NEBuilder® HiFi DNA Assembly

NEBuilder HiFi DNA Assembly enables virtually error-free joining of DNA fragments, even those with 5´- and 3´-end mismatches. This flexible kit enables simple and fast seamless cloning utilizing a proprietary high-fidelity polymerase. Find out why NEBuilder HiFi is the next generation of DNA assembly and cloning.

5 Reasons to choose NEBuilder HiFi 

  1. Save time – Enjoy simple and fast seamless cloning in as little as 15 minutes.
  2. Flexibility – Use one system for both "standard-size" cloning and larger gene assembly products, up to 6 fragments.
  3. Compatible with downstream applications – DNA can be used immediately for transformation or as template for PCR or RCA.
  4. Adaptable – Adapts easily for multiple DNA manipulations, including site-directed mutagenesis. 
  5. Save money - No licensing fees from NEB for NEBuilder products

NEBuilder HiFi DNA assembly offers unique features over other DNA assembly methods, including:

How does NEBuilder HiFi stack up?

graph iconPerformance data and advantages against NEB Gibson Assembly
graph iconPerformance data and advantages against In-Fusion HD
table iconSynthetic Biology/DNA Assembly Selection Chart
table iconComparison of DNA Assembly Reaction Types

Not your average DNA assembly reagent

NEBuilder HiFi DNA Assembly can be used for a variety of DNA assembly methods.


NEBuilder HiFi products

NEBuilder HiFi DNA Assembly Cloning Kit (NEB #E5520)
  • Includes NEB 5-alpha competent cells
  • NEB recommends NEB 5-alpha Competent E. coli (High Efficiency, NEB #C2987) for routine assemblies of 15 kb or less
  • For repetitive sequences, NEB recommends NEB Stable Competent E. coli (NEB #C3040)
NEBuilder HiFi DNA Assembly Bundle for Large Fragments (NEB #E2623)
  • Includes NEB 10-beta competent cells
  • NEB recommends NEB 10-beta Competent E. coli (High Efficiency, NEB #C3019) or NEB 10-beta Electrocompetent E. coli (NEB #C3020) for assemblies larger than 15 kb
NEBuilder HiFi DNA Assembly Master Mix (NEB #E2621)
  • Does not include competent cells 



One or more of these products are covered by patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc. For more information, please email us at [email protected]. The use of these products may require you to obtain additional third party intellectual property rights for certain applications. 

GIBSON ASSEMBLY® is a registered trademark of Synthetic Genomics, Inc.

IN-FUSION® is a registered trademark of ClonTech Laboratories, Inc.

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Protocols for NEBuilder® HiFi DNA Assembly
Application Notes NEBuilder® HiFi DNA Assembly