木薯糖转运蛋白MeSWEET18的克隆与功能分析  

Cloning and Functional Analysis of Sugar Transporter MeSWEET18 in Cassava

作　　者：薛晶晶[1] 安飞飞[1] 朱文丽[1] 罗秀芹[1] XUE Jingjing;AN Feifei;ZHU Wenli;LUO Xiuqin(Tropical Crops Genetic Resources Institute,Chinese Academy of Tropical Agricultural Sciences/Key Laboratory of Conservation and Utilization for Cassava Germplasm Resources,Haikou,Hainan 571101,China)

机构地区：[1]中国热带农业科学院热带作物品种资源研究所/农业农村部木薯种质资源保护与利用重点实验室,海南海口571101

出　　处：《热带作物学报》2023年第6期1083-1090,共8页Chinese Journal of Tropical Crops

基　　金：海南省基础与应用基础研究计划(自然科学领域)高层次人才项目(No.2019RC314)。

摘　　要：木薯(Manihot esculenta Crantz)是热带亚热带地区重要的粮食作物。而SWEET家族基因在植物运输糖类、生殖和发育、植物逆境、与病原体互作等方面发挥着重要作用。为了明确SWEET家族基因在木薯生长发育过程中的功能,本研究以木薯华南9号(SC9)作为实验材料,克隆糖转运蛋白基因MeSWEET18并进行生物信息学分析,通过酵母实验验证其糖转运能力;采用qRT-PCR分析该基因在木薯不同器官及不同发育时期的表达情况以及在非生物胁迫下的表达趋势。结果表明:MeSWEET18基因开放阅读框为714 bp,编码237个氨基酸,蛋白分子质量为25.94 kDa,理论等电点为6.57,不稳定系数为37.50,属于稳定类蛋白。MeSWEET18蛋白N端含有保守结构域MtN3_slv,C端含有PQ-loop super family保守结构域,且具有7个跨膜结构域,是典型的膜蛋白。ProtScale预测表明MeSWEET18蛋白属于亲水性蛋白。系统进化树分析发现MeSWEET18属于CladeⅣ亚类,与AtSWEET16、AtSWEET17亲缘关系最近;氨基酸序列同源分析显示,MeSWEET18和AtSWEET16的同源性达到53.23%,MeSWEET18和AtSWEET17的同源性达到56.05%。酵母功能互补试验显示MeSWEET18主要转运果糖。qRT-PCR结果表明,随着木薯的生长发育,MeSWEET18在膨大期的块根中表达水平最高,成熟期时急剧下降;而在叶片、叶柄和茎杆中的表达水平随着木薯的发育呈上升趋势,至成熟期达到最大值。黑暗条件下,蔗糖、葡萄糖和果糖溶液对SC9水培苗的处理结果显示,MeSWEET18受果糖影响明显。对SC9水培苗进行高盐(8 g/L NaCl)、干旱(100 mmol/L甘露醇)、氧化(10%H_(2)O_(2))和低温(15℃24 h,后降至4℃24 h)等非生物胁迫,结果发现,MeSWEET18在叶片、叶柄、茎杆和根等不同器官的表达出现不同程度的变化。推测MeSWEET18在木薯的非生物胁迫中起着重要作用。Cassava(Manihot esculenta Crantz)is an important food crop in tropical and subtropical areas.SWEETs par-ticipate in a variety of plant activities,including sugar transport,reproduction and development,plant stress,and inter-action with pathogens,which play an important role in plant development.In order to clarify the function of SWEET in the growth and development of cassava,sugar transporter MeSWEET18 was cloned from Cassava South China 9(SC9)for bioinformatics analysis and verified sugar transport capacity by yeast experiment in this study.The expression trend of MeSWEET18 in cassava in different organ,different developmental stages and under abiotic stress were determined via qRT-PCR method.MeSWEET18 contained a 714 bp open reading frame(ORF)encoding a protein of 237 amino acids residues with predicted molecular mass of 25.94 kDa and theoretical isoelectric point of 6.57.Through instability index(II)analysis by ProtParam,the instability index(II)of the protein encoded by MeSWEET18 was 37.50,indicating the protein of MeSWEET18 as stable.MeSWEET18 was a typical membrane protein with a conserved domain MtN3_slv at the N-terminal and a PQ-Loop Super family conserved domain at the C-terminal and seven transmembrane domains.ProtScale predicted that MeSWEET18 was a hydrophilic protein.Phylogenetic tree analysis revealed that MeSWEET18 belonged to Clade IV.MeSWEET18 was in the same evolutionary tree as AtSWEET16 and AtSWEET17.Alignment of amino acid sequences revealed 53.23%homology between MeSWEET18 and AtSWEET16,and 56.05%homology between MeSWEET18 and AtSWEET17.Yeast functional complementarity tests showed that MeSWEET18 primarily transported fructose.qRT-PCR analysis showed that MeSWEET18 was highly expressed in tuberous roots during the expansion and decreased sharply at maturity of cassava tuberous roots,while the expression levels in leaf,petiole and stem increased with the development of cassava,and reached the maximum at the maturity stage of cassava.qRT-PCR analysis showed that MeSWEET18 was significantly a

关 键 词：木薯 糖转运蛋白 功能分析 糖转运能力 非生物胁迫 

分 类 号：S533[农业科学—作物学]