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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...

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...

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Related Experiment Video

Updated: Oct 31, 2025

Delivery of the Cas9/sgRNA Ribonucleoprotein Complex in Immortalized and Primary Cells via Virus-like Particles "Nanoblades"

Delivery of the Cas9/sgRNA Ribonucleoprotein Complex in Immortalized and Primary Cells via Virus-like Particles "Nanoblades"

Published on: March 31, 2021

Efficient Peptide-Mediated In Vitro Delivery of Cas9 RNP.

Oskar Gustafsson¹, Julia Rädler¹, Samantha Roudi¹

¹Centre for Biomolecular and Cellular Medicine, Department of Laboratory Medicine, Karolinska Institutet, 141 57 Stockholm, Sweden.

|July 2, 2021

Summary

This summary is machine-generated.

A novel cell-penetrating peptide, PepFect14 (PF14), efficiently delivers CRISPR-Cas9 ribonucleoprotein (RNP) complexes for gene editing. This non-toxic delivery method achieves high editing rates and demonstrates stability under various storage conditions.

Keywords:

CRISPR/Cas9 PepFect14 RNP cell-penetrating peptide (CPP)drug delivery gene editing nanoparticle non-viral

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Area of Science:

Molecular Biology
Biotechnology
Gene Editing

Background:

CRISPR gene editing tools offer significant clinical potential but face delivery challenges.
Efficient and non-toxic delivery of CRISPR machinery to the cell nucleus is crucial.

Purpose of the Study:

To repurpose the cell-penetrating peptide PepFect14 (PF14) for delivering Cas9 ribonucleoprotein (RNP) complexes.
To evaluate the efficiency, toxicity, and stability of this novel delivery system.

Main Methods:

Formation of Cas9-RNP complexes with PepFect14 (PF14).
Assessment of editing efficiency in HEK293T and reporter cells using restriction enzyme digest and next-generation sequencing.
Evaluation of storage stability including freeze-thaw cycles and freeze-drying.

Main Results:

Achieved high editing rates (up to 80% in HEK293T cells) at low nanomolar concentrations.
Demonstrated comparable or superior efficiency to commercial reagents (RNAiMAX, CRISPRMax) without apparent toxicity.
Cas9-RNP-CPP complexes maintained editing efficiency after various storage conditions, including freeze-drying.

Conclusions:

PepFect14 (PF14) provides an efficient, non-toxic, and stable method for Cas9 RNP delivery.
This CPP-based strategy enhances CRISPR delivery options and may be applicable to other gene editors.