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Article

I-SceI Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System

1
Institute of Agrobiological Sciences, NARO, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, Japan
2
Gene Research Center, Ibaraki University, 3-21-1 Ami, Inashiki, Ibaraki 300-0393, Japan
*
Author to whom correspondence should be addressed.
Plants 2020, 9(9), 1070; https://doi.org/10.3390/plants9091070
Received: 30 July 2020 / Revised: 13 August 2020 / Accepted: 18 August 2020 / Published: 20 August 2020
(This article belongs to the Special Issue Plant Systems and Application)
Xanthomonas campestris is one of bacteria carrying a type III secretion system which transports their effector proteins into host plant cells to disturb host defense system for their infection. To establish a genome editing system without introducing any foreign gene, we attempted to introduce genome editing enzymes through the type III secretion system. In a test of protein transfer, X. campestris pv. campestris (Xcc) transported a considerable amount of a reporter protein sGFP-CyaA into tobacco plant cells under the control of the type III secretion system while maintaining cell viability. For proof of concept for genome editing, we used a reporter tobacco plant containing a luciferase (LUC) gene interrupted by a meganuclease I-SceI recognition sequence; this plant exhibits chemiluminescence of LUC only when a frameshift mutation is introduced at the I-SceI recognition site. Luciferase signal was observed in tobacco leaves infected by Xcc carrying an I-SceI gene which secretes I-SceI protein through the type III system, but not leaves infected by Xcc carrying a vector control. Genome-edited tobacco plant could be regenerated from a piece of infected leaf piece by repeated selection of LUC positive calli. Sequence analysis revealed that the regenerated tobacco plant possessed a base deletion in the I-SceI recognition sequence that activated the LUC gene, indicating genome editing by I-SceI protein transferred through the type III secretion system of Xcc. View Full-Text
Keywords: bacterial type III secretion system; Xanthomonas campestris; I-SceI; protein transportation; genome editing bacterial type III secretion system; Xanthomonas campestris; I-SceI; protein transportation; genome editing
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MDPI and ACS Style

Yanagawa, Y.; Takeuchi, K.; Endo, M.; Furutani, A.; Ochiai, H.; Toki, S.; Mitsuhara, I. I-SceI Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System. Plants 2020, 9, 1070. https://doi.org/10.3390/plants9091070

AMA Style

Yanagawa Y, Takeuchi K, Endo M, Furutani A, Ochiai H, Toki S, Mitsuhara I. I-SceI Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System. Plants. 2020; 9(9):1070. https://doi.org/10.3390/plants9091070

Chicago/Turabian Style

Yanagawa, Yuki, Kasumi Takeuchi, Masaki Endo, Ayako Furutani, Hirokazu Ochiai, Seiichi Toki, and Ichiro Mitsuhara. 2020. "I-SceI Endonuclease-Mediated Plant Genome Editing by Protein Transport through a Bacterial Type III Secretion System" Plants 9, no. 9: 1070. https://doi.org/10.3390/plants9091070

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