Development and testing of a versatile genome editing application reporter (V-GEAR) system

Area of Science:

• Molecular Biology
• Genetics
• Biotechnology

Background:

• CRISPR-Cas9 and related technologies have transformed genome editing.
• Efficacy of genome editing tools depends on variants, guide RNAs, and DNA context.
• Current screening methods lack flexibility for comparing diverse editing strategies.

Purpose of the Study:

• To develop a versatile and rapid screening system for evaluating genome editing reagents and designs.
• To enable direct comparison of different genome editing approaches like base editing, prime editing, and HDR.
• To facilitate the acceleration of genome editing technology development.

Main Methods:

• Developed the versatile genome editing application reporter (V-GEAR) system.
• V-GEAR utilizes a detectable synthetic cell surface protein (RQR8) and customizable target sequences.
• Validated V-GEAR with base editors, prime editors, and Cas9-mediated homology-directed repair.

Main Results:

• V-GEAR allows for reliable identification and enrichment of cells with desired edits.
• The system demonstrated versatility in transient screening or stable integration at the AAVS1 safe harbor locus.
• Immunomagnetic isolation enabled rapid V-GEAR system application.

Conclusions:

• The V-GEAR system provides a flexible and efficient platform for screening genome editing technologies.
• This system accelerates the development and testing of novel genome editing approaches.
• V-GEAR facilitates direct comparative analysis of base editing, prime editing, and HDR.