利用CRISPR/Cas9技术创制抗稻瘟病水稻

doi:10.3969/j.issn.1674-7968.2025.12.015

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Abstract Rice (Oryza sativa) blast is one of the most important diseases that threatens rice yield stability. In recent years, the incidence and severity of rice blast have been increasing in Northeast China. The inactivation of the function of Bsr-d1 (broad spectrum resistance digul) can weaken the degradation of hydrogen peroxide and improve rice resistance to Magnaporthe oryzae; Bsr-k1 (broad-spectrum resistance Kitaake-1) encodes a TPR protein, and its functional inactivation mutant bsr-k1 leads to the enrichment of OsPAL (phenylalanine ammonia-lyase) family gene mRNA and increased lignin synthesis, thereby enhancing the immune response and giving rice broad-spectrum resistance. The study designed targets for the first exon of the 2 broad-spectrum disease resistance genes Bsr-dl and the third exon of Bsr-kl and constructed gene editing vectors. Agrobacterium tumefaciens-mediated rice transformation technology was used to introduce the gene editing vector into the recipient rice variety 'TH899', and T0 generation plants with various different variants of the Bsr-dl and Bsr-kl genes were obtained. Analysis of the T₂ generation gene-edited materials that did not contain transgenic components showed that both single and simultanecous mutations of the Bsr-dl and Bsr-kl genes showed enhanced resistance to rice blast, among which the selected double-gene-edited rice materials had significantly better disease resistance than single-gene-edited materials. In addition, the single-plant yield and seed length-to-width ratio of some gene-edited lines also changed. In summary, this study used gene editing technology to simultaneously edit the Bst-dl and Bsr-kl genes, which was able to significantly enhance the resistance of 'T1899' to rice blast more than single-gene editing, and the yield traits of some mutants were also changed. The results of this study showed that combinatorial editing of rice blast resistance genes regulating different physiological pathways could effectively improve rice disease resistance. This study demonstrates the great potential of multi-gene editing in improving rice disease resistance and provides new material resources for breeding rice blast-resistant japonica rice.

Key words： Rice blast Disease resistance Rice Gene editing

Received: 06 March 2025

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	Articles by authors
	LI Yan-Hao
	DUAN Yi-Tong
	YIN Xiao-Yu
	XU Ya-Feng
	XU Yang
	YU Xiao-Ming

Cite this article:

LI Yan-Hao,DUAN Yi-Tong,YIN Xiao-Yu, et al. Using CRISPR/Cas9 Technology to Create Blast-resistant Rice (Oryza sativa)[J]. 农业生物技术学报, 2025, 33(12): 2722-2730.

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https://journal05.magtech.org.cn/Jwk_ny/EN/10.3969/j.issn.1674-7968.2025.12.015 OR https://journal05.magtech.org.cn/Jwk_ny/EN/Y2025/V33/I12/2722

