甜菜磷脂酰肌醇转运蛋白基因SbSEC14的克隆及低温胁迫下的表达分析

磷脂酰肌醇转运蛋白(PITPs)广泛存在于真核生物细胞中,能够在体外膜脂质双层之间调控磷脂酰肌醇(PtdIns)或者磷脂酰胆碱(PtdCho)单体的独立转运。参与磷酸肌醇代谢、膜运输、极性生长、信号转导、逆境胁迫、胞浆运动和细胞周期调节等多种重要的生命过程,在植物的逆境响应以及发育调节中具有重要的作用。为了研究甜菜磷脂酰肌醇转运蛋白基因及其在低温胁迫下的表达情况。本研究以甜菜基因组数据库中一条预测的磷脂酰肌醇转运蛋白基因CRS1为模板,用基因克隆的方法得到一条全长765 bp,开放阅读框596 bp,编码198个氨基酸的甜菜SEC14基因,命名为SbSEC14。理化性质分析表明,该蛋白质为不稳定亲水蛋白;蛋白质二、三级结构分析表明,该蛋白质α-螺旋所占的比例最高,为47.47%,β-转角所占比例最低,为5.56%;蛋白质保守结构分析表明,该蛋白质有典型的SEC14结构域;蛋白质系统进化树分析表明,甜菜SbSEC14基因与菠菜、藜麦的磷脂酰肌醇运转蛋白基因亲缘关系最近;实时荧光定量结果表明,SbSEC14基因在甜菜中组成型表达,在甜菜叶中的表达量最高,在甜菜根中的表达量最低,在叶中表达量约为根中的2.5倍。甜菜幼苗4℃处理0、2、6、12、24 h,该基因在植株处理0~2 h时表达量呈上升趋势,在植株处理2~6 h时表达量呈下降趋势,在植株处理6~24 h时表达量趋于平稳状态。因此,预测该基因与甜菜抗低温胁迫相关。

Phosphatidylinositol Transporters Gene SbSEC14 C in Sugarbeet: Cloning and Expression Analysis Under Low Temperature Stress

Phosphatidylinositol transporters (PITPs) are widely present in eukaryotic cells and can regulate the independent transport of phosphatidylinositol (PtdIns) or phosphatidylcholine (PtdCho) monomers between lipid bilayers in vitro. They are involved in many important life processes such as phosphoinositide metabolism, membrane transport, polar growth, signal transduction, stress, cytoplasmic movement, and cell cycle regulation. They play an important role in plant stress response and development regulation. In order to study the sugarbeet phosphatidylinositol transporter gene and its expression under low temperature stress, in this study, using a predicted phosphatidylinositol transporter gene CRS1 from sugarbeet genome database as a template, a beet SEC14 gene with a length of 765 bp, an open reading frame of 596 bp, and a coding of 198 amino acids was obtained by gene cloning. It was named SbSEC14. The analysis of the physicochemical properties showed that the protein was an unstable hydrophilic protein. The secondary and tertiary structure analysis of the protein showed that the protein's α-helix accounted for the highest proportion, 47.47%, and the β-turn angle accounted for the lowest, 5.56%. The conserved structural analysis of the protein showed that the protein had a typical SEC14 domain. The phylogenetic tree analysis of the protein showed that the beet SbSEC14 gene was most closely related to spinach and quinoa's. Real-time fluorescence quantitative results showed that the constitutive expression of SbSEC14 gene appeared in sugarbeet, the highest expression was in sugarbeet leaves, the lowest expression was in sugarbeet roots, and the expression in leaves was about 2.5 times that of roots. Sugarbeet seedlings were treated at 4℃ for 0, 2, 6, 12, and 24 h, the expression level of this gene showed an upward trend when the plants were treated for 0-2 h, the expression level was decreased when the plants were treated for 2-6 h and the expression level tended to be steady for 6-24 h. Therefore, it is predicted that this gene is related to the resistance of sugarbeet to low temperature stress.