A fluorescence-based high-throughput screening method for cytokinin translocation mutants

Plant Methods

Guodong Yang¹  Bingli Ding²  Mengyuan Zhang²  Yujia Li²  Kewei Zhang²  Jiangzhe Zhao²  Penghong Zhang² 

[1] College of Life Sciences, Shandong Agricultural University, 271018, Taian, Shandong, China;Institute of Plant Genetics and Developmental Biology, College of Chemistry and Life Sciences, Zhejiang Normal University, 321004, Jinhua, Zhejiang, China;Zhejiang Provincial Key Laboratory of Biotechnology on Specialty Economic Plants, Zhejiang Normal University, 321004, Jinhua, Zhejiang, China;

关键词: Cytokinin;    Translocation;    Arabidopsis;    EMS mutagenesis;    Fluorescence reporter;   

DOI  :  10.1186/s13007-020-00676-4

来源: Springer

【 摘 要 】

BackgroundCytokinins are one kind of phytohormones essential for plant growth, development and stress responses. In the past half century, significant progresses have been made in the studies of cytokinin signal transduction and metobolic pathways, but the mechanism of cytokinin translocation is poorly understood. Arabidopsis (Arabidopsis thaliana) response regulator 5 (ARR5) is a type-A response factor in cytokinin signaling which is induced by cytokinins and has been used as a reporter gene for the endogenous cytokinins in Arabidopsis. Here, we report a fluorescence-based high-throughput method to screen cytokinin translocation mutants using an ethyl methyl sulfone (EMS) mutagenesis library generated with ARR5::eGFP transgenic plants.ResultsThe seedlings with enhanced green fluorescent protein (GFP) signal in roots were screened in a luminescence imaging system (LIS) in large scale to obtain mutants with over-accumulated cytokinins in roots. The selected mutants were confirmed under a fluorescence microscopy and then performed phenotypic analysis. In this way, we obtained twelve mutants with elevated GFP signal in the roots and further found three of them displayed reduced GFP signal in the aerial tissues. Two of the mutants were characterized and proved to be the atabcg14 allelic mutants which are defective in the long-distance translocation of root-synthesized cytokinins.ConclusionsWe provide a strategy for screening mutants defective in cytokinin translocation, distribution or signaling. The strategy can be adapted to establish a system for screening mutants defective in other hormone transporters or signaling components using a fluorescence reporter.

【 授权许可】

CC BY