NEBuilder HiFi DNA Assembly enables virtually error-free joining of DNA fragments, even those with 5´ and 3´ restriction enzyme mismatches.
This flexible kit enables simple and fast Seamless Cloning utilizing a new proprietary high-fidelity polymerase.
Find out why NEBuilder HiFi is the next generation of DNA assembly and cloning.
NEBuilder HiFi DNA Assembly Workflow
Illustration NEBuilder Method
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Illustration NEBuilder Workflow
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Benefits over Gibson Assembly® Master Mix
- Enjoy less screening/re-sequencing of constructs, with virtually error-free, high-fidelity assembly
- Join DNA fragments together more efficiently, even with larger fragments or low DNA inputs
- Use NEBuilder HiFi in successive rounds of assembly, because it removes 5´- and 3´- restriction enzyme mismatches
- Bridge two double-stranded fragments with a synthetic single-stranded DNA oligo for simple and fast construction (e.g. linker insertion or gRNA libraries)
- Switch from other systems easily, because NEBuilder HiFi is compatible with Gibson Assembly-designed (and other) fragments
- Save time with simple and fast Seamless Cloning
- No licensing fee requirements from NEB for the NEBuilder products
Comparison NEBuilder and Gibson Assembly
NEBuilder HiFi DNA Assembly Master Mix offers improved efficiency and accuracy over Gibson Assembly with different numbers of fragments.
Reactions were set up in a 2- and 6- fragment assembly reaction according to recommended reaction conditions.
NEBuilder HiFi DNA Assembly Master Mix offers improved efficiency and accuracy over Gibson Assembly, with lower amounts of DNA by increasing overlaps.
Reactions were set up in a 4- fragment assembly reaction according to recommended reaction conditions. Amount of DNA and size of overlap is shown.
NEBuilder HiFi DNA Master Mix offers improved fidelity over Gibson Assembly Master Mix
Fidelity of assembled products was compared between NEBuilder HiFi DNA Assembly Master Mix (NEB #E2621) and Gibson Assembly Master Mix (NEB #E2611). Experiments were performed using various fragment and vector sizes following suggested protocols. Experiments D and E did not yield any colonies with Gibson Assembly. Experiments H and J were performed with fragments containing 3´ restriction enzyme mismatches. Experiments B through G vary because sequences of fragments are different.
Introduction to NEBuilder HiFi DNA Assembly
Primer Design and Fragment Assembly Using NEBuilder HiFi DNA Assembly or Gibson Assembly
NEBuilder HiFi DNA Assembly: Bridging dsDNA with a ssDNA Oligo