顺乌头酸酶AtACO3参与调控Zn稳态的研究

Involvement of AtACO3in Zn homeostasis

从拟南芥中克隆得到AtACO3基因的全长cDNA序列.半定量PCR实验结果表明,200μmol/L CuCl2处理能明显抑制该基因的表达,而200μmol/L ZnCl2处理1 h则可显著促进其表达.为验证AtACO3蛋白N端的功能,构建AtACO3 5'端1632 bp基因片段的原核表达载体pET30a-AtACO3-N,并转化大肠杆菌.SDS-聚丙烯酰胺凝胶电泳检测结果显示,该基因片段可以在大肠杆菌中稳定表达.进一步的抗性实验表明,异源表达基因AtACO3的N端序列能提高大肠杆菌的Zn2+耐受性.

A full-length cDNA, designated as AtAUO3, was isolated from AraOtctopsts tttattana. Semi-quantitative RT-PCR revealed that AtACO3 was significantly activated under treatment with 200 p, mol/L ZnCl2 for 1 hour while inhibited under treatment with 200 μmol/L CuCl2. To investigate the function of the N-terminus of AtACO3, the prokaryotic expression vector pET30a with a fragment of AtAC03 including 1632 bp from the initiation codon was constructed and successfully transformed E. colt. SDS-PAGE indicated that the recombinant N-terminus of AtAC03 was soluble and stable when expressed in E. colt. Additionally, the tolerance of E. colt against Zn^2＋ was found to be enhanced by expressing the N-terminus fragment of AtACO3.