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Open Access Green as soon as Postprint is submitted to ZB.
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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Publication type
Article: Journal article
Document type
Scientific Article
Keywords
Damage Response; Break Repair; Dna; Receptor; Protein; Kinase; Ku70
ISSN (print) / ISBN
1087-0156
e-ISSN
1546-1696
Journal
Nature Biotechnology
Quellenangaben
Volume: 33,
Issue: 5,
Pages: 543-548
Publisher
Nature Publishing Group
Publishing Place
New York, NY
Reviewing status
Peer reviewed
Institute(s)
Institute of Developmental Genetics (IDG)