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

Transcription Attenuation in Prokaryotes02:42

Transcription Attenuation in Prokaryotes

18.2K
Transcriptional attenuation occurs when RNA transcription is prematurely terminated due to the formation of a terminator mRNA hairpin structure.  Bacteria use these hairpins to regulate the transcription process and control the synthesis of several amino acids including histidine, lysine, threonine, and phenylalanine. Transcription attenuation takes place in the non-coding regions of mRNA.
There are several different mechanisms used to attenuate transcription. In ribosome mediated...
18.2K
Cell Specific Gene Expression01:58

Cell Specific Gene Expression

16.3K
Multicellular organisms contain a variety of structurally and functionally distinct cell types, but the DNA in all the cells originated from the same parent cells. The differences in the cells can be attributed to the differential gene expression. Liver cells, whose functions include detoxification of blood, production of bile to metabolize fats, and synthesis of proteins essential for metabolism, must express a specific set of genes to perform their functions. Gene expression also varies with...
16.3K
Cell Specific Gene Expression01:58

Cell Specific Gene Expression

5.4K
5.4K
Cleavage and Blastulation01:33

Cleavage and Blastulation

49.9K
After a large-single-celled zygote is produced via fertilization, the process of cleavage occurs while zygotes travel through the uterine tube. Cleavage is a mitotic cell division that does not result in growth. With each round of successive cell division, daughter cells get increasingly smaller.
49.9K
RNA Editing02:23

RNA Editing

9.8K
RNA editing is a post-transcriptional modification where a precursor mRNA (pre-mRNA) nucleotide sequence is changed by base insertion, deletion, or modification. The extent of RNA editing varies from a few hundred bases, in mitochondrial DNA of trypanosomes, to a just single base, in nuclear genes of mammals. Even a single base change in the pre-mRNA can convert a codon for one amino acid into the codon for another amino acid or a stop codon. This type of re-coding can significantly affect the...
9.8K
Gene Conversion02:08

Gene Conversion

10.6K
Other than maintaining genome stability via DNA repair, homologous recombination plays an important role in diversifying the genome. In fact, the recombination of sequences forms the molecular basis of genomic evolution. Random and non-random permutations of genomic sequences create a library of new amalgamated sequences. These newly formed genomes can determine the fitness and survival of cells. In bacteria, homologous and non-homologous types of recombination lead to the evolution of new...
10.6K

You might also read

Related Articles

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

Sort by
Same author

Retargeted serine integrases for one-step, precise integration of large DNA sequences in human cells.

Nature biotechnology·2026
Same author

Functional Genomics Screening in <i>Chlamydomonas reinhardtii</i> Maps the Genetic Landscape of Tolerance to Paraquat and Diuron.

Environmental science & technology·2026
Same author

Comparison Between Perfusion and Non-Perfusion CT Neuroimaging in the Diagnosis of Acute Stroke-Can We Achieve a Similar Clinical Outcome?

Journal of medical imaging and radiation oncology·2026
Same author

Analytical Assessment of sgRNA Impurities and Their Impact on Functional Performance.

Analytical chemistry·2026
Same author

Detrimental effects associated with the Anterior Growth Guidance Appliance.

Journal of clinical orthodontics : JCO·2026
Same author

The fallacy of "extraction regret syndrome".

Journal of clinical orthodontics : JCO·2026

Related Experiment Video

Updated: Jan 21, 2026

Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage
06:51

Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage

Published on: May 6, 2020

4.3K

Enhancing gene editing specificity by attenuating DNA cleavage kinetics.

Jeffrey C Miller1, Deepak P Patil1, Danny F Xia1

  • 1Sangamo Therapeutics, Inc., Richmond, CA, USA.

Nature Biotechnology
|July 31, 2019
PubMed
Summary
This summary is machine-generated.

Engineered nucleases like zinc finger nucleases (ZFNs) can edit genomes precisely. This study enhances ZFN specificity by modifying the FokI domain, significantly reducing off-target mutations for safer gene editing applications.

More Related Videos

CIRCLE-Seq for Interrogation of Off-Target Gene Editing
08:23

CIRCLE-Seq for Interrogation of Off-Target Gene Editing

Published on: November 1, 2024

1.4K
Author Spotlight: Advancing Gene Editing in Bamboo Leaves for Sustainable Plastic Alternatives
06:57

Author Spotlight: Advancing Gene Editing in Bamboo Leaves for Sustainable Plastic Alternatives

Published on: August 18, 2023

2.2K

Related Experiment Videos

Last Updated: Jan 21, 2026

Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage
06:51

Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage

Published on: May 6, 2020

4.3K
CIRCLE-Seq for Interrogation of Off-Target Gene Editing
08:23

CIRCLE-Seq for Interrogation of Off-Target Gene Editing

Published on: November 1, 2024

1.4K
Author Spotlight: Advancing Gene Editing in Bamboo Leaves for Sustainable Plastic Alternatives
06:57

Author Spotlight: Advancing Gene Editing in Bamboo Leaves for Sustainable Plastic Alternatives

Published on: August 18, 2023

2.2K

Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Engineered nucleases, particularly zinc finger nucleases (ZFNs), are powerful tools for genome editing.
  • Concerns regarding off-target cleavage by ZFNs limit their therapeutic applications due to potential mutagenicity.
  • Improving the specificity of ZFNs is crucial for safe and effective gene editing therapies.

Purpose of the Study:

  • To develop an approach for enhancing the specificity of zinc finger nucleases (ZFNs).
  • To engineer the FokI catalytic domain to reduce cleavage at off-target sites.
  • To create highly specific ZFNs for therapeutic gene editing applications.

Main Methods:

  • Engineered single-residue substitutions in the FokI catalytic domain of ZFNs.
  • Tested ZFN variants for on-target activity and off-target cleavage efficiency.
  • Combined FokI domain engineering with reduced zinc finger affinity modifications.
  • Assessed ZFN specificity for the TRAC locus in T cells.

Main Results:

  • Engineered FokI domain substitutions preserved on-target activity.
  • Off-target indels were reduced by up to 3,000-fold with modified FokI domains.
  • Developed highly specific ZFNs for the TRAC locus, achieving 98% knockout in T cells.
  • No detectable off-target activity was observed at an assay background of ~0.01%.

Conclusions:

  • Modifying the FokI catalytic domain is an effective strategy to enhance ZFN specificity.
  • The developed FokI variants enable the generation of highly specific nucleases for gene editing.
  • This approach facilitates the routine creation of gene editing tools with no detectable off-target activity, advancing therapeutic potential.