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Increasing the efficiency of homology-directed repair for CRISPR-Cas9-induced precise gene editing in mammalian cells.
Nat. Biotechnol. 33, 543-548 (2015)
The insertion of precise genetic modifications by genome editing tools such as CRISPR-Cas9 is limited by the relatively low efficiency of homology-directed repair (HDR) compared with the higher efficiency of the nonhomologous end-joining (NHEJ) pathway. To enhance HDR, enabling the insertion of precise genetic modifications, we suppressed the NHEJ key molecules KU70, KU80 or DNA ligase IV by gene silencing, the ligase IV inhibitor SCR7 or the coexpression of adenovirus 4 E1B55K and E4orf6 proteins in a 'traffic light' and other reporter systems. Suppression of KU70 and DNA ligase IV promotes the efficiency of HDR 4-5-fold. When co-expressed with the Cas9 system, E1B55K and E4orf6 improved the efficiency of HDR up to eightfold and essentially abolished NHEJ activity in both human and mouse cell lines. Our findings provide useful tools to improve the frequency of precise gene modifications in mammalian cells.
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Publikationstyp
Artikel: Journalartikel
Dokumenttyp
Wissenschaftlicher Artikel
Schlagwörter
Damage Response; Break Repair; Dna; Receptor; Protein; Kinase; Ku70
ISSN (print) / ISBN
1087-0156
e-ISSN
1546-1696
Zeitschrift
Nature Biotechnology
Quellenangaben
Band: 33,
Heft: 5,
Seiten: 543-548
Verlag
Nature Publishing Group
Verlagsort
New York, NY
Nichtpatentliteratur
Publikationen
Begutachtungsstatus
Peer reviewed
Institut(e)
Institute of Developmental Genetics (IDG)