木薯金属硫蛋白基因的克隆和表达分析

Cloning and Expression Analysis of Cassava Metallothionein Gene

金属硫蛋白(metallothionein,MT)分子量小,半胱氨酸含量高,具有消除离子自由基,减弱重金属毒性和减少体内活性氧积累等功能。为探究木薯金属硫蛋白基因(MeMT)的原核表达能力和对活性氧的耐受性,本研究通过木薯cDNA克隆获取MeMT并将其连接至pET-28a,在BL21(DE3)菌株中成功诱导其表达。H2O2耐受性分析发现,与空载菌株相比,表达MeMT蛋白的DE3菌株对H2O2的耐受水平较高。RT-qPCR技术检测显示,在H2O2胁迫下,3 h时木薯叶片中MeMT的表达水平明显升高,但6 h时恢复甚至略低于对照组,证明MeMT在木薯中参与ROS应答过程。该结果对研究木薯抗逆性和产量的提高具有重要意义。

Metallothionein,a low-molecular-weight protein with rich cysteine,involves in scavenging ion radicals,reducing the toxicity of heavy metals and the accumulation of reactive oxygen species.This study aimed to investigate the prokaryotic expression level of metallothionein from cassava and its tolerance to reactive oxygen species.MeMT was cloned and linked in pET-28a to obtain fusion expression vector pET-28a-MeMT,then the pET-28a-MeMT vector was transferred to BL21(DE3)strain.Analysis of H2O2 tolerance showed that MeMT can increased tolerance to H2O2.Furthermore,RT-qPCR analysis showed that under H2O2 stress,the expression level of MeMT was significantly increased at 3 h,but recovered at 6 h.These results indicate that the MeMT gene is involved in the ROS response in cassava.These results is of great significance for increasing the resistance and the yield of cassava.