利用CRISPR/Cas9系统定向改良水稻粒长和穗粒数性状  

Orientation Improvement of Grain Length and Grain Number in Rice by Using CRISPR/Cas9 System

作　　者：沈兰[1,2] 李健[3] 付亚萍[2] 王俊杰[2] 华宇峰 焦晓真[2] 严长杰[1] 王克剑[2] SHEN Lan;LI Jian;FU Yaping;WANG Junjie;HUA Yufeng;JIAO Xiaozhen;YAN Changjie;WANG Kejian(Jiangsu Key Laboratory of Crop Genetics and Physiology/Co-Innovation Center for Modern Production Technology of Grain Crops/Key Laboratory of Plant Functional Genomics of the Ministry of Education/College of Agronomy, Yangzhou University, Yangzhou 225009, China;State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006, China;Lianyungang Academy of Agricultural Sciences; Lianyungang 222000, China)

机构地区：[1]扬州大学农学院/江苏省作物遗传生理国家重点实验室培育点/江苏省现代粮食作物生产协同创新中心/教育部植物功能基因组学重点实验室,江苏扬州225009 [2]中国水稻研究所水稻生物学国家重点实验室,杭州310006 [3]连云港市农业科学院,江苏连云港222000

出　　处：《中国水稻科学》2017年第3期223-231,共9页Chinese Journal of Rice Science

基　　金：国家自然科学基金资助项目(31371233);高校自然科学基金重大项目(14KJA210002);中国农业科学院科技创新工程资助项目;江苏省农业科技自主创新资金资助项目[CX(13)5075]

摘　　要：【目的】基因组定点编辑技术已成为分子育种的重要手段。本研究拟对GS3和Gn1a功能缺失突变对目标性状的改良效应进行分析,以期为培育高产水稻提供理论基础。【方法】利用CRISPR/Cas9系统,以控制粒型基因GS3和控制每穗粒数基因Gn1a为编辑对象,构建了共敲除载体p C1300-2×35S::Cas9-g^(GS3)-g^(Gn1a)a,用农杆菌介导法转化4个优质水稻品种,分析了基因突变的特征和相应农艺性状。【结果】构建的敲除载体成功地实现了对GS3和Gn1a基因的定点编辑。在4个转化受体的T_0代均分别获得了gs3和gs3gn1a的移码突变体。对T_1 代中无选择标记突变体的农艺性状分析表明,突变体gs3和gs3gn1a与野生型相比粒长变长,千粒重增加;突变体gs3gn1a与突变体gs3相比,每穗粒数显著增加。【结论】利用CRISPR/Cas9系统进行水稻基因编辑可以快速改良品种的目标性状,在水稻品种的定向改良方面具有巨大的潜力。【Objective】Gene orientation editing has become an important way for molecular breeding.We evaluated the improvement effects on the target traits following the construction of GS3and Gn1a loss-of-function mutants so as to lay a solid foundation for high-yielding rice breeding.【Method】GS3and Gn1a were selected as targets for gene editing,which control the grain size and grain number in rice,respectively.The pC1300-2×35S::Cas9-gGS3-gGn1a expression vector was constructed for knocking out both GS3and Gn1a by using CRISPR/Cas9system,and transformed into four good quality rice varieties by the Agrobacterium-mediated method.And the properties of mutations and the target traits were analyzed in the transgenic lines.【Result】The sequencing results showed the GS3and Gn1a in four rice varieties were successfully edited.In T0generation,we obtained mutants with frame shift mutations in GS3and Gn1a in four rice varieties.In T1generation,the marker-free plants were screened for analyzing the agronomic traits in four genetic backgrounds.For the agronomic characters,the gs3and gs3gn1a mutants had the longer grain length and the increased1000-grain weight compared to the wild type,and the gs3gn1a mutants had more grains per panicle compared to the gs3mutants.【Conclusion】CRISPR/Cas9-mediated gene editing can improve rice target traits,which has great potential in orientation improvement of rice varieties.

关 键 词：CRISPR/Cas9 基因编辑 水稻 GS3 Gn1a 

分 类 号：S511[农业科学—作物学]