DNA Assembly and Cloning

cloning owl

For the purposes of cloning, DNA assembly refers to a method to physically link together multiple fragments of DNA, in an end-to-end fashion, to achieve a desired, higher-order assembly prior to joining to a vector. This process is the cornerstone of the synthetic biology movement, and allows the construction of novel biological systems and devices using defined components. The methods are carried out in vitro, and are typically enzymatically driven with the final constructions being maintained in microbial host cells.

To help select the best DNA assembly method for your needs, please use our Synthetic Biology/DNA Assembly Selection Chart.

Learn more about the various DNA Assembly methods: NEBuilder HiFi DNA Assembly, Gibson AssemblyBioBrick AssemblyGolden Gate Assembly

  1. NEBuilder® HiFi DNA Assembly: Removal of 3´-end Mismatches

    Did you know that one of the advantages of NEBuilder HiFi is that it removes 3’ and 5’ -end mismatch sequences prior to fragment assembly? Learn how!

  2. Construction of an sgRNA-Cas9 Expression Vector via an ssOligo Bridge

    Learn how you can use NEBuilder HiFi to generate an sgRNA-Cas9 expression vector with a single-stranded oligo bridge.

  3. Introduction to NEBuilder HiFi DNA Assembly

    Find out how NEBuilder® HiFi DNA Assembly can reliably join DNA fragments in a single tube, isothermal reaction, with advantages over NEB Gibson Assembly®.

  4. Introduction to Gibson Assembly® E5510 E2611

    Watch an interactive tutorial that details the process by which Gibson Assembly® joins DNA fragments in a single tube, isothermal reaction.

  5. Primer Design and Fragment Assembly Using NEBuilder® HiFi DNA Assembly or Gibson Assembly® E5510 E5520

    Watch an interactive tutorial on primer design to see how simple it really is to clone with either NEBuilder HiFi DNA Assembly or the Gibson Assembly Cloning Kit.

Featured Products

DNA Assembly and Cloning includes these areas of focus:

NEBuilder® HiFi DNA Assembly
Gibson Assembly®
BioBrick® Assembly
Golden Gate Assembly

Protocols for DNA Assembly and Cloning

    Publications related to DNA Assembly and Cloning:

  1. Binder A, Lambert J, Morbitzer R, Popp C, Ott T, Lahaye T, Parniske M (2014). A Modular Plasmid Assembly Kit for Multigene Expression, Gene Silencing and Silencing Rescue in Plants PLoS One. 9(2), e88218. PubMedID: 24551083, DOI: 10.1371/journal.pone.0088218
  2. Feng Y, Zhang S, Huang X (2014). A robust TALENs system for highly efficient mammalian genome editing Sci Rep. 4, 3632. PubMedID: 24407151, DOI: 10.1038/srep03632
  3. Abil Z, Denard CA, Zhao H (2014). Modular assembly of designer PUF proteins for specific post-transcriptional regulation of endogenous RNA J Biol Eng. 8(1), 7. PubMedID: 24581042, DOI: 10.1186/1754-1611-8-7

Overview of Gibson Assembly®

Gibson Assembly® (NEB #E2611) was developed by Dr. Daniel Gibson and his colleagues at the J. Craig Venter Institute and licensed to NEB by Synthetic Genomics, Inc. It allows for successful assembly of multiple DNA fragments, regardless of fragment length or end compatibility. It has been rapidly adopted by the synthetic biology community due to its ease-of-use, flexibility and suitability for large DNA constructs.
Gibston Assembly® is a registered trademark of Synthetic Genomics, Inc.

Overview of BioBrick® Assembly Kit

The BioBrick™ Assembly Kit (NEB #E0546) provides a streamlined method for assembly of BioBrick parts into multi-component genetic systems. BioBrick parts are DNA sequences that encode a defined biological function and can be readily assembled with any other BioBrick part. The process for assembling any two BioBrick parts is identical and results in a new composite BioBrick part.
The BioBrick Assembly Kit was developed in partnership with Ginkgo BioWorks.
BioBrick™ is a trademark of The BioBricks Foundation.