Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Asymmetric synthesis of alkyl fluorides <i>via</i> biocatalytic reduction of α-fluoroenones and α-fluoroenoates.

Green chemistry : an international journal and green chemistry resource : GC·2026
Same author

Transaminase and Norcoclaurine Synthase One-Pot Cascades Towards (1S)-Tetrahydroisoquinolines.

Chembiochem : a European journal of chemical biology·2026
Same author

The discovery of new metagenomic urethanases utilising a novel colorimetric assay for applications in the biodegradation of polyurethanes.

Green chemistry : an international journal and green chemistry resource : GC·2025
Same author

Using hyperspectral imaging and machine learning to identify food-contaminated compostable and recyclable plastics.

UCL open. Environment·2025
Same author

Functional Enrichment and Sequence-Based Discovery Identify Promiscuous and Efficient Poly Lactic Acid Degrading Enzymes.

Environmental science & technology·2025
Same author

Expanding the Enzymatic Toolbox for Carboligation: Increasing the Diversity of the 'Split' Transketolase Sequence Space.

Chembiochem : a European journal of chemical biology·2025
Same journal

Investigating the interactomic landscape of survival motor neuron (SMN) and the SMNΔ7 truncated protein.

BioTechniques·2026
Same journal

Antigen retrieval-immunofluorescence on free floating sections to visualize the liver lobule and its cellular makeup.

BioTechniques·2026
Same journal

Special approach of droplet digital polymerase chain reaction (ddPCR) for transgene stability of a Chinese hamster ovary (CHO) cell line.

BioTechniques·2026
Same journal

Strand-specific quantification of L1 ORF0 and related transcripts by multiplex reverse transcription with tagged primers.

BioTechniques·2026
Same journal

Why and when should we choose digital PCR?

BioTechniques·2026
Same journal

Quantitative and unbiased lung alveolar septum assessment in an LPS experimental mouse model using 2D-spatial correlation image analysis from hematoxylin and eosin slides.

BioTechniques·2026
See all related articles

Related Experiment Video

Updated: Dec 2, 2025

Author Spotlight: Exploring Cloning Techniques for Full-Length DNA Fragments
04:18

Author Spotlight: Exploring Cloning Techniques for Full-Length DNA Fragments

Published on: May 17, 2024

917

pET expression vector customized for efficient seamless cloning.

Dragana Dobrijevic1, Lily A Nematollahi2, Helen C Hailes3

  • 1Department of Biochemical Engineering, University College London, Gower Street, London, WC1E 6BT, UK.

Biotechniques
|November 2, 2020
PubMed
Summary
This summary is machine-generated.

Researchers modified the pET29 expression vector for faster parallel cloning using gene replacement and Golden Gate methods. This enables efficient cloning of diverse bacterial enzyme variants for biocatalysis applications.

Keywords:
Golden Gateexpression in E. colipET vectorssacBseamless cloning

More Related Videos

Generation of Plasmid Vectors Expressing FLAG-tagged Proteins Under the Regulation of Human Elongation Factor-1&#945; Promoter Using Gibson Assembly
10:18

Generation of Plasmid Vectors Expressing FLAG-tagged Proteins Under the Regulation of Human Elongation Factor-1α Promoter Using Gibson Assembly

Published on: February 9, 2015

37.7K
Author Spotlight: Unveiling the Potential of Unpurified Recombinant AAVs in Cell Culture Research
06:41

Author Spotlight: Unveiling the Potential of Unpurified Recombinant AAVs in Cell Culture Research

Published on: October 20, 2023

3.7K

Related Experiment Videos

Last Updated: Dec 2, 2025

Author Spotlight: Exploring Cloning Techniques for Full-Length DNA Fragments
04:18

Author Spotlight: Exploring Cloning Techniques for Full-Length DNA Fragments

Published on: May 17, 2024

917
Generation of Plasmid Vectors Expressing FLAG-tagged Proteins Under the Regulation of Human Elongation Factor-1&#945; Promoter Using Gibson Assembly
10:18

Generation of Plasmid Vectors Expressing FLAG-tagged Proteins Under the Regulation of Human Elongation Factor-1α Promoter Using Gibson Assembly

Published on: February 9, 2015

37.7K
Author Spotlight: Unveiling the Potential of Unpurified Recombinant AAVs in Cell Culture Research
06:41

Author Spotlight: Unveiling the Potential of Unpurified Recombinant AAVs in Cell Culture Research

Published on: October 20, 2023

3.7K

Area of Science:

  • Molecular Biology
  • Biotechnology
  • Synthetic Biology

Background:

  • The pET29 expression vector is widely used for protein expression in bacteria.
  • Efficient cloning strategies are crucial for discovering and engineering enzymes.
  • Genomic and metagenomic libraries offer vast potential for novel biocatalyst discovery.

Purpose of the Study:

  • To present a modified pET29 expression vector enabling rapid parallel cloning.
  • To demonstrate the utility of a gene replacement and Golden Gate strategy for vector modification.
  • To facilitate the cloning of numerous bacterial enzyme variants from diverse genetic sources.

Main Methods:

  • Modification of the pET29 expression vector.
  • Implementation of a gene replacement strategy.
  • Application of Golden Gate cloning for parallel assembly.
  • Cloning of enzyme variants from genomic and metagenomic DNA.

Main Results:

  • Successful modification of the pET29 vector for enhanced cloning efficiency.
  • Demonstration of rapid and straightforward parallel cloning capabilities.
  • Generation of a library of bacterial enzyme variants.
  • Application of the method to diverse genetic sources.

Conclusions:

  • The modified pET29 vector facilitates high-throughput cloning of enzyme variants.
  • The gene replacement and Golden Gate strategy is effective for vector engineering.
  • This approach accelerates the discovery of novel biocatalysts for industrial applications.