甘蔗金属硫蛋白基因(ScMT2-1-4)的克隆及表达分析

Cloning and Expression Analysis of Metallothionein Gene Sc MT2-1-4 in Sugarcane

从甘蔗热带种Badila(Saccharum officinarum L.)中克隆获得1个2型金属硫蛋白基因的c DNA序列,命名为Sc MT2-1-4(Gen Bank登录号:KJ504375)。生物信息学分析显示,Sc MT2-1-4基因c DNA长459 bp,开放读码框为243 bp,编码80个氨基酸,富含14个半胱氨酸残基。推导的Sc MT2-1-4蛋白为亲水性蛋白,分子量为7.82 k D,等电点为5.59,该蛋白的二级结构由无规则卷曲和延伸链构成。实时荧光定量PCR检测结果显示,Sc MT2-1-4基因是重金属胁迫的快速响应基因,其对Cu2+、Zn2+和Cd2+胁迫的应答模式提示了:甘蔗不同组织中Sc MT2-1-4对Cu2+胁迫应答的分功不同;Sc MT2-1-4对甘蔗抵御Zn2+胁迫起积极的作用;但该基因不直接参与甘蔗对Cd2+的螯合和解毒的过程。研究结果有助于进一步深入探究MT2基因在甘蔗应答重金属胁迫过程中的作用,为阐明甘蔗富集和耐受重金属的分子机制研究奠定基础。

A c DNA sequence of type 2 metallothionein（MT） gene, termed as ScMT2-1-4（Gen Bank Accession number： KJ504375）, was isolated from sugarcane tropical variety of Badila（Saccharum officinarum L.）. The c DNA length of the ScMT2-1-4 is 459 bp containing a 243 bp open reading frame and encoding a polypeptide of 80 amino acids that is rich in cysteines（14）. According to the analysis of bioinformatics, the ScMT2-1-4 is a hydrophilic protein with a size of 7.82 kD and a p I of 5.59. The protein is mainly consisted of random coil and extended strand.Real-time quantitative PCR（q RT-PCR） analysis revealed that ScMT2-1-4 should be a positive response gene under heavy metal stress（Cu^2＋, Zn^2＋and Cd^2＋）. Its expression profile in the Cu^2＋treatment indicated that ScMT2-1-4 in different sugarcane tissues work differently in response to Cu^2＋stress. ScMT2-1-4 exhibited a significant up-regulated expression under the stress of Zn^2＋. However, ScMT2-1-4 is not directly involved in the mechanisms of Cd^2＋accumulation and tolerance in sugarcane. These results formed a basis upon which to further explore the role of MT2 gene in the response process of heavy metal stress and elucidated molecular mechanisms of heavy metal accumulation and tolerance.