Search-and-replace genome editing without double-strand breaks or donor DNA.

Area of Science:

• Molecular Biology
• Genetics
• Biotechnology

Background:

• Genetic variants are difficult to correct efficiently and without byproducts.
• Existing genome editing methods have limitations in precision and scope.

Purpose of the Study:

• To describe prime editing, a versatile and precise genome editing method.
• To demonstrate prime editing's ability to correct various genetic mutations in human cells.

Main Methods:

• Prime editing utilizes a catalytically impaired Cas9 fused to a reverse transcriptase.
• A prime editing guide RNA (pegRNA) programs the system to target specific DNA sites and encode edits.
• Performed over 175 edits in human cells, including insertions, deletions, and all 12 point mutation types.

Main Results:

• Successfully corrected genetic causes of sickle cell disease and Tay-Sachs disease.
• Installed a protective transversion in PRNP and precisely inserted tags/epitopes.
• Prime editing demonstrated higher efficiency and fewer byproducts than homology-directed repair.
• Showed lower off-target editing compared to standard Cas9 nuclease.

Conclusions:

• Prime editing expands the capabilities of genome editing.
• This method can potentially correct up to 89% of known disease-associated genetic variants.
• Prime editing offers a precise and versatile approach for genetic disease correction.