Rapid genome editing by CRISPR-Cas9-POLD3 fusion

eLife

Salla Keskitalo¹  Markku Varjosalo¹  Monika Szymanska²  Ganna Reint²  Katariina Mamia²  Xian Hu²  Zhuokun Li²  Artur Cieslar-Pobuda³  Judith Staerk⁴  Inkeri Soppa⁵  Emma Haapaniemi⁶  Diana L Bordin⁷  Susanne Lorenz⁸  Leonardo A Meza-Zepeda⁸  Eivind Valen⁹  Kornel Labun⁹  Bernhard Schmierer^1,10  Jussi Taipale^1,11  Eero Tolo^1,12 

[1] Center for Biotechnology, University of Helsinki, Helsinki, Finland;Centre for Molecular Medicine Norway, University of Oslo, Oslo, Norway;Centre for Molecular Medicine Norway, University of Oslo, Oslo, Norway;Department of Cancer Immunology, Institute of Cancer Research, Oslo University Hospital, Oslo, Norway;Centre for Molecular Medicine Norway, University of Oslo, Oslo, Norway;Department of Haematology, Oslo University Hospital, Oslo, Norway;Centre for Molecular Medicine Norway, University of Oslo, Oslo, Norway;Stem Cells and Metabolism Research Program, Faculty of Medicine, University of Helsinki, Oslo, Finland;Centre for Molecular Medicine Norway, University of Oslo, Oslo, Norway;Stem Cells and Metabolism Research Program, Faculty of Medicine, University of Helsinki, Oslo, Finland;Department of Pediatrics, Oslo University Hospital, Oslo, Norway;Department of Clinical Molecular Biology, Akershus University Hospital, Oslo, Norway;Department of Core Facilities, Institute for Cancer Research, Oslo University Hospital, Oslo, Norway;Department of Informatics, Computational Biology Unit, University of Bergen, Bergen, Norway;Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden;Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden;Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom;Genome-Scale Biology Program, University of Helsinki, Oslo, Norway;Faculty of Social Sciences, University of Helsinki, Oslo, Finland;

关键词: CRISPR-Cas9;    gene editing;    molecular biology;    Other;   

DOI  :  10.7554/eLife.75415

来源: eLife Sciences Publications, Ltd

【 摘 要 】

Precision CRISPR gene editing relies on the cellular homology-directed DNA repair (HDR) to introduce custom DNA sequences to target sites. The HDR editing efficiency varies between cell types and genomic sites, and the sources of this variation are incompletely understood. Here, we have studied the effect of 450 DNA repair protein-Cas9 fusions on CRISPR genome editing outcomes. We find the majority of fusions to improve precision genome editing only modestly in a locus- and cell-type specific manner. We identify Cas9-POLD3 fusion that enhances editing by speeding up the initiation of DNA repair. We conclude that while DNA repair protein fusions to Cas9 can improve HDR CRISPR editing, most need to be optimized to the cell type and genomic site, highlighting the diversity of factors contributing to locus-specific genome editing outcomes.

【 授权许可】

CC BY