Simultaneous precise editing of multiple genes in human cells. Issue 19 (8th August 2019)

• Record Type: Journal Article
• Authors: Riesenberg, Stephan; Chintalapati, Manjusha; Macak, Dominik; Kanis, Philipp; Maricic, Tomislav; Pääbo, Svante
• Is Part Of: Nucleic acids research. Volume 47:Issue 19(2019)
• Page Start: e116
• Page End: e116
• Publication Date: 2019-08-08
• Subjects: Nucleic acids -- Periodicals; Molecular biology -- Periodicals; 572.805
• DOI: 10.1093/nar/gkz669
• Languages: English
• ISSNs: 0305-1048

Abstract

Abstract: When double-strand breaks are introduced in a genome by CRISPR they are repaired either by non-homologous end joining (NHEJ), which often results in insertions or deletions (indels), or by homology-directed repair (HDR), which allows precise nucleotide substitutions to be introduced if a donor oligonucleotide is provided. Because NHEJ is more efficient than HDR, the frequency with which precise genome editing can be achieved is so low that simultaneous editing of more than one gene has hitherto not been possible. Here, we introduced a mutation in the human PRKDC gene that eliminates the kinase activity of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). This results in an increase in HDR irrespective of cell type and CRISPR enzyme used, sometimes allowing 87% of chromosomes in a population of cells to be precisely edited. It also allows for precise editing of up to four genes simultaneously (8 chromosomes) in the same cell. Transient inhibition of DNA-PKcs by the kinase inhibitor M3814 is similarly able to enhance precise genome editing.