桃PpRBD2基因的克隆与表达分析

Cloning and Expression Analysis of PpRBD2 Gene in Peach

为了研究桃中RBD家族基因与低温胁迫之间的关系,本试验采用同源克隆方法,从桃中克隆出了一个新的RBD基因,命名为PpRBD2,并对该基因进行了生物信息学、理化性质和表达分析。试验结果表明,该基因全长2 401 bp,有4个外显子,3个内含子,编码区全长897 bp,编码298个氨基酸,蛋白分子质量为33.14 k D,理论等电点(pI)为4.78;PpRBD2具有两个RRM结构域,是典型的RBD家族蛋白。进一步的聚类分析表明该蛋白与樱桃(Prunus avium)、苹果(Malus domestica)和白梨(Pyrus×bretschneideri)中的RBD蛋白聚类在一起。RT-qPCR结果显示,除‘中华寿桃’外,叶片在经过低温处理后,PpRBD2基因在‘珲春’‘、毛桃1-1’和‘21世纪’等三个桃品种中的相对表达量总体呈现出先下降后上升的趋势,但随着低温处理时间的延长,该基因在两个抗寒性桃品种‘珲春’和‘毛桃1-1’中表达量显著高于两个不抗寒性桃品种‘21世纪’与‘中华寿桃’。综上,桃PpRBD2基因与抗寒性密切相关,该试验结果将为今后桃抗寒育种工作提供理论参考。

In order to understand the relationship between RBD family genes and low temperature stress in peach,a new RBD gene was cloned from peaches, named PpRBD2 by homologous cloning method, and the bioinformatics and physicochemical properties of the gene were analyzed. The results showed that the total length of the gene was 2 401 bp, with 4 exons separated by 3 introns, the coding region was 897 bp, 299 amino acids were encoded,the molecular weight of the protein was 33.14 kD, and the theoretical isoelectric point(pI) was 4.78. PpRBD2 is a typical RBD family protein with two RRM domains. A clustering analysis showed that PpRBD2 protein was clustered with RBD protein from Prunus avium, Malus domestica and Pyrus ×bretschneideri. RT-qPCR results showed that, except for ’Zhonghuashoutao’, the relative expression level of PpRBD2 gene in the peach cultivars of ’Huichun’, ’Maotao1-1’ and ’21Shiji’ showed a trend of decreasing at first and then increasing after low temperature treatment. However, with the increase of low temperature treatment time, the expression level of PpRBD2 gene in’Huichun’ and ’Maotao1-1’ was significantly higher than that in ’Zhonghuashoutao’ and ’21Shiji’, which was consistent with their cold resistance grading. The PpRBD2 gene found in this experiment will provide a theoretical reference for peach cold resistance breeding in the future.