In-Fusion PCR Cloning Kits enable directional, seamless cloning of any PCR fragment—or multiple fragments—into any linearized vector in a single 15-minute reaction. No additional treatment of the PCR fragment is required (such as restriction digestion, ligation, phosphorylation, or blunt-end polishing). The efficiency of In-Fusion HD cloning kits is over 95%.
How In-Fusion PCR Cloning Works
The In-Fusion Enzyme premix fuses PCR-generated sequences and linearized vectors efficiently and precisely, utilizing a 15 bp overlap at their ends. This 15 bp overlap can be engineered by designing custom primers for amplification of the desired sequences. This method can be used to clone single or multiple fragments into a single vector without subcloning.
In-Fusion HD Cloning Plus System Components
All In-Fusion HD Cloning Plus kits contain the In-Fusion HD enzyme premix and reagents for control experiments. In-Fusion HD Cloning Plus kits also include:
- CloneAmp HiFi High Fidelity PCR Premix
CloneAmp HiFi PCR Premix is a convenient 2X master mix that provides exceptionally accurate, efficient and fast DNA amplification. The premix contains dNTPs and an optimized buffer, allows rapid set-up of PCR reactions, and facilitates successful cloning.
- Stellar Competent Cells
An E. coli HST08 strain is included that provides high transformation efficiency (greater than 5 x 108 cfu/µg) and compliments the efficiency of the In-Fusion HD Cloning Plus kit.
- Cloning Enhancer or NucleoSpin Gel and PCR Clean-Up
- Cloning Enhancer (CE) is an easy-to-use enzyme premix that removes background plasmid DNA and PCR residue, eliminating the need for PCR insert purification prior to cloning. In-Fusion HD Cloning Plus CE kits are ideal for cloning when there is a single PCR product on the gel.
- Alternatively, NucleoSpin Gel and PCR Clean-Up is suitable for gel extraction as well as PCR purification.