High Frequency Targeted Mutagenesis Using Engineered Endonucleases and DNA-End Processing Enzymes

• DOI: 10.1371/journal.pone.0053217
• Publication Date: 2013-01-24T22:42:30Z
• Authors: Fabien Delacôte; Christophe Perez; Valérie Guyot; Marianne Duhamel; Christelle Rochon; Nathalie Ollivier; Rachel Macmaster; George H. Silva; Frédéric Pâques; Fayza Daboussi; Philippe Duchateau
• Keywords: Gene targeting

Targeting DNA double-strand breaks is a powerful strategy for gene inactivation applications. Without the use of repair plasmid, targeted mutagenesis can be achieved through Non-Homologous End joining (NHEJ) pathways. However, many produced by engineered nucleases may subject to precise re-ligation without loss genetic information and thus are likely unproductive. In this study, we combined endonucleases DNA-end processing enzymes increase efficiency mutagenesis, providing robust efficient method (i) greatly improve frequency up 30-fold, and; (ii) control nature mutagenic events using meganucleases in conjunction with human primary cells.