Improvement of Genome Editing Efficiency in Mammalian Cells by a Refined Uni-Vector Prime Editing System.

Prime editing is an advanced technology in the CRISPR/Cas genome editing field. The original multi-vector method utilized in prime editing has less efficiency and precision and is also inherently difficult in generating homozygous mutations in mammalian cells. To overcome these technical difficulties, a Uni-vector prime editing system containing the major components for prime editing has been constructed in two all-in-one plasmids, pPE3-pPuro and pePE5max-pPuro. The Uni-vector prime editing plasmids enhance the editing efficiency of prime editing and improve the generation of homozygous mutated mammalian cell lines. The editing efficiency depends on the transfection efficiency. Remarkably, the Uni-vector ePE5max system can achieve a high editing rate even in cell lines that are traditionally difficult to transfect. This Uni-vector approach simplifies the delivery of all the necessary editing components and greatly improves the editing efficiency. This approach represents an advanced progress in the field of prime editing.