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

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

Single-Cell Transcriptomics and Mendelian Randomization Analysis Reveal Key Genes in Atrial Fibrillation.

Journal of cardiovascular electrophysiology·2026
Same author

Sensitive detection of PFOA in environmental water based on Pt/MnO<sub>2</sub> nanozymes with engineered d-band structure.

Journal of hazardous materials·2026
Same author

Monocyte Dll4 Contributes to Systemic Inflammation in People Living With HIV.

Circulation research·2026
Same author

Lignin nanoparticles: Preparation strategies, surface engineering, and emerging applications-A review.

International journal of biological macromolecules·2026
Same author

A feeder-free culture system supports long-term expansion and germline competence of bovine formative embryonic stem cells†.

Biology of reproduction·2026
Same author

Exploration and validation of potential diagnostic biomarkers of PANoptosis in periodontitis using bioinformatics approaches.

Biochemical and biophysical research communications·2026

Related Experiment Video

Updated: Jul 1, 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

Scaffold-Proximal DNA Extensions Enhance Cas12a Trans-cleavage for Direct and Broad-Scope Nucleic Acid Detection.

Wenjuan Xin1, Zijie Tang1, Shumin Wang1

  • 1Key Laboratory of Optic-electric Sensing and Analytical Chemistry for Life Science, MOE, Shandong Key Laboratory of Biochemical Analysis, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, China.

Analytical Chemistry
|February 16, 2026
PubMed
Summary
This summary is machine-generated.

We developed a novel DNA activator for CRISPR/Cas12a diagnostics, enabling direct RNA detection without reverse transcription. This Proximal-Extended Activator (PEA) system enhances sensitivity and specificity for microRNAs, mRNAs, and SNPs.

More Related Videos

Using Next Generation Sequencing to Identify Mutations Associated with Repair of a CAS9-induced Double Strand Break Near the CD4 Promoter
06:59

Using Next Generation Sequencing to Identify Mutations Associated with Repair of a CAS9-induced Double Strand Break Near the CD4 Promoter

Published on: March 31, 2022

2.9K
Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a
09:03

Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a

Published on: December 23, 2022

3.2K

Related Experiment Videos

Last Updated: Jul 1, 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
Using Next Generation Sequencing to Identify Mutations Associated with Repair of a CAS9-induced Double Strand Break Near the CD4 Promoter
06:59

Using Next Generation Sequencing to Identify Mutations Associated with Repair of a CAS9-induced Double Strand Break Near the CD4 Promoter

Published on: March 31, 2022

2.9K
Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a
09:03

Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a

Published on: December 23, 2022

3.2K

Area of Science:

  • Molecular Biology
  • Biotechnology
  • Diagnostic Technology

Background:

  • CRISPR/Cas12a systems offer powerful nucleic acid diagnostics but face limitations for direct RNA detection, including the need for reverse transcription, lower sensitivity, and poor performance with diverse RNA targets like microRNAs (miRNAs) and messenger RNAs (mRNAs).
  • Existing CRISPR diagnostics struggle with detecting single nucleotide polymorphisms (SNPs) and require amplification steps, limiting their direct applicability.

Purpose of the Study:

  • To engineer a novel DNA activator architecture for the CRISPR/Cas12a system to enhance its trans-cleavage activity and expand its utility for direct RNA detection.
  • To overcome limitations of sensitivity, amplification requirements, and target compatibility in CRISPR-based diagnostics.

Main Methods:

  • Development of a rationally engineered DNA activator architecture, specifically a Proximal-Extended Activator (PEA), with a focus on the positional dependence of activator extensions.
  • Investigating the mechanism of Cas12a activation by PEA, including its impact on ribonucleoprotein (RNP) complex stabilization and interdomain motions.
  • Utilizing a Split-PEA format for enhanced SNP discrimination.

Main Results:

  • The PEA system significantly boosted Cas12a trans-cleavage activity, enabling direct, amplification-free RNA detection with high sensitivity (1.3 fM for miRNA, 93 fM for mRNA) without reverse transcription.
  • Positional dependence of activator extensions was critical: scaffold-proximal extensions enhanced activation, while distal extensions were inhibitory due to steric hindrance.
  • The Split-PEA format demonstrated exceptional SNP discrimination, successfully identifying the EGFR T790 M mutation at a 0.1% allelic frequency.

Conclusions:

  • The engineered PEA system provides a facile and versatile platform for highly sensitive and specific CRISPR-based diagnostics of diverse nucleic acid targets, including RNA and SNPs.
  • This approach overcomes significant hurdles in CRISPR diagnostics, enabling direct detection without reverse transcription and improving performance for challenging targets.
  • The mechanistic understanding of activator design offers a pathway for further optimization of CRISPR diagnostic tools.