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MxIrt1是从苹果属植物小金海棠中克隆出的二价阳离子转运膜蛋白基因。为进一步研究该基因的功能,利用MxIrt1基因位于第3和第4跨