茶树异胡豆苷合酶基因CsSTR1克隆及表达分析

Cloning and expression analysis of CsSTR1 gene in tea plant

【目的】探明茶树(Camellia sinensis)STR基因(CsSTR)的分子特征及其表达调控模式,为其功能研究以及茶叶药用价值的挖掘奠定基础。【方法】利用RACE克隆技术对茶树CsSTR1基因进行克隆,利用信息学分析方法对其理化性质、蛋白质结构、亚细胞定位以及系统进化关系进行分析,通过qPCR技术检测其表达模式。【结果】CsSTR1基因cDNA全长1382 bp,其中5′UTR长92 bp, 3′UTR长159 bp, CDS长1131 bp,编码376个氨基酸,分子量为41.5 kD,理论等电点为6.36,不稳定指数为29.74,脂肪族氨基酸指数为88.94,亲水性平均系数为-0.077。CsSTR1定位于液泡,其二级结构中无规卷曲、延伸链、α螺旋和β转角占比分别为42.82%、29.79%、16.76%、10.64%,其三维结构呈六叶β-螺旋浆状。系统发育树分析显示,CsSTR1与水稻OsSTRL2蛋白具有较近的亲缘关系。CsSTR1基因在茶树不同组织器官中均有表达,其中在芽和第一、二、三叶中表达量较高,在花和果中表达量较低。低温、PEG模拟干旱胁迫以及MeJA和SA处理均诱导CsSTR1基因上调表达;ABA处理后24~48 h,CsSTR1基因呈现下调表达。【结论】CsSTR1在茶树芽叶发育以及逆境防御反应中具有重要作用,植物激素MeJA,SA和ABA能调控CsSTR1基因表达。

【Objective】The study aimed to explore the molecular characteristics and the expression regulation pattern of STR gene(CsSTR) in tea plant(Camellia sinensis), so as to lay the foundation for further functional analysis of CsSTR gene and for mining the medicinal value of tea. 【Method】By employing the RACE technology, CsSTR1 gene was cloned. The physicochemical properties, protein structure, subcellular localization and phylogenetic relationship of CsSTR1 protein were studied by the methods of bioinformatics. Expression pattern of CsSTR1 gene was analyzed by qPCR. 【Result】The full length of CsSTR1 cDNA was 1382 bp, containing 92 bp 5′ UTR, 159 bp 3′ UTR, 1131 bp CDS. The CsSTR1 protein had 376 aa, with the molecular weight being 41.5 kD, the theoretical isoelectric point being 6.36, the instability index being 29.74, the aliphatic index being 88.94, and the grand average of hydropathicity being-0.077. Subcelluar analysis revealed that the CsSTR1 protein was localized in vacuole. The proportions of random coil, extended strand, α-helix and β-turn in the secondary structure of CsSTR1 protein were 42.82%, 29.79%, 16.76%, and 10.64%, respectively. The three-dimensional structure of CsSTR1 protein displayed as six-bladed β-propeller. The phylogenetic analysis revealed a close genetic relationship between CsSTR1 and rice OsSTRL2 protein. qPCR results showed that CsSTR1 gene was expressed in various organs of tea plant, with higher expression levels in buds, the first leaf, the second leaf and the third leaf, with lower expression levels in flowers and fruits. Up-regulation of CsSTR1 gene was seen under low temperature, PEG simulated drought stress, MeJA and SA treatments;Expression of CsSTR1 gene was down-regulated by ABA after 24-48 hours of the treatment. 【Conclusion】The results suggest that CsSTR1 has an important role in both the buds and leaves development of tea plant and defense response to adversity stress, MeJA, SA and ABA can regulate the CsSTR1 gene expression.