小麦TaDHN2基因及其启动子的克隆与功能研究

Cloning and Functional Analysis of Coding and Promoter Regions of TaDHN2 Gene in Wheat

为给深入研究Ta DHN2基因在小麦抗旱机制中的作用机理奠定基础,并为进一步丰富小麦DHN基因研究内容提供参考,本研究通过筛选石麦15基因组BAC文库和BAC克隆测序方法克隆了Ta DHN2基因及其启动子,并对Ta DHN2基因序列特征、表达模式和启动子功能等进行了分析和探讨。结果表明,Ta DHN2基因含有1个88 bp的内含子,开放读码框长为696 bp,编码1个含有231个氨基酸的脱水素蛋白。Ta DHN2蛋白具有Y-segment、S-segment和K-segment结构域,属于YSK2类型脱水素蛋白。此外,该蛋白含有明显的核定位信号序列S-segment和基序RRKK。Ta DHN2基因受渗透胁迫诱导表达,在根和叶中表达模式类似,叶中表达量显著高于根中。Ta DHN2基因启动子序列长为2 025 bp,预测含有9个脱水响应顺式元件。在转基因拟南芥中,Ta DHN2基因启动子能够启动GUS基因表达,并在渗透胁迫下诱导GUS基因上调表达。以上结果说明,Ta DHN2基因为脱水响应基因,其启动子为渗透胁迫强诱导启动子。

In order to study the mechanism of Ta DHN2 gene in wheat drought resistance and enrich the knowledge of DHN genes in common wheat,the coding and promoter sequences of Ta DHN2 gene have been cloned by BAC library screening of Shimai 15 and sequence walking. The sequence character,expression pattern and promoter function of Ta DHN2 gene were analyzed in this study. Only an intron with 88 bp was identified in Ta DHN2 gene. The open reading frame of Ta DHN2 gene is 696 bp long encoding a putative 231-amino-acid dehydrin protein. Ta DNH2 protein contained Y-segment,S-segment and K-segment,belonging to YSK2-type dehydrin protein. Ta DHN2 gene contained putative nuclear localization signals,S-segment and RRKK. The expression of Ta DHN2 gene can be induced by osmotic stress. The expression pattern in root and leaf were similar,but expression level in leaf was higher than that in root. Nine cis-elements responding to dehydration were predicted in the 2 025 bp-long promoter of Ta DHN2 gene.Ta DHN2 promoter could lead to the expression of GUS gene and be induced by osmotic stress in transgenic Arabidopsis. The results indicated Ta DHN2 gene could be a dehydration-response gene and its promoter was strongly induced by osmotic stress.