药学学报, 2016, 51(6): 991-997
引用本文:
张岗, 刘思思, 杨新杰, 陈莹, 刘亮亮, 郭顺星. 一个全新的铁皮石斛DoSWEET1基因的分子克隆与特性分析[J]. 药学学报, 2016, 51(6): 991-997.
ZHANG Gang, LIU Si-si, YANG Xin-jie, CHEN Ying, LIU Liang-liang, GUO Shun-xing. Molecular cloning and characterization of a novel DoSWEET1 gene from Dendrobium officinale[J]. Acta Pharmaceutica Sinica, 2016, 51(6): 991-997.

一个全新的铁皮石斛DoSWEET1基因的分子克隆与特性分析
张岗1,2, 刘思思1, 杨新杰2, 陈莹2, 刘亮亮2, 郭顺星1
1. 中国医学科学院、北京协和医学院药用植物研究所, 北京 100193;
2. 陕西中医药大学药学院, 陕西 西安 712046
摘要:
SWEET (sugars will be eventually exported transporters)是真核生物中普遍存在的一类多基因家族糖转运蛋白,在植物生理代谢、生长发育及植物-微生物互作过程中起重要调控作用。本研究利用RT-PCR、RACE技术,从珍稀药用铁皮石斛(Dendrobium officinale Kimura et Migo)中分离到一个全新的SWEET基因,命名为DoSWEET1(GenBank注册号KT957550)。DoSWEET1基因cDNA全长1150 bp,编码1条262个氨基酸组成的多肽,分子质量29.18 kD,等电点9.49。推定的DoSWEET1蛋白无信号肽,具有7个跨膜域,包含植物SWEET家族2个保守MtN3_slv结构域(11-94、130-212)。多序列比对表明, DoSWEET1与多种植物SWEET蛋白一致性较高(45%~54.6%)。进化树结果显示, DoSWEET1与水稻OsSWEET13、OsSWEET14和OsSWEET15亲缘关系最近,隶属于SWEET蛋白家族分子进化树的ClassⅡ分支。qPCR分析揭示, DoSWEET1在石斛3个器官中差异表达,该转录本在根中相对表达量最高,叶中次之,分别为茎中的9.88倍和2.85倍;在胶膜菌Tulasnella sp.侵染石斛共生萌发的3级种子中,该基因转录本剧烈上调为对照的1359.06倍,说明其参与种子接菌共生萌发过程。DoSWEET1克隆和分子特征为进一步研究该基因在铁皮石斛糖转运及种子共生萌发过程中的调控作用奠定基础。
关键词:    铁皮石斛      SWEET      表达      定量PCR      共生     
Molecular cloning and characterization of a novel DoSWEET1 gene from Dendrobium officinale
ZHANG Gang1,2, LIU Si-si1, YANG Xin-jie2, CHEN Ying2, LIU Liang-liang2, GUO Shun-xing1
1. Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China;
2. College of Pharmacy, Shaanxi University of Chinese Medicine, Xi'an 712046, China
Abstract:
SWEET (sugars will be eventually exported transporters) constitute a large and conserved gene family of sugar transporters in eukaryotes, which are important in the cellular metabolisms, growth and development, and plant-microbe interaction in plants. In the present study, a full length cDNA of SWEET encoding gene, designed as DoSWEET1(GenBank accession No. KT957550), was identified in Dendrobium officinale using RT-PCR and RACE approaches. DoSWEET1 was 1150 bp in length and encoded a 262-aa protein with a molecular weight of 29.18 kD and an isoelectric point of 9.49. The deduced DoSWEET1 protein contained seven transmembrane regions and two conserved MtN3-slv domains (11-94, 130-212). Multiple sequence alignment revealed that DoSWEET1 had high identities (45%-54.6%) with SWEET proteins from various plants. A neighbor joining phylogenetic analysis suggests that DoSWEET1 belonged to the class II subgroup of the SWEET evolutionary tree, and was closely related to rice OsSWEET13, OsSWEET14, and OsSWEET15. qPCR analysis demonstrated that DoSWEET1 gene was differentially expressed in the three included organs of D. officinale, and the expression was most abundant in the roots at 9.88 fold over that of the stems, followed by that of the leaves with 2.85 fold higher. In the 3rd symbiotic germinating seeds infected by Tulasnella sp., the transcipts were dramatically induced by 1359.06 fold over that in the ungerniamted control seeds, suggesting a vital role of the gene in the D. officinale symbiotic germination process. Molecular cloning and characterization of the novel DoSWEET1 gene provides a foundation for the functional study of the gene in sugar translocation during the D. officinale symbiotic germination process.
Key words:    Dendrobium officinale    SWEET    expression    quantitative PCR    symbiosis   
收稿日期: 2015-11-30
DOI: 10.16438/j.0513-4870.2015-1092
基金项目: 国家自然科学基金资助项目(81473331,81503195);陕西省青年科技新星项目(2012KJXX-44).
通讯作者: 郭顺星,Tel/Fax:86-10-62829619,E-mail:sxguo1986@163.com
Email: sxguo1986@163.com
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