苹果糖转运蛋白基因MdSWEET17的功能鉴定

杨官显1,2, 许海峰1,2, 张静1,2, 王楠1,2, 房鸿成1,2, 邹琦1,2, 王意程1,2, 姜生辉1,2, 陈学森1,2,*
1山东农业大学园艺科学与工程学院, 山东泰安271018; 2山东农业大学作物生物学国家重点实验室, 山东泰安271018

通信作者:陈学森;E-mail: xhfdl1991@163.com

摘 要:

苹果糖转运蛋白基因MdSWEET17在液泡膜糖转运过程中发挥着重要的作用。本文以新疆红肉苹果杂种一代优系‘红脆1号’为试材, 克隆MdSWEET17并原核诱导获得其重组蛋白, 对其进行生物信息学分析; 测定该基因在不同组织及不同发育时期的表达, 通过亚细胞定位及转基因鉴定其功能。结果显示, MdSWEET17基因编码区长度为786 bp, 蛋白大小为29.34 kDa左右, 定位于15号染色体, 由5个外显子和4个内含子组成。糖代谢相关转运蛋白系统进化树分析发现MdSWEET17与AtSWEET17在同一个进化树枝上; MdSWEET17定位于苹果‘王林’愈伤组织原生质体的液泡膜上。定量表达分析表明, MdSWEET17在苹果的根、叶和幼果中均有较高的表达, 且在果实发育中, MdSWEET17的表达水平与果糖含量显著负相关。MdSWEET17启动子含有与糖信号及激素信号相关的顺式作用元件。低温和糖处理‘王林’愈伤组织后发现MdSWEET17响应果糖和低温诱导; 而在‘王林’愈伤组织中过表达MdSWEET17, 发现它对葡萄糖和蔗糖含量没有影响, 但能减少‘王林’愈伤组织的果糖含量。

关键词:苹果; 糖转运蛋白; SWEET17; 功能鉴定; 亚细胞定位

收稿:2018-02-05   修定:2018-11-01

资助:国家自然科学基金(31572091)和国家公益性行业(农业)科研专项(201303093)。

Functional identification of a sugar transporter gene MdSWEET17 in apple

YANG Guan-Xian1,2, XU Hai-Feng1,2, ZHANG Jing1,2, WANG Nan1,2, FANG Hong-Cheng1,2, ZOU Qi1,2, WANG Yi-Cheng1,2, JIANG Sheng-Hui1,2, CHEN Xue-Sen1,2,*
1College of Horticulture Science and Engineering, Shandong Agricultural University, Tai’an, Shandong 271018, China; 2State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai’an, Shandong 271018, China

Corresponding author: CHEN Xue-Sen; E-mail: xhfdl1991@163.com

Abstract:

The apple sugar transporter gene MdSWEET17 played an important role in tonoplast sugar transport. We cloned the MdSWEET17 in apple ‘Hongcui No.1’ and obtained the recombinant protein by the prokaryotic induction technology, then, analysed the bio-information of it. The expression levels of MdWEET17 in different tissues and different development stages were studied by the qRT-PCR, and we also identificated the fuction of MdWEET17 by the subcellular localization and transgenosis. The results showed that the length of coding sequence was 786 bp and the molecular mass of MdSWEET17 was about 29.34 kDa, it was located in the chromosome 15 of the apple genome, included 5 exons and 4 introns. A phylogenetic tree of related transporter in sugar metabolism indicated MdSWEET17 and AtSWEET17 were located in the same evolutionary branch. Md-SWEET17 was located in the tonoplast of orin callus. The qRT-PCR results showed that the MdSWEET17 had the higher expression level in roots, leaves and young fruits of apples, and its expression had a significant negative correlation with the content of fructose during the development stage of fruits. The promoter of MdSWEET17 contained several typical cis-acting elements, including sugar signaling responsive elements and phytohormone responsive elements. After low temperature and sugar treatment of apple ‘Orin’ callus MdSWEET17 was found in response to fructose and low temperature induction. When overexpressing MdSWEET17 in the orin callus, We found it could reduce the contents of fructose, however, it had no influence on the contents of sucrose and glucose.

Key words: apple; sugar transporter; SWEET17; functional identification; subcellular localization

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