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Related Experiment Video

Updated: Jun 4, 2025

Enhanced Genome Editing with Cas9 Ribonucleoprotein in Diverse Cells and Organisms

Enhanced Genome Editing with Cas9 Ribonucleoprotein in Diverse Cells and Organisms

Published on: May 25, 2018

Improved split prime editors enable efficient in vivo genome editing.

Rongwei Wei¹, Zhenxing Yu², Lihong Ding¹

¹Westlake Genetech, Ltd., No. 1 Yunmeng Road, Cloud Town, Hangzhou 310024, China.

|January 2, 2025

Summary

This summary is machine-generated.

Researchers developed a new split prime editor (PE) system delivered via dual adeno-associated viruses (AAVs). This novel split-PE architecture shows high in vivo editing efficiency, advancing gene therapy potential.

Keywords:

CP: Genomics PCSK9 editing dual AAV split prime editors

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Area of Science:

Molecular Biology
Gene Editing Technologies
Biotechnology

Background:

Efficient in vivo delivery of prime editors (PEs) is crucial for gene therapy and disease modeling.
Dual adeno-associated virus (AAV) delivery of split PEs shows variable editing efficiency depending on the chosen split sites.
Limited efficient split sites exist within the Cas9 nickase (Cas9n) component of PEs.

Purpose of the Study:

To identify and validate efficient split sites for dual AAV-mediated PE delivery.
To develop a novel split-PE architecture with improved Cas9n cleavage efficiency.
To assess the in vivo editing efficiency of the optimized split-PE system.

Main Methods:

Verification of the 1115 (Asn) split site for dual AAV-delivered PEs.
Design of split sites within the Cas9n moiety, detaching the reverse transcriptase.
Packaging of the optimized split-ePE3-367 into AAV9 vectors for in vivo testing.

Main Results:

The 1115 (Asn) site was confirmed as an efficient split site for dual AAV-delivered PEs.
The novel split-PE-367 architecture demonstrated high editing efficiency, particularly with Rma intein.
In vivo testing in mice showed 17.5% precise editing using split-ePE3-367 delivered via AAV9.

Conclusions:

An alternative split-PE architecture has been established, enhancing editing efficiency.
This optimized system provides a robust tool for gene editing applications.
The findings facilitate advancements in disease modeling and therapeutic gene correction strategies.