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Updated: Apr 14, 2026

A Standard Methodology to Examine On-site Mutagenicity As a Function of Point Mutation Repair Catalyzed by CRISPR/Cas9 and SsODN in Human Cells

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Efficient genome editing with chimeric oligonucleotide-directed editing.

Long T Nguyen^1,2, Noah R Rakestraw³, Brianna L M Pizzano⁴

¹Department of Molecular Biology, Princeton University, Princeton, NJ, USA.

Nature Communications

|April 12, 2026

Summary

This summary is machine-generated.

A new genome editing system, chimeric oligonucleotide-directed editing (CODE), offers precise gene editing. This CODE system, including CODEMax(exo+), shows improved efficiency in cell lines and successfully modifies embryos without double-strand breaks.

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Mouse Genome Engineering Using Designer Nucleases

Mouse Genome Engineering Using Designer Nucleases

Published on: April 2, 2014

Area of Science:

Molecular Biology
Genetics
Biotechnology

Background:

Prime editing represents a significant advancement in genome editing precision.
Existing Cas9-based methods have limitations that newer approaches aim to overcome.

Purpose of the Study:

To develop and characterize a novel, efficient two-component genome editing system called CODE (chimeric oligonucleotide-directed editing).
To engineer fusion proteins of nCas9 and DNA polymerase for enhanced editing capabilities.
To evaluate the performance of CODE and its variant, CODEMax(exo+), in various cell types and embryos.

Main Methods:

Creation of nCas9-DNA polymerase fusion proteins and chimeric pegRNAs (cpegRNAs).
Engineering of Bst DNA polymerase for improved thermostability and processivity.
Incorporation of 5' to 3' exonuclease activity in CODEMax(exo+) for enhanced strand invasion.
Testing editing efficiency for small insertions, deletions, and substitutions in HEK293T cells using plasmid and RNP delivery.
Assessing precise editing rates in mouse and bovine embryos.

Main Results:

CODE systems demonstrated improved efficiency for small insertions, deletions, and substitutions compared to PEMax in HEK293T cells.
CODEMax(exo+) achieved up to 9.3% precise editing in mouse and bovine embryos.
The CODE system effectively performs genome editing without inducing double-stranded breaks.

Conclusions:

CODE and CODEMax(exo+) represent a valuable addition to the genome editing toolbox, offering enhanced precision and efficiency.
These systems expand the possibilities for genome manipulation, particularly in challenging contexts like embryos.
Further optimization holds promise for even greater editing outcomes in diverse applications.