铁皮石斛己糖转运蛋白基因DoHT1的分离和表达分析

Isolation and expression analysis of a hexose transporter gene Do HT1 in Dendrobium officinale

该研究借助RACE技术克隆了铁皮石斛己糖转运蛋白（hexose transporter，HT）基因DoHTl，对其序列特征进行生物信息分析，并采用定量PCR分析基因表达模式。结果表明，DoHTl基因（GenBank注册号KUl60469）eDNA全长1586bp，ORF长l536bp，编码l条由511个氨基酸组成的多肽，56．18kDa，等电点9．08。DoHTl蛋白含有糖转运蛋白协助扩散超家族保守结构域（22～483）、糖转运基序1（139-164）、糖转运基序2（338～355）和功能基序；无信号肽，含11个跨膜域，预测定位在质膜，与多种植物HTs蛋白有较高的一致性（54．7％～80．7％），隶属于MST（单糖转运蛋白）分支，与小兰屿蝴蝶兰亲缘关系最近。DoHTt基因具有组织表达特异性，其转录本在石斛叶中显著最高，茎中次为，分别为根中的19．36，1．82倍。该研究获得DoHTl基因全长及分子特征，为进一步研究其在铁皮石斛糖代谢调控中的作用奠定基础。

In this study, RACE technology was employed to isolate the full length cDNA of DoHT1 in Denclrobium oJficinale, fol- lowed by bioinformatics analysis of the sequence characteristics. And the expression pattern of the gene was also analyzed by quantita- tive PCR. The full length cDNA of DoHT1 was 1 586 bp in length, containing a 1 536 bp ORF, which encoded a 511-aa protein with molecular weight of 56. 18 kD and isoelectric point of 9.08. The deduced DoHT1 protein had the major facilitator superfamily conserved domain （22-483）, SUGAR_TRANSPORT_I （ 139-164）, and SUGAR TRANSPORT_2 （338-355）, typical for sugar transporter; DoHTI, without a single peptide had 11 transmembrane regions, and was predicted to locate in the plasma membrane ; DoHT1 had high identities （54. 7%-80. 7% ） with HTs proteins from various plants. DoHT1 belonged to the MST （monosaccharide transporter） group of the evolutionary tree, and was closely related to the Phalaenopsis equestris. DoHT1 was differentially expressed in the three included organs. The transcripts were significantly the most abundant in the leaves with 19.36 fold than roots, then 1.82 fold in the stems than the roots. The identification and molecular characterization of the full length DoHT1 will be essential for further function study of the gene during the regulation of sugar metabolism of D. offtcinale.