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液泡型H+-ATP酶(V-ATPase)是植物生长、发育及应对逆境胁迫的关键酶,全酶由13种不同亚基组成,而V-ATPase c亚基(VHA-c)是分子量为16 k D的高度保守的脂质蛋白,对V-ATPase全酶的组装和活性具有重要作用。本研究中,利用RT-PCR方法从马蔺(Iris lactea)中克隆出Irl VHA-c基因全长读码框,将其构建到原核表达载体pGEX上并在大肠杆菌BL21中诱导表达。结果发现在0.1 mmol/L的IPTG终浓度下诱导8hGST-IrlVHA-c融合蛋白高效表达,并利用层析方法纯化获得了GST-IrlVHA-c融合蛋白,进一步为研究IrlVHA-c蛋白的结构与功能提供帮助。
The vacuolar H + -ATPase (V-ATPase) is a key enzyme in plant growth, development and response to stress. The holoenzyme consists of 13 different subunits, while the V-ATPase c subunit (VHA-c) The highly conserved lipid protein of kD plays an important role in the assembly and activity of the V-ATPase holoenzyme. In this study, the full-length reading frame of Irl VHA-c gene was cloned from Iris lactea by RT-PCR and cloned into prokaryotic expression vector pGEX and induced to express in E. coli BL21. The results showed that the high expression of 8 hGST-IrlVHA-c fusion protein was induced at the final concentration of IPTG of 0.1 mmol / L, and the GST-IrlVHA-c fusion protein was purified by chromatography. The structure and function of IrlVHA-c protein provide help.