Generation of AAVS1 integrated doxycycline-inducible CRISPR-Prime Editor human induced pluripotent stem cell line

Prime editing uses the Cas9 nickase fused to a reverse transcriptase to copy a DNA sequence into a specific locus from a 'prime editing' guide RNA (pegRNA), eliminating the need for double-stranded DNA breaks and donor DNA templates. To facilitate prime editing in human induced pluripotent stem cells (iPSCs), we integrated a doxycycline-inducible Prime Editor protein (PE2) into the AAVS1 genomic safe harbor locus. Prime editing of iPSCs resulted in precise insertion of three nucleotides in HEK3 locus with high efficiency, demonstrating the utility of this approach. This engineered cell line can be used to edit a single or multiple genomic loci by introducing a target-specific pegRNA for precise and effective genome editing to facilitate disease modeling and functional genetics studies.

Automated summary

Prime editing involves using a fused Cas9 nickase and reverse transcriptase to copy a DNA sequence into a specific locus without the need for double-stranded DNA breaks or donor DNA templates. By integrating a doxycycline-inducible Prime Editor protein into safe harbor genomic loci, precise nucleotide insertions can be achieved in human induced pluripotent stem cells with high efficiency, showing promise for disease modeling and functional genetics studies.