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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(Gen Bank注册号KT957550)。DoSWEET1基因cDNA全长1 150 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1 359.06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。
SWEET (sugars will be eventually exported transporters) is a ubiquitous multi-gene family sugar transporter in eukaryotes and plays an important regulatory role in plant physiological metabolism, growth and development and plant-microorganism interaction. In this study, a new SWEET gene was isolated from the rare medicinal Dendrobium officinale Kimura et Migo by RT-PCR and RACE technology and named as DoSWEET1 (GenBank accession number KT957550). The cDNA of DoSWEET1 gene is 1 150 bp in length and encodes a polypeptide of 262 amino acids with a molecular mass of 29.18 kD and an isoelectric point of 9.49. The putative DoSWEET1 protein has no signal peptide with seven transmembrane domains and comprises two conserved MtN3_slv domains (11-94, 130-212) of the plant SWEET family. Multiple sequence alignment showed that DoSWEET1 was highly consistent with SWEET proteins of various plants (45% -54.6%). Phylogenetic tree analysis showed that DoSWEET1 has the closest genetic relationship with rice OsSWEET13, OsSWEET14 and OsSWEET15 and belongs to the ClassⅡ branch of phylogenetic tree of SWEET protein family. qPCR analysis revealed that DoSWEET1 was differentially expressed in three organs of Dendrobium. The transcripts had the highest relative expression levels in roots, followed by leaves at 9.88 and 2.85 folds, respectively. In Tulasnella sp. Dendrobium symbiotic germination of the three seeds, the gene transcription was drastically increased 359.06 times that of the control, indicating that their participation in the germination of the seed germination of bacteria. The cloning and molecular characterization of DoSWEET1 laid the foundation for the further study on the regulation of this gene in the transport of dendrobium saccharide and the symbiotic germination of seed.