- 著者
- Shuhei Yasumoto Satoru Sawai Hyoung Jae Lee Masaharu Mizutani Kazuki Saito Naoyuki Umemoto Toshiya Muranaka
- 出版者
- Japanese Society for Plant Biotechnology
- 雑誌
- Plant Biotechnology (ISSN:13424580)
- 巻号頁・発行日
- vol.37, no.2, pp.205-211, 2020-06-25 (Released:2020-06-25)
- 参考文献数
- 24
- 被引用文献数
- 22
Genome editing using site-specific nucleases, such as transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeat–CRISPR-associated protein 9 (CRISPR-Cas9), is a powerful technology for crop breeding. For plant genome editing, the genome-editing reagents are usually expressed in plant cells from stably integrated transgenes within the genome. This requires crossing processes to remove foreign nucleotides from the genome to generate null segregants. However, in highly heterozygous plants such as potato, the progeny lines have different agronomic traits from the parent cultivar and do not necessarily become elite lines. Agrobacteria can transfer exogenous genes on T-DNA into plant cells. This has been used both to transform plants stably and to express the genes transiently in plant cells. Here, we infected potato, with Agrobacterium tumefaciens harboring TALEN-expression vector targeting sterol side chain reductase 2 (SSR2) gene and regenerated shoots without selection. We obtained regenerated lines with disrupted-SSR2 gene and without transgene of the TALEN gene, revealing that their disruption should be caused by transient gene expression. The strategy using transient gene expression by Agrobacterium that we call Agrobacterial mutagenesis, developed here should accelerate the use of genome-editing technology to modify heterozygous plant genomes.
- 著者
- Naoyuki Umemoto Shuhei Yasumoto Muneo Yamazaki Kenji Asano Kotaro Akai Hyoung Jae Lee Ryota Akiyama Masaharu Mizutani Yozo Nagira Kazuki Saito Toshiya Muranaka
- 出版者
- Japanese Society for Plant Biotechnology
- 雑誌
- Plant Biotechnology (ISSN:13424580)
- 巻号頁・発行日
- vol.40, no.3, pp.211-218, 2023-09-25 (Released:2023-09-25)
- 参考文献数
- 19
- 被引用文献数
- 1
Genome editing is highly useful for crop improvement. The method of expressing genome-editing enzymes using a transient expression system in Agrobacterium, called agrobacterial mutagenesis, is a shortcut used in genome-editing technology to improve elite varieties of vegetatively propagated crops, including potato. However, with this method, edited individuals cannot be selected. The transient expression of regeneration-promoting genes can result in shoot regeneration from plantlets, while the constitutive expression of most regeneration-promoting genes does not result in normally regenerated shoots. Here, we report that we could obtain genome-edited potatoes by positive selection. These regenerated shoots were obtained via a method that combined a regeneration-promoting gene with the transient expression of a genome-editing enzyme gene. Moreover, we confirmed that the genome-edited potatoes obtained using this method did not contain the sequence of the binary vector used in Agrobacterium. Our data have been submitted to the Japanese regulatory authority, the Ministry of Education, Culture, Sports, Science and Technology (MEXT), and we are in the process of conducting field tests for further research on these potatoes. Our work presents a powerful method for regarding regeneration and acquisition of genome-edited crops through transient expression of regeneration-promoting gene.