Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Related Concept Videos

Conservative Site-specific Recombination and Phase Variation

Conservative Site-specific Recombination and Phase Variation

Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...

CRISPR/Cas9 Genome Editing

CRISPR/Cas9 Genome Editing

The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...

You might also read

Related Articles

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

Sort by

Same author

Tracking Synthetic Adhesins on Bacterial Surfaces with Immunofluorescence Microscopy.

Methods in molecular biology (Clifton, N.J.)·2026

Same author

Emergent bioengineering.

Current opinion in biotechnology·2026

Same author

Social relevance of microbiology literacy.

FEMS microbiology letters·2026

Same author

Precision Proteolysis of Triosephosphate Isomerase of <i>Escherichia coli</i> Boosts Dihydroxyacetone Phosphate Biosynthesis.

ACS synthetic biology·2026

Same author

Synthetically primed growth of Pseudomonas putida on 2,4-dinitrotoluene as sole carbon and nitrogen source.

Metabolic engineering·2026

Same author

The disputed identity of <i>Pseudomonas putida</i> KT2440: when taxonomists rename your favorite microbe.

mBio·2026

Search research articles

Related Experiment Video

Updated: Sep 22, 2025

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Published on: April 11, 2019

High-Efficiency Multi-site Genomic Editing (HEMSE) Made Easy.

Tomás Aparicio¹, Víctor de Lorenzo², Esteban Martínez-García¹

¹Systems Biology Department, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.

Methods in Molecular Biology (Clifton, N.J.)

|May 18, 2022

Summary

This summary is machine-generated.

This study details a new method for bacterial genome engineering in Pseudomonas putida. The High-Efficiency Multi-site genomic Editing (HEMSE) workflow enables simple and efficient introduction of multiple genetic mutations.

Keywords:

Cycled recombineering HEMSE Multiplex genome editing Pseudomonas putida Synthetic biology ssDNA

More Related Videos

Author Spotlight: Efficient CRISPR/Cas9 Genome Editing in Bone Marrow-Derived Macrophages for Precise Gene Disruption

Author Spotlight: Efficient CRISPR/Cas9 Genome Editing in Bone Marrow-Derived Macrophages for Precise Gene Disruption

Published on: August 4, 2023

Establishment of Genome-edited Human Pluripotent Stem Cell Lines: From Targeting to Isolation

Establishment of Genome-edited Human Pluripotent Stem Cell Lines: From Targeting to Isolation

Published on: February 2, 2016

Related Experiment Videos

Last Updated: Sep 22, 2025

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Published on: April 11, 2019

Author Spotlight: Efficient CRISPR/Cas9 Genome Editing in Bone Marrow-Derived Macrophages for Precise Gene Disruption

Author Spotlight: Efficient CRISPR/Cas9 Genome Editing in Bone Marrow-Derived Macrophages for Precise Gene Disruption

Published on: August 4, 2023

Establishment of Genome-edited Human Pluripotent Stem Cell Lines: From Targeting to Isolation

Establishment of Genome-edited Human Pluripotent Stem Cell Lines: From Targeting to Isolation

Published on: February 2, 2016

Area of Science:

Microbiology
Synthetic Biology
Genomics

Background:

Bacterial genome engineering is vital for biotechnology.
Single-stranded (ss) DNA recombineering is an effective method for introducing genomic mutations.
This technology has been adapted for various bacterial species, including Pseudomonas putida.

Purpose of the Study:

To present a detailed protocol for performing recombineering experiments in P. putida.
To enable the introduction of single and multiple genomic changes efficiently.
To implement the High-Efficiency Multi-site genomic Editing (HEMSE) workflow for P. putida.

Main Methods:

Utilizing a phage recombinase effective in the target strain.
Facilitating the introduction of ssDNA oligonucleotides carrying desired mutations.
Transiently suppressing the endogenous mismatch repair (MMR) system using a dominant-negative mutL allele (MutL_E36K^PP).

Main Results:

A protocol for easily performing recombineering experiments in P. putida is detailed.
The HEMSE workflow allows for the simultaneous introduction of multiple genomic mutations.
The method facilitates virtually any type of genomic edit.

Conclusions:

The described protocol and HEMSE workflow provide an efficient and straightforward method for engineering the P. putida genome.
This advancement is significant for various bio-related technologies relying on precise bacterial genetic modification.
The technique enhances the versatility of P. putida for metabolic and environmental applications.