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细胞分裂素响应调节子（RR）是细胞分裂素信号转导途径中的重要组分，行使细胞分裂素信号输出与调控下游基因信号应答的重要功能。为了探究小麦RR基因在植物低温胁迫应答中的作用，以强抗寒性冬小麦品种东农冬麦1号（Dn1）为材料，克隆了小麦A型细胞分裂素响应调节基因（TaRRA6-D），对其进行生物信息学、亚细胞定位及低温下启动子活性分析，并转化拟南芥野生型创建了过表达株系，进行根长表型观察及抗冻实验。结果表明，TaRRA6-D基因（TraesCS5D02G140200）全长1 088 bp，编码序列696 bp，该基因被定位于小麦5D染色体，其编码蛋白为不稳定疏水性蛋白。经系统进化树分析，小麦TaRRA6-D蛋白与粗山羊草和大麦RR蛋白的亲缘关系较近。TaRRA6-D蛋白定位于细胞核。TaRRA6-D启动子区含有响应低温、干旱、光及多种激素的顺式作用元件；TaRRA6-D启动子的活性受低温诱导表达。与野生型相比，无论是否添加6-BA，过表达株系根长均显著增长。与野生型相比，过表达株系幼苗抗冻性显著增强，6-BA处理对幼苗抗冻性提高更明显。

英文摘要:

Cytokinin response regulators(RR), as important components of the cytokinin signaling pathway, perform critical functions in cytokinin signaling output and regulation of downstream gene signaling responses. In order to investigate the role of wheat RR genes in response to low-temperature stress, we cloned the A-type cytokinin responsive regulator gene TaRRA6-D from the wheat cultivar Dongnongdongmai 1(Dn1), performed bioinformatics analysis, subcellular localization analysis, and promoter activity analysis under low-temperature. We also genetically transformed Arabidopsis wild type to create overexpression lines for phenotypic observation of root length and cold resistance evaluation. Bioinformatics analysis showed that the TaRRA6-D gene(TraesCS5D02G140200) was 1 088 bp in total length, with a coding sequence of 696 bp, which was localized to wheat chromosome 5D, and the encoded protein was an unstable hydrophobic protein. The phylogenetic tree analysis showed that the wheat TaRRA6-D protein was most closely related to the RR proteins of Aegilops tauschii and Hordeum vulgare. Transient expression analysis of transformed tobacco showed that TaRRA6-D protein was localized in the nucleus. Analysis of cis-acting elements in the promoter region of the gene showed that the TaRRA6-D promoter region contained cis-acting elements responsive to low-temperature, drought, light, and a variety of hormones. Expression pattern analysis of the transformed Arabidopsis thaliana lines showed that TaRRA6-D promoter activity was induced to express by low-temperature. Phenotypic observations showed that root length of the overexpression lines increased significantly compared with the wild type, with or without the addition of 6-BA. Seedling freezing resistance assay showed that seedlings of overexpression lines were significantly resistant to low-temperature compared with wild type, and 6-BA treatment significantly improved the freezing resistance of seedlings.

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