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水分对于植物的生长代谢非常重要,植物水通道蛋白(Aquaporins)是一类能快速促进植物水分运输的蛋白质。豇豆拥有顽强的耐旱品质,但在豇豆在水通道蛋白方面的研究还是空白。根据水通道蛋白进化的保守性,本研究从豇豆的表达标签序列(ESTs)中筛选并利用c DNA末端快速扩增技术(RACR)成功从两种豇豆根系中克隆到五个细胞质膜水通道蛋白基因。使用Prot Param、MEGA、Antheprot等生物信息学软件分析了他们的序列的理化性质、进化关系和蛋白二级结构等。利用定量PCR技术首先分析了正常水培下五个基因在两种豇豆根中的表达量,并重点关注了4%浓度的聚乙二醇6000处理24 h后两个表达量高的基因在两种豇豆根中的响应。结果表明,五个水通道蛋白基因编码284~287个氨基酸,理论等电点在7.63~9.08之间,主要定位于质膜上,属于跨膜蛋白。五个水通道蛋白均由无规则卷曲α螺旋、延伸链、和β转角组成。正常水培下两种豇豆根中均以Vu PIP1-1和Vu PIP2-3表达量最高,Vu PIP2-2其次,以Vu PIP1-2和Vu PIP2-1表达量最低。PEG处理下,Vu PIP1-1和Vu PIP2-3在抗旱品种根中无显著变化,而在非抗旱品种根中的表达显著下调。本研究认为,豇豆在应对聚乙二醇诱导的渗透胁迫时,Vu PIP1-1和Vu PIP2-3两个水通道蛋白基因有重要的作用。
Moisture is very important for the growth and metabolism of plants. Aquaporins, a plant protein, are a type of protein that can rapidly promote plant water transport. Cowpea has a stubborn drought-tolerant quality, but the research on cowpea in aquaporin is still blank. According to the conservatism of aquaporins evolution, we screened from cowpea expression tag sequences (ESTs) and successfully cloned five cytoplasmic membrane aquaporins from two kinds of cowpea roots using rapid amplification of cDNA ends (RACR) gene. Bioinformatics software such as Prot Param, MEGA and Antheprot was used to analyze the physicochemical properties, evolutionary relationships and protein secondary structure of their sequences. Quantitative PCR was firstly used to analyze the expression of five genes in two kinds of cowpea roots under normal hydroponics. The focus was on the two genes with high expression level after treatment with 4% polyethylene glycol 6000 for 24 h. Response of cowpea roots. The results showed that the five aquaporin genes encoded 284 ~ 287 amino acids, the theoretical isoelectric point was between 7.63 ~ 9.08, mainly located on the plasma membrane, which belongs to the transmembrane protein. The five water channel proteins are composed of random curly alpha helices, extended chains, and beta turns. Vu PIP1-1 and Vu PIP2-3 showed the highest expression levels in two kinds of cowpea roots under normal hydroponics. Vu PIP2-2 followed by Vu PIP1-2 and Vu PIP2-1 had the lowest expression levels. Under PEG treatment, Vu PIP1-1 and Vu PIP2-3 had no significant changes in the roots of drought-tolerant cultivars, but significantly down-regulated in roots of non-drought-tolerant cultivars. In this study, Vu PIP1-1 and Vu PIP2-3 two aquaporin genes play an important role in cowpea’s response to PEG-induced osmotic stress.