紫花苜蓿MsHB1基因的克隆及表达分析

Cloning and Expression Analysis of MsHB1 Gene from Medicago sativa

HD-Zip(Homologous structural Domain-leucine Zip,同源结构域-亮氨酸拉链)蛋白是植物特有的转录因子,其中HB1属于HD-ZIP I亚族。通过调节植物体内的内源激素含量变化,HB1转录因子可以提高植物对干旱、高盐等不利环境的抵抗能力。本文为探究紫花苜蓿HB1基因的生物学功能,采用RT-PCR技术成功克隆了MsHB1基因。其编码区长867 bp,编码288个氨基酸。氨基酸序列分析结果表明,HB1蛋白为不稳定蛋白。系统发育树分析表明HB1蛋白与蒺藜苜蓿(Medicago truncatula)同源蛋白的亲缘关系接近。构建MsHB1的原核表达载体,在大肠杆菌BL21中成功诱导紫花苜蓿MsHB1蛋白表达,Western Blot也证实MsHB1蛋白表达成功。表达分析显示:MsHB1基因在紫花苜蓿根、茎、叶中均有表达,在不同发育阶段中,幼嫩的叶片中表达最高。MsHB1在NaCl和ABA处理时的基因表达水平升高,推测MsHB1可能在紫花苜蓿高盐等胁迫应答方面发挥着重要的作用。

HD-Zip(Homologous structural Domain-leucine Zip)proteins are plant-specific transcription factors,among which HB1 belongs to the HD-ZIP I subfamily.By regulating variations in the content of endogenous hormones within the plant,the HB1 transcription factor can enhance the plant’s resistance to adverse environments such as drought and high salinity.In this study,to explore the biological function of the HB1 gene in alfalfa(Medicago sativa),the MsHB1 gene was successfully cloned using RT-PCR technology.Its coding region was 867 bp in length,encoding 288 amino acids.Amino acid sequence analysis revealed that the HB1 protein was an unstable protein.Phylogenetic tree analysis indicated a close evolutionary relationship between the HB1 protein and its homologous protein in Medicago truncatula.The prokaryotic expression vector of MsHB1 was constructed,and the expression of MsHB1 protein in Escherichia coli BL21 was successfully induced,which was also confirmed by Western Blot.Expression analysis showed that the MsHB1 gene was expressed in the roots,stems,and leaves of alfalfa,with higher expression levels in young leaves during different leaf development stages.The gene expression levels of MsHB1 increased under NaCl and ABA treatment.Therefore,MsHB1 might play an important role in responding to salinity stress in alfalfa.