TY - JOUR
T1 - Fusion of the Cas9 endonuclease and the VirD2 relaxase facilitates homology-directed repair for precise genome engineering in rice
AU - Ali, Zahir
AU - Shami, Ashwag
AU - Sedeek, Khalid Elwy Mohamed
AU - Kamel, Radwa
AU - Alhabsi, Abdulrahman
AU - Tehseen, Muhammad
AU - Hassan, Norhan
AU - Butt, Haroon
AU - Kababji, Ahad
AU - Hamdan, Samir
AU - Mahfouz, Magdy M.
N1 - KAUST Repository Item: Exported on 2020-10-01
Acknowledgements: We thank members of the Laboratory for Genome Engineering and Synthetic Biology at King Abdullah University of Science and Technology for helpful discussions and comments on the manuscript. This research work is supported by King Abdullah University of Science and Technology (KAUST) baseline funding to Magdy Mahfouz
PY - 2020/1/23
Y1 - 2020/1/23
N2 - Precise genome editing by systems such as clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) requires high-efficiency homology-directed repair (HDR). Different technologies have been developed to improve HDR but with limited success. Here, we generated a fusion between the Cas9 endonuclease and the Agrobacterium VirD2 relaxase (Cas9-VirD2). This chimeric protein combines the functions of Cas9, which produces targeted and specific DNA double-strand breaks (DSBs), and the VirD2 relaxase, which brings the repair template in close proximity to the DSBs, to facilitate HDR. We successfully employed our Cas9-VirD2 system for precise ACETOLACTATE SYNTHASE (OsALS) allele modification to generate herbicide-resistant rice (Oryza sativa) plants, CAROTENOID CLEAVAGE DIOXYGENASE-7 (OsCCD7) to engineer plant architecture, and generate in-frame fusions with the HA epitope at HISTONE DEACETYLASE (OsHDT) locus. The Cas9-VirD2 system expands our ability to improve agriculturally important traits in crops and opens new possibilities for precision genome engineering across diverse eukaryotic species.
AB - Precise genome editing by systems such as clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) requires high-efficiency homology-directed repair (HDR). Different technologies have been developed to improve HDR but with limited success. Here, we generated a fusion between the Cas9 endonuclease and the Agrobacterium VirD2 relaxase (Cas9-VirD2). This chimeric protein combines the functions of Cas9, which produces targeted and specific DNA double-strand breaks (DSBs), and the VirD2 relaxase, which brings the repair template in close proximity to the DSBs, to facilitate HDR. We successfully employed our Cas9-VirD2 system for precise ACETOLACTATE SYNTHASE (OsALS) allele modification to generate herbicide-resistant rice (Oryza sativa) plants, CAROTENOID CLEAVAGE DIOXYGENASE-7 (OsCCD7) to engineer plant architecture, and generate in-frame fusions with the HA epitope at HISTONE DEACETYLASE (OsHDT) locus. The Cas9-VirD2 system expands our ability to improve agriculturally important traits in crops and opens new possibilities for precision genome engineering across diverse eukaryotic species.
UR - http://hdl.handle.net/10754/661143
UR - http://www.nature.com/articles/s42003-020-0768-9
UR - http://www.scopus.com/inward/record.url?scp=85078088411&partnerID=8YFLogxK
U2 - 10.1038/s42003-020-0768-9
DO - 10.1038/s42003-020-0768-9
M3 - Article
C2 - 31974493
SN - 2399-3642
VL - 3
JO - Communications Biology
JF - Communications Biology
IS - 1
ER -