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

CRISPR and crRNAs

CRISPR and crRNAs

Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...

CRISPR

CRISPR

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...

You might also read

Related Articles

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

Sort by

Same author

Reduced eccDNA content in idiopathic asthenozoospermia sperm is associated with compromised DNA repair capacity and elevated DNA damage.

Cellular & molecular biology letters·2026

Same author

Energy metabolism alteration and gene expression reprogramming in a cell model of high fat load non-alcoholic fatty liver disease.

BMC gastroenterology·2026

Same author

Liquid biopsy epigenetics: establishing a molecular profile based on cell-free DNA.

Molecular oncology·2025

Same author

Diagnostic and Monitoring Strategies for VEXAS Syndrome: Evaluating Sanger Sequencing, NGS, and the SWIM-Score.

Journal of clinical immunology·2025

Same author

Spatial-Temporal Diversity of Extrachromosomal DNA Shapes Urothelial Carcinoma Evolution and the Tumor Immune Microenvironment.

Cancer discovery·2025

Same author

An Overview and Comparative Analysis of CRISPR-SpCas9 gRNA Activity Prediction Tools.

The CRISPR journal·2025

Search research articles

Related Experiment Video

Updated: Mar 21, 2026

A Customizable Protocol for String Assembly gRNA Cloning STAgR

A Customizable Protocol for String Assembly gRNA Cloning STAgR

Published on: December 26, 2018

Golden Gate Assembly of CRISPR gRNA expression array for simultaneously targeting multiple genes.

Johan Vad-Nielsen¹, Lin Lin¹, Lars Bolund¹

¹Department of Biomedicine, Aarhus University, Wilhelm Meyers Alle 4, Building 1242, 8000, Aarhus C, Denmark.

Cellular and Molecular Life Sciences : CMLS

|May 15, 2016

Summary

This study presents a new Golden Gate Assembly method for creating CRISPR gene editing arrays, enabling simultaneous targeting of multiple genes in human cells within two weeks. This breakthrough overcomes a key challenge in multiplexed CRISPR applications.

Keywords:

CRISPR Cas9 Genome editing Golden Gate Assembly Simultaneous multiple gene inhibition Simultaneous multiple gene knockout

More Related Videos

Generation of Defined Genomic Modifications Using CRISPR-CAS9 in Human Pluripotent Stem Cells

Generation of Defined Genomic Modifications Using CRISPR-CAS9 in Human Pluripotent Stem Cells

Published on: September 25, 2019

Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

Published on: January 7, 2019

Related Experiment Videos

Last Updated: Mar 21, 2026

A Customizable Protocol for String Assembly gRNA Cloning STAgR

A Customizable Protocol for String Assembly gRNA Cloning STAgR

Published on: December 26, 2018

Generation of Defined Genomic Modifications Using CRISPR-CAS9 in Human Pluripotent Stem Cells

Generation of Defined Genomic Modifications Using CRISPR-CAS9 in Human Pluripotent Stem Cells

Published on: September 25, 2019

Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

Published on: January 7, 2019

Area of Science:

Molecular Biology
Genetics
Biotechnology

Background:

CRISPR/Cas9 is a powerful genome editing tool.
Simultaneously targeting multiple genes in single cells presents a technical hurdle.

Purpose of the Study:

To develop an efficient method for generating CRISPR gRNA expression arrays for multiplexed gene targeting.
To enable simultaneous targeting of multiple genomic loci or inhibition of multiple genes.

Main Methods:

Utilized Golden Gate Assembly for constructing CRISPR gRNA expression arrays.
Designed multiplexed gRNA expression array vectors.

Main Results:

Developed a method to generate CRISPR gRNA expression arrays in 2 weeks, accommodating up to 30 gRNA expression cassettes.
Successfully demonstrated simultaneous targeting of 10 genomic loci in human cells.
Achieved simultaneous inhibition of multiple endogenous genes using the multiplexed vector.

Conclusions:

The Golden Gate Assembly method provides an efficient solution for multiplexed CRISPR gene editing.
The developed vectors facilitate simultaneous targeting and gene inhibition in human cells.
Plasmids are available via Addgene for research use.