拟南芥种皮色素形成突变体的筛选与表型鉴定  

Screening and Phenotypic Identification of Seed Coat Pigmentation Mutants in Arabidopsis thaliana

作　　者：李娜 王潇楠 LI Na;WANG Xiaonan(Faculty of Basic Medicine,Zhengzhou Shuqing MedicalCollege,Zhenzhou 450000;Kaifeng Institute of Vegetable Science,Kaifeng 475003)

机构地区：[1]郑州澍青医学高等专科学校基础医学部,郑州450000 [2]开封市蔬菜科学研究所,开封475003

出　　处：《植物研究》2023年第1期59-68,共10页Bulletin of Botanical Research

基　　金：国家自然科学基金项目(31871419,31570294)。

摘　　要：类黄酮在植物应答各种环境胁迫和种皮发育调控中起着重要作用。通过甲基磺酸乙酯(EMS)诱变筛选获得1个透明种皮突变体,与野生型拟南芥(Arabidopsis thaliana)(Col-0)相比,突变体成熟的种子颜色为黄色,其表型性状由隐性单基因控制。利用图位克隆和精细定位技术将突变基因定位于5号染色体MAH20的BAC上,是TT4(At5G13930)基因的第1 299位碱基C突变为T,使得第324位氨基酸甘氨酸突变为谷氨酸。TT4(transparent testa 4)编码1个类黄酮合成的结构基因查尔酮合酶(CHS),突变后种皮透明,种子颜色为黄色,突变体命名为tt4-1。利用功能回补突变体恢复褐色种皮表型,进一步证明了TT4在调节种皮颜色发育过程的重要作用。启动子偶联GUS基因组织表达分析显示TT4基因在植株幼苗的根、茎、叶和花中均有表达,生理表型分析结果显示与野生型相比,突变体tt4-1种子萌发早,幼苗主根短、侧根和根毛较多,成苗叶片气孔开度大和失水率高等特性。该研究将为进一步阐述TT4基因功能奠定理论依据。Flavonols play important roles in plant responses to various environmental stresses and in the regulation of seed coat development.In this study,a transparent seed coat mutant was obtained through ethyl methane sulfonate(EMS)mutagenesis screening,compared with wild-type Arabidopsis thaliana(Col-0),the mature seed color of the mutant was yellow,and its phenotypic trait was controlled by a recessive single gene.Using map-based cloning and fine-mapping technology,the mutant gene TT4(At5G13930)was mapped to BAC of MAH20 on chromosome 5,and the 1 299th base C of the TT4 was mutated to T,resulting in the 324th amino acid glycine mutated to glutamic acid.TT4(transparent testa 4)encoded a structural gene for flavonoid synthesis,chalcone synthase(CHS),and after mutation,the seed coat was transparent and the seed color was yellow,then the mutant was named tt4-1.The phenotype of brown seed coat was restored by functional complement mutants,which further demonstrated the important role of TT4 in regulating the development of seed coat color.The tissue expression analysis of TT4 was conducted by promoter coupling GUS assay,and it was found that TT4 gene was expressed in the roots,stems,leaves of plant seedlings and flowers respectively.Physiological phenotypic analysis showed that the mutant tt4-1 seeds germinated earlier,and the seedlings had shorter taproots,as well as more lateral roots and root hairs,larger stomatal opening and higher water loss rate in mature leaves compared with the wild type.This study would lay a theoretical basis for further elaborating the function of TT4 gene.

关 键 词：TT4 筛选 类黄酮 图位克隆 

分 类 号：S664.9[农业科学—果树学]