玉米 ZmGST 基因的克隆及表达特性分析

Cloning and Expression Characteristics of ZmGST Gene in Maize

谷胱甘肽硫转移酶基因(GST)可以参与植物应对非生物胁迫的过程。为了解ZmGST在玉米中响应盐和干旱胁迫的能力,以玉米自交系CA66为材料,通过RT-PCR克隆获得ZmGST基因并对该基因进行了生物信息学分析,通过实时荧光定量分析该基因在玉米不同组织以及在模拟干旱和盐胁迫下的相对表达量,构建原核表达载体后进行干旱和盐处理。结果表明,ZmGST基因CDS序列全长为384 bp,编码127个氨基酸,生物信息学分析结果表明,该基因属于Tau家族,为亲水不稳定蛋白,不存在跨膜结构,存在12个磷酸化修饰位点,未发现有信号肽,为非分泌型蛋白,亚细胞定位预测ZmGST位于细胞核和细胞质。同源序列结果表明,ZmGST与高粱的亲缘关系最近,与ZmGSTU14基因相似度最高。启动子分析结果发现,含有TC-rich repeats等元件,参与防御和应激反应。实时荧光定量结果显示,ZmGST基因在玉米根中的表达量最高,并受盐胁迫诱导上调表达,在处理24 h后达到最高。在原核水平的盐和干旱胁迫结果显示,重组质粒pET28a-ZmGST的大肠杆菌菌体在不同盐浓度的培养基中均能正常生长,模拟干旱被抑制。推测ZmGST基因在玉米响应盐胁迫方面有重要作用。

The glutasidyl thiotransferase gene(GST)can be involved in the process of plants responding to abiotic stresses.In order to understand the ability of ZmGST in response to salt and drought stress in maize,the ZmGST gene was obtained by RT-PCR cloning using the inbred line CA66 of maize as the material,and the gene was bioinformatically analyzed,and the relative expression of the gene in different tissues of maize and simulated drought and salt stress was analyzed by Real-time fluorescence,and the prokaryotic expression vector was constructed and treated with drought and salt.The results showed that the CDS sequence of ZmGST gene was 384 bp in length and encoded 127 amino acids,and the bioinformatics analysis showed that the gene belonged to the Tau family,was a hydrophilic unstable protein,did not have a transmembrane structure,had 12 phosphorylation modification sites,found no signal peptide,was a non-secreted protein,and subcellular localization predicted that ZmGST was located in the nucleus and cytoplasm.The homologous sequence results showed that ZmGST was the closest related to Sorghum bicolor,and had the highest similarity with ZmGSTU14 gene.The promoter analysis results were found to contain TC-rich repeats and other elements,which are involved in defense and stress response.Real-time fluorescence quantitative results showed that the expression of ZmGST gene was the highest in maize roots and was upregulated by salt stress,which reached the highest level after 24 h of treatment.The results of salt and drought stress at the prokaryotic level showed that the recombinant plasmid pET28a-ZmGST could grow normally in medium with different salt concentrations,and the simulated drought was inhibited.It is speculated that ZmGST gene plays an important role in maize response to salt stress.