水稻短根突变体ksr8的表型分析和基因克隆  

Phenotypic Analysis and Gene Cloning of a Short Root Mutant ksr8 in Rice (Oryza sativa L.)

作　　者：林静霞 朱俊兆 虞洁 陈星月 李浩然 郑文娟[1] 朱世华[1] 丁沃娜[1] LIN Jingxia;ZHU Junzhao;YU Jie;CHEN Xingyue;LI Haoran;ZHENG Wenjuan;ZHU Shihua;DING Wona(College of Science and Technology,Ningbo University,Ningbo,Zhejiang 315212; School of Marine Science,Ningbo University,Ningbo,Zhejiang 315211)

机构地区：[1]宁波大学科学技术学院,浙江宁波315212 [2]宁波大学海洋学院,浙江宁波315211

出　　处：《核农学报》2020年第7期1378-1386,共9页Journal of Nuclear Agricultural Sciences

基　　金：浙江省自然科学基金(LY17C020002);宁波市自然科学基金(2017A610291、2017A610292)。

摘　　要：本研究前期由籼稻品种Kasalath经EMS诱变获得一个短根突变体ksr8,为揭示该突变体根系发育的分子机制,对其进行了表型鉴定、遗传分析、基因定位、转基因互补验证以及外源精氨酸处理。表型分析表明,ksr8幼苗的株高、主根、侧根和不定根长度均变短,成熟期植株较野生型矮小、分蘖数与每穗实粒数都减少。根尖树脂半超薄切片及醋酸洋红染色显示,ksr8的短根表型与伸长区细胞变短和根尖细胞分裂有关。遗传分析结果表明,ksr8的突变表型受隐性单基因控制,利用图位克隆技术将突变基因定位于3号染色体的分子标记InD3和RM3280之间,物理距离约106 kb,在该定位区间内包含一个编码催化精氨酸合成通路最后一个步骤的精氨酰琥珀酸裂解酶基因OsASL1(LOC_Os03g19280)。测序结果表明,突变体ksr8中OsASL1基因第3外显子内(CDS 653 bp处)的G突变成A,导致编码的218位精氨酸(R)突变为赖氨酸(K);转基因互补试验证实ksr8的表型是由该基因突变引起;进一步分析发现,外源精氨酸添加可以恢复ksr8的根系缺陷表型。本研究结果证明了突变基因ksr8是OsASL1的新等位基因,进一步明确了OsASL1在水稻根系发育过程中的重要作用,为解析水稻根系发育的分子机制提供了基因资源和理论依据。In the early study, a rice short-root mutant ksr8(Oryza sativa kasalath short root 8) was isolated from an ethylmethane sulfonate(EMS)-mutagenized rice library of indica Kasalath. Phenotype characterization, genetic analysis, gene mapping and cloning, complementation analysis and exogenous application of arginine were conducted to clarify the molecular mechanism of ksr8 root development. Phenotyping analysis showed that the elongation of primary, lateral and adventitious roots and plant height were all inhibited in ksr8 compared with the wild type Kasalath. At the maturation stage, the tiller number, the panicle length and the number of grains per panicle of ksr8 were also reduced. Acetocarmine staining and resin semi-thin sectioning analysis revealed defects in the meristematic cell division activity and cell elongation in the elongation region of the root tip of ksr8. Results of genetic analyses showed that the mutant phenotype was controlled by a recessive nuclear gene, which was mapped to a 106 kb region between markers InDel3 and RM3280 on chromosome 3. One root related gene OsASL1(LOCOs03 g19280) was found in the region, which encodes an argininosuccinate lyase catalyzing the last step of the arginine biosynthesis pathway. Sequencing analysis confirmed a point mutation(G653 to A) in the CDS sequence of OsASL1, resulting in an amino acid substitution(Arg 218 to Lys). Transgenic complementation analysis confirmed that the mutation in OsASL1 was responsible for the phenotype of ksr8. Further analysis showed that the root defects of ksr8 can be restored by exogenous application of arginine. Our results confirmed that the ksr8 gene was a new allele of OsASL1 and further clarified the important role of OsASL1 in root development of rice, providing genetic resources and theoretical basis for further exploration of the molecular mechanism of rice root development.

关 键 词：水稻 短根突变体 OsASL1基因 遗传分析 基因克隆 

分 类 号：S511[农业科学—作物学]