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

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

1.6K
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...
1.6K
Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

6.6K
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...
6.6K
CRISPR01:59

CRISPR

57.4K
Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...
57.4K

You might also read

Related Articles

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

Sort by
Same author

Optimal dismantling of directed networks.

Nature communications·2026
Same author

IL-22 promotes genesis of small intestinal secretory cells that protect against cholera in mice.

Nature microbiology·2026
Same author

Machine learning-based Personalized Dietary Recommendations to Achieve Desired Gut Microbial Compositions.

bioRxiv : the preprint server for biology·2026
Same author

Early-life Wnt4 expressing colon stromal cells orchestrate lifelong mucosal homeostasis via BMP-driven iNKT cell imprinting.

Nature communications·2026
Same author

A pro-carcinogenic bacterial toxin binds claudin-4 to cleave E-cadherin.

Nature·2026
Same author

IL-22 promotes genesis of small intestinal secretory cells that protect against cholera in mice.

bioRxiv : the preprint server for biology·2026

Related Experiment Video

Updated: Jan 10, 2026

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format
08:25

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

Published on: April 8, 2017

14.4K

Adapting CRISPR-associated transposons for rapid and high-throughput reverse genetics.

David W Basta1,2,3, Franz G Zingl2,3, Yiyan Yang4

  • 1Department of Pathology, Brigham and Women's Hospital, Boston, MA, 02115, USA.

Biorxiv : the Preprint Server for Biology
|November 24, 2025
PubMed
Summary
This summary is machine-generated.

MultiCAST enables high-throughput bacterial genetic screening using CRISPR-associated transposons (CAST) and guide RNA barcoding. This platform streamlines genome engineering and identifies condition-specific fitness genes efficiently.

More Related Videos

A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins
08:37

A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins

Published on: April 30, 2018

8.1K
High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots
12:59

High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots

Published on: April 30, 2016

18.6K

Related Experiment Videos

Last Updated: Jan 10, 2026

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format
08:25

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

Published on: April 8, 2017

14.4K
A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins
08:37

A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins

Published on: April 30, 2018

8.1K
High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots
12:59

High-throughput CRISPR Vector Construction and Characterization of DNA Modifications by Generation of Tomato Hairy Roots

Published on: April 30, 2016

18.6K

Area of Science:

  • Molecular Biology
  • Genomics
  • Synthetic Biology

Background:

  • CRISPR-associated transposons (CAST) are programmable tools for bacterial genome engineering.
  • Existing CAST systems are not optimized for high-throughput genetic screening applications.

Purpose of the Study:

  • To develop MultiCAST, a streamlined platform for rapid, scalable, guide RNA-directed transposon insertion in bacteria.
  • To enable pooled, high-throughput genetic screens using amplicon sequencing for molecular barcoding.
  • To identify factors influencing CAST transposition efficiency and optimize screening protocols.

Main Methods:

  • Conjugative delivery of plasmids encoding CAST machinery and a mini-transposon with a guide RNA.
  • Utilizing guide RNA sequences as molecular barcodes for pooled screening.
  • Developing a machine learning model to predict active guide RNAs based on target sequence features.
  • Investigating the role of nucleoid-associated protein H-NS in CAST activity.

Main Results:

  • MultiCAST enables single-step, targeted transposon insertions for scalable genetic screens.
  • Optimized conjugation ratios minimize guide-transposon crosstalk.
  • Machine learning model accurately predicts active guide RNAs.
  • H-NS was identified as an inhibitor of CAST activity, explaining variable insertion frequencies.
  • A large-scale screen in *E. coli* identified condition-specific fitness genes.

Conclusions:

  • MultiCAST significantly enhances the accessibility, speed, and throughput of bacterial genome-scale functional screens.
  • The platform facilitates the identification of genes with condition-specific fitness effects.
  • MultiCAST is adaptable for diverse bacterial species, broadening its applicability in research and biotechnology.