Liu Xun, He Bo-Wen, Zhang Yi-Zheng
College of Life Sciences, Sichuan University, Sichuan Key Laboratory of Molecular Biology and Biotechnology, Chengdu 610064, China.
Yi Chuan. 2009 Aug;31(8):859-64. doi: 10.3724/sp.j.1005.2009.00859.
A prokaryotic expression plasmid pET-IbGST, which contains the full encoding region of a glutathione-S- transferase (GST) gene of sweet potato, was constructed. The recombinant IBGSTU1 protein was expressed in Escherichia coli and found in the soluble fraction, as well as in insoluble inclusion bodies of lysed cells. Its enzymatic activity was detected using UV spectrophotometer. The protein was purified and used to prepare antibody. Semi-quantitative RT-PCR and Western blotting analyses demonstrated that IBGSTU1 gene was not expressed under normal conditions. When subjected to some environ-mental stresses such as cold-stress or heavy-medal stress, the organism switches on the expression of IBGSTU1 at both mRNA and protein levels, and its expression has tissue specificity.
构建了一个原核表达质粒pET-IbGST，其包含甘薯谷胱甘肽-S-转移酶（GST）基因的完整编码区。重组IBGSTU1蛋白在大肠杆菌中表达，并且在裂解细胞的可溶部分以及不溶性包涵体中均被发现。使用紫外分光光度计检测其酶活性。该蛋白被纯化并用于制备抗体。半定量RT-PCR和蛋白质印迹分析表明，IBGSTU1基因在正常条件下不表达。当受到诸如冷胁迫或重金属胁迫等一些环境胁迫时，生物体在mRNA和蛋白质水平上均开启IBGSTU1的表达，并且其表达具有组织特异性。
