which is beneficial to the complete degradation of the original methylation template.
• DpnI eliminates contamination of the original template and decreases background.
• Amplified products can be directly used in the recombination reaction after DpnI treatment.
• The reaction mixture can be used to directly transform chemically competent E.coli cells.

Figure 2: Schematic of introducing single base or continuous multi-bases site-directed mutation into plasmid

Note: use methylase non-defective host strains (eg. Top10, DH5α, JM109) to extract the original plasmids for use with this kit.

Store the product at -20°C, and it will be valid for 1 year.