中国农业科学 ›› 2021, Vol. 54 ›› Issue (10): 2179-2191.doi: 10.3864/j.issn.0578-1752.2021.10.013

• 园艺 • 上一篇    下一篇

森林草莓FveD27的表达特性和功能

孙洪影1(),王岩1,李伟佳1,2,朱天姝1,姜颖1,许妍1,吴清悦1,张志宏1()   

  1. 1沈阳农业大学园艺学院/辽宁省草莓育种与优质栽培重点实验室,沈阳 110866
    2山西大同大学炭材料研究所,山西大同 037009
  • 收稿日期:2020-08-05 接受日期:2020-11-14 出版日期:2021-05-16 发布日期:2021-05-24
  • 通讯作者: 张志宏
  • 作者简介:孙洪影,E-mail: 2019200077@stu.syau.edu.cn
  • 基金资助:
    国家自然科学基金(31672113);辽宁省“兴辽英才计划”项目(XLYC1902069)

Expression Characteristics and Function of FveD27 in Woodland Strawberry

SUN HongYing1(),WANG Yan1,LI WeiJia1,2,ZHU TianShu1,JIANG Ying1,XU Yan1,WU QingYue1,ZHANG ZhiHong1()   

  1. 1College of Horticulture, Shenyang Agricultural University/Liaoning Key Laboratory of Strawberry Breeding and Cultivation, Shenyang 110866
    2Institute of Carbon Materials Science, Shanxi Datong University, Datong 037009, Shanxi
  • Received:2020-08-05 Accepted:2020-11-14 Online:2021-05-16 Published:2021-05-24
  • Contact: ZhiHong ZHANG

摘要:

【目的】新型植物激素独脚金内酯是调控植物分枝发育的关键因子,但独脚金内酯在草莓生长发育中的作用尚不清楚。揭示森林草莓(Fragaria vesca)中独脚金内酯生物合成关键基因DWARF27D27)的表达特性及主要功能,探索FveD27在草莓分枝以及生长发育过程中的作用,为研究草莓植株构型奠定理论基础。【方法】本研究选用森林草莓作为试验材料,利用RT-PCR方法克隆FveD27的编码区序列,利用MEGA 6.0分析草莓FveD27与苹果、拟南芥等物种中D27的系统进化关系;构建FveD27GFP的融合载体,利用注射烟草叶片的方法对FveD27进行亚细胞定位分析;通过qRT-PCR技术对FveD27在森林草莓不同器官的表达水平进行定量分析,同时构建FveD27启动子与GUS融合表达载体并通过农杆菌介导法将其转化到森林草莓中,利用GUS染色进一步分析FveD27的表达特性;构建FveD27过表达载体并通过农杆菌介导的叶盘法进行稳定遗传转化,获得FveD27过表达草莓株系。【结果】从森林草莓中克隆出编码区长度为789 bp的FveD27序列;烟草的亚细胞定位结果表明FveD27定位在叶绿体;FveD27在森林草莓组织器官中的表达量由高至低依次为幼叶、茎尖、叶柄、成熟叶、根;克隆出长度为1 670 bp的FveD27启动子序列,GUS活性分析结果显示转pFveD27::GUS融合基因草莓植株的幼叶和芽等部位GUS活性较强,而成熟叶与叶柄部位GUS活性较弱,根部几乎无GUS活性,GUS染色分析揭示的基因表达结果与qRT-PCR结果相符;构建了FveD27过表达载体并通过农杆菌介导获得了过表达FveD27的草莓转基因株系,表型调查结果表明过表达FveD27能够显著抑制新茎分枝的形成,同时增加花序数量。【结论】FveD27具有调控森林草莓新茎分枝发育、花序数量等功能。研究结果可为调控草莓新茎分枝数量和产量等提供新思路。

关键词: 森林草莓, 独脚金内酯, D27, 亚细胞定位, 分枝

Abstract:

【Objective】It is known that the new plant hormone strigolactone is a key factor regulating plant branching development, but the role of strigolactone in the growth and development of strawberry plants is unclear. In this study, the expression characteristics and function of the key gene DWARF27 (D27) for strigolactone biosynthesis in woodland strawberry (Fragaria vesca) was revealed, and the role of FveD27 in strawberry branching growth and development was explored, which would lay a theoretical foundation for studying strawberry plant architecture. 【Method】The coding sequence region of strigolactone synthesis key gene FveD27 was cloned by RT-PCR from woodland strawberry. The phylogenetic relationship between FveD27 and D27 in apple, Arabidopsis and other species was analyzed by using MEGA 6.0. A fusion vector of FveD27 and GFP was constructed to inject tobacco leaves for analyzing the subcellular location of FveD27. The expression level of FveD27 in different organs of woodland strawberry was analyzed by qRT-PCR technology. The FveD27 promoter and GUS fusion expression vector was constructed and transformed into woodland strawberry by Agrobacterium-mediated method, and the expression characteristic of FveD27 was further analyzed by GUS staining. The overexpression vector of FveD27 was constructed and the FveD27 overexpression strawberry lines were obtained by Agrobacterium-mediated transformation with leaf disks of woodland strawberry as explants. 【Result】The coding sequence region of FveD27 with the length of 789 bp was cloned from woodland strawberry. The subcellular localization in tobacco showed that FveD27 was located in the chloroplast. The expression levels of FveD27 in woodland strawberry organs from high to low were followed by young leaves, shoot tips, petiole, mature leaves and roots. The FveD27 promoter sequence with a length of 1670 bp was cloned, and transgenic plants with pFveD27::GUS gene showed that the young leaves and buds had strong GUS activity, the mature leaves and petioles had weak GUS activity, while the roots had little GUS activity. The result of FveD27 expression revealed by GUS analysis was consistent with the result by qRT-PCR. The FveD27 overexpression vector was constructed and the transgenic woodland strawberry lines overexpressing FveD27 were obtained with Agrobacterium-mediated transformation. Overexpressing FveD27 in woodland strawberry inhibited significantly the formation of branch crowns and increased the number of inflorescences. 【Conclusion】FveD27 had the functions of regulating the development of branch crowns and the number of inflorescences in woodland strawberry, and this study provided new ideas for regulating the number of branch crowns and yield of strawberry.

Key words: Fragaria vesca, strigolactone, D27, subcellular localization, branching