| 敲除OsNramp5基因创制低镉优质粳稻新材料的应用评价 |
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| 引用本文： | 裴峰,王广达,高鹏,冯志明,胡珂鸣,陈宗祥,陈红旗,崔傲,左示敏.敲除OsNramp5基因创制低镉优质粳稻新材料的应用评价[J].中国水稻科学,2023,37(1):16-28. |
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| 作者姓名： | 裴峰 王广达 高鹏 冯志明 胡珂鸣 陈宗祥 陈红旗 崔傲 左示敏 |
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| 作者单位： | 1.扬州大学 江苏省作物基因组学和分子育种重点实验室/植物功能基因组学教育部重点实验室，江苏 扬州 225009;2.扬州大学 江苏省粮食作物现代产业技术协同创新中心/江苏省作物遗传生理重点实验室，江苏 扬州 225009;3.中国水稻研究所 水稻生物学国家重点实验室，杭州 311400;4.镇江禾下土农业科技有限公司，江苏 镇江 212008;5.扬州大学 教育部农业与农产品安全国际合作联合实验室，江苏 扬州 225009 |
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| 基金项目： | 江苏省重点研发计划资助项目(BE2019339);江苏省农业重大新品种创制项目(PZCZ201703);扬州市重点研发计划资助项目(现代农业YZ2020031(现代农业YZ2020031) |
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| 摘 要： | 【目的】探究在粳稻中敲除Os Nramp5基因对镉等金属元素积累、产量和品质的影响，为科学高效地生产优质健康粳米提供新材料和理论参考。【方法】利用CRISPR/Cas9技术，在优良食味高产粳稻品种南粳46背景下对OsNramp5进行基因敲除，结合标记辅助选择，获得了无外源基因的Os Nramp5敲除突变系，考查其产量和品质性状变化特征，以及在不同镉含量土壤中和锌肥、硒肥喷施后的籽粒镉、锰、硒等七种金属元素的含量变化。【结果】在T0代获得了5种OsNramp5不同变异类型的转基因植株，设计开发了其中3种突变型的特异分子标记，进而结合标记辅助选择，在各植株的T2代中获得不含潮霉素选择标记基因和CAS9蛋白基因的纯合敲除系。与野生型相比，无论在高浓度还是低浓度镉含量土壤中，敲除系整株包括籽粒中的镉含量和地上部组织中锰含量均显著低于南粳46；敲除系的株高略微下降，穗粒数显著降低并导致其单株产量明显下降，但在低分蘖肥处理中产量下降未达显著。敲除系稻米外观总体好于对照，蛋白质和直链淀粉含量明显高于对照，导致其食味值明显降低。敲除系籽粒中铜、锰和硒的含量显著低于对照，但在施用锌肥和硒肥后可以显著提高...
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| 关 键 词： | 粳稻 低镉 基因编辑 品质 产量 育种价值 |
| 收稿时间： | 2022-05-06 |
| 修稿时间： | 2022-07-31 |
Evaluation of New japonica Rice Lines with Low Cadmium Accumulation and Good Quality Generated by Knocking Out OsNramp5 |
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| PEI Feng,WANG Guangda,GAO Peng,FENG Zhiming,HU Keming,CHEN Zongxiang,CHEN Hongqi,CUI Ao,ZUO Shimin.Evaluation of New japonica Rice Lines with Low Cadmium Accumulation and Good Quality Generated by Knocking Out OsNramp5[J].Chinese Journal of Rice Science,2023,37(1):16-28. |
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| Authors: | PEI Feng WANG Guangda GAO Peng FENG Zhiming HU Keming CHEN Zongxiang CHEN Hongqi CUI Ao ZUO Shimin |
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| Abstract: | 【Objective】 It is crucial to elucidate the effects of OsNramp5 gene knockout on cadmium and other metal elements accumulation, yield and quality in japonica rice, which can provide new materials and theoretical references for scientific and efficient production of high-quality and green japonica rice.【Methods】 Using CRISPR/Cas9 technology, the OsNramp5 was knocked out under the background of Nanjing 46 (NJ46), a japonica rice variety with good taste and high yield. Combined with molecular marker-assisted selection, some OsNramp5 knockout lines without exogenous genes were obtained. The changes in yield and quality traits and seven metal elements contents such as cadmium, manganese and selenium in grains after spraying zinc fertilizer or selenium fertilizer and cultivated in soils with different cadmium content were investigated. 【Results】 Five transgenic plants with different mutations of OsNramp5 were obtained in the T0 generation, and specific molecular markers of three of them were designed. Then, combined with marker-assisted selection, homozygous knockout lines without hygromycin selective marker gene and CAS9 protein gene were obtained in the T2 generation of each plant. Compared with the wild type, the cadmium content in the whole plant including the grain of the knockout lines and the manganese content in shoots were significantly lower than those of the wild type in both high and low concentrations of cadmium soil. The plant height and grain number per panicle of the knockout lines decreased slightly, and the yield per plant decreased significantly in the knockout lines at high and medium tillering fertilizer levels while no changes were observed at the low tillering fertilizer level. The appearance of the knockout lines was better than that of the control, and the contents of protein and amylose were significantly higher than those of the control. The contents of copper, manganese and selenium in grains of knockout lines were significantly lower than those of the control NJ46, but zinc and selenium fertilizer could significantly increase the contents of zinc and selenium, and further reduce the content of cadmium in grains. Application of selenium was beneficial to rice appearance quality, but application of zinc was contrary. Spraying both fertilizers reduced the edible value of rice, worsening taste quality. The change of seed setting rate of the knockout lines under extreme high and low temperature conditions was the same as that of the control, but the resistance level to rice sheath blight was significantly reduced.【Conclusion】 We obtained marker-free OsNramp5 knockout lines under the background of japonica variety with high quality and yield, and proved that OsNramp5 was of great value in the production of low-cadmium, Se- and zinc-rich, though their yield and eating quality were decreased to different extents. |
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| Keywords: | japonica rice low cadmium accumulation gene editing grain quality grain yield breeding value |
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