低温胁迫下外源油菜素内酯对冬小麦REVEILLE 7-like基因表达的影响

Effects of exogenous brassinosteroids on REVEILLE 7-like gene expression in winter wheat under low temperature stress

关键词:  寒地冬小麦, 油菜素内酯, REVEILLE 7-like, 基因克隆, 低温胁迫

Key words:  winter wheat in cold zone, brassinolide, REVEILLE 7-like, gene cloning, low temperature stress

摘要:

‘东农冬麦1号(Dn1)'是首个能在严寒地带大面积种植、返青率超过85%的强抗寒冬小麦品种。为探索冬小麦中REVEILLE 7-like基因的功能,以及低温胁迫下外源油菜素内酯(BR)对该基因表达的影响,该研究利用RT-PCR法克隆了冬小麦(Dn1)REVEILLE 7-like基因的cDNA序列,并对其进行了生物信息学分析,在冬小麦三叶期时叶面喷施BR,并分别于4、0、-10、-25 ℃时对冬小麦叶片和分蘖节进行取样,用qRT-PCR技术分析该基因在以上不同处理组冬小麦叶片和分蘖节中的表达模式。结果表明:(1)REVEILLES属于MYB类转录因子家族,REVEILLE 7-like基因全长1 251 bp,共编码416个氨基酸,编码蛋白为不稳定亲水性蛋白,主要定位于细胞核。(2)REVEILLE 7-like与节节麦、二粒小麦亲缘关系最近,蛋白序列总相似度78.62%。(3)REVEILLE 7-like基因启动子包含参与逆境响应、激素响应及MYB结合位点的顺式作用元件。(4)表达模式分析发现,对照组中REVEILLE 7-like基因的表达量在0、-10、-25 ℃时与4 ℃时相比较均有显著提高。处理组与对照组相比,该基因的表达量在-10 ℃和-25 ℃时均显著提高。综上推测,该基因在冬小麦受到低温胁迫时起重要作用,并且BR可以提高该基因的表达,从而进一步提高冬小麦的耐寒性。

Abstract:

‘Dongnong Dongmai No.1'(Dn1)is the first hardy cold-resistant variety that can be planted in a large area in the frigid regions, and the regreening rate is over 85%. To explore the function of REVEILLE 7-like gene in winter wheat and effect of exogenous brassinolide(BR)on the expression of this gene under low temperature stress. In this study, the cDNA sequence of REVEILLE 7-like gene of winter wheat(Dn1)was cloned by RT-PCR, and its bioinformatics analysis was carried out. At the three-leaf stage of winter wheat, BR was sprayed on the leaves, and leaves and tillering nodes of winter wheat were sampled at 4, 0, -10, -25 ℃. The expression patterns of this gene in the leaves and tillers of winter wheat in the above different treatment groups were analyzed by qRT-PCR. The results were as follows:( 1 )REVEILLES belonged to MYB transcription factor family, and the full length of REVEILLE 7-like gene was 1 251 bp, which encoded 416 amino acids. The encoded protein was an unstable hydrophilic protein, mainly located in the nucleus.( 2 )REVEILLE 7-like had the closest genetic relationship with Aegilops tauschii and emmer wheat, showing 78.62% sequence identity.( 3 )REVEILLE 7-like gene promoter contained cis-acting elements involved in adversity response, hormone response and MYB binding site.( 4 )The expression pattern analysis showed that the expression level of REVEILLE 7-like gene in the control group was significantly higher at 0 ℃, -10 ℃ and -25 ℃ than at 4 ℃, suggesting that the gene might play an important role in winter wheat under low temperature stress. At -10 ℃ and -25 ℃, the expression level of the gene both in leaves and tillering nodes of winter wheat treated with BR was significantly higher than that in the control group. In summary, these results suggest that the REVEILLE 7-like gene plays an important role in the response of winter wheat to low temperature stress. Exogenous BR application enhances the expression of this gene, particularly in tillering nodes and at lower temperatures(-10 ℃ and -25 ℃), where its expression level is significantly higher than in leaves. This BR-induced upregulation of REVEILLE 7-like expression further improves the cold tolerance of winter wheat, with a more pronounced effect observed in tillering nodes as temperatures decrease..