[1] 陈晓军, 惠建, 马洪文, 等. 2021. 广谱抗稻瘟病基因BSR-d1在75份北方粳稻品种中的多态性分布与表达分析[J]. 种子, 40(11): 113-116+131.
(Chen X J, Hui J, Ma H W, et al.2021. Polymorphism distribution and expression analysis of broad-spectrum blast resistance gene BSR-d1 in 75 northern Japonica rice varieties[J]. Seed, 40(11): 113-116+131.)
[2] 韩政宏, 段宇轩, 徐善斌, 等. 2022. 利用CRISPR/Cas9技术敲除GS3和GS9基因改良水稻粒型性状[J]. 华北农学报, 37(02): 9-17.
(Han Z H, Duan Y X, Xu S B, et al.2022. Improvement of grain shape in rice by knocking GS3 and GS9 via CRISPR/Cas9 system[J]. Acta Agriculturae Boreali-Sinica, 37(02): 9-17.)
[3] 李彦利, 孟令君, 贾玉敏,等. 2016. 水稻新品种通禾899选育报告[J]. 北方水稻, 46(04): 54-55.
(Li Y L, Meng L J, Jia Y M, et al.2016. Breeding report on the new rice variety Tonghe-899[J]. Northern Rice, 46(04): 54-55.)
[4] 毛洧, 陈学伟, 王静. 2022. 水稻抗稻瘟病机制的研究进展[J]. 中国科学: 生命科学, 52(10): 1495-1510.
(Mao W, Chen X W, Wang J.2022. Breeding report on the new rice variety Tonghe-899[J]. Scientia Sinica (Vitae), 52(10): 1495-1510.)
[5] 王芳权, 范方军, 李文奇,等. 2016. 利用CRISPR/Cas9技术敲除水稻Pi21基因的效率分析[J]. 中国水稻科学, 30(05): 469-478.
(Wang F Q, Fan F J, Li W Q, et al.2016. Knock-out efficiency analysis of Pi21 gene using CRISPR/Cas9 in rice[J]. Chinese Journal of Rice Science, 30(05): 469-478.)
[6] 王军, 赵婕宇, 许扬, 等. 2018. 水稻稻瘟病抗性基因Bsr-d1功能标记的开发和利用[J]. 作物学报, 44(11): 1612-1620.
(Wang J, Zhao J Y, Xu Y, et al.2018. Development and application of functional markers for rice blast resistance gene Bsr-d1 in rice[J]. Acta Agronomica Sinica, 44(11): 1612-1620.)
[7] 王亚, 王越涛, 申关望, 等. 2022. 聚合R基因Pigm和非R基因bsr-d1改良水稻稻瘟病抗性[J]. 华北农学报, 37(05): 157-165.
(Wang Y, Wang Y T, Shen G W, et al.2022. Improvement of rice blast resistance by Pyramiding the R gene Pigm and the non-R gene bsr-d1[J]. Acta Agriculturae Boreali-Sinica, 37(05): 157-165.)
[8] 曾栋昌, 马兴亮, 谢先荣,等. 2018. 植物CRISPR/Cas9多基因编辑载体构建和突变分析的操作方法[J]. 中国科学: 生命科学, 48(7): 783-794.
(Zeng D C, Ma X L, Xie X R, et al.2018. A protocol for CRISPR/Cas9-based multi-gene editing and sequence decoding of mutant sites in plants[J]. Scientia Sinica (Vitae), 48(7): 783-794.)
[9] 张海旺, 房文文, 刘翠翠, 等. 2014. 离体水稻叶片划伤接种鉴定稻瘟菌的致病型[J]. 植物保护, 40(05): 121-125.
(Zhang H W, Fang W W, Liu C C, et al.2014. Identification of Magnaporthe oryzae pathotypes by wounding inoculation of detached rice leaves[J]. Plant Protection, 40(05): 121-125.)
[10] Belhaj K, Chaparro-Garcia A, Kamoun S, et al.2015. Editing plant genomes with CRISPR/Cas9[J]. Current Opinion in Biotechnology, 32: 76-84.
[11] Chen X, Shang J, Chen D, et al.2006. A B-lectin receptor kinase gene conferring rice blast resistance[J]. The Plant Joural, 46(5): 794-804.
[12] Cho S W, Kim S, Kim J M, et al.2013. Targeted genome engineering in human cells with the Cas9 RNA-guided endonuclease[J]. Nature Biotechnology, 31(3): 230-232.
[13] Huang Y, Cui K, Zhang Z, et al.2023. Identification and fine-mapping of quantitative trait loci (QTL) conferring rice false smut resistance in rice[J]. Journal of Genetics and Genomics, 50(4): 276-279.
[14] Li W, Zhu Z, Chern M, et al.2017. A natural allele of a transcription factor in rice confers broad-spectrum blast resistance[J]. Cell, 170(1): 114-126.e115.
[15] Ma X, Zhang Q, Zhu Q, et al.2015. A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants[J]. Molecular Plant, 8(8): 1274-1284.
[16] Shi X, Long Y, He F, et al.2018. The fungal pathogen Magnaporthe oryzae suppresses innate immunity by modulating a host potassium channel[J]. PLOS Pathogens, 14(1): e1006878.
[17] Zhang C, Fang H, Wang J, et al.2024. The rice E3 ubiquitin ligase-transcription factor module targets two trypsin inhibitors to enhance broad-spectrum disease resistance[J]. Developmental Cell, 59(15): 2017-2033.e2015.
[18] Zhao H, Wang X, Jia Y, et al.2018. The rice blast resistance gene Ptr encodes an atypical protein required for broad-spectrum disease resistance[J]. Nature Communications, 9(1): 2039.
[19] Zhou X, Liao H, Chern M, et al.2018. Loss of function of a rice TPR-domain RNA-binding protein confers broad-spectrum disease resistance[J]. Proceedings of the National Academy of Sciences of the USA, 115(12): 3174-3179.
[20] Zhu Z, Yin J, Chern M, et al.2020. New insights into bsr-d1-mediated broad-spectrum resistanceto rice blast[J]. Molecular Plant Pathology, 21(7): 951-960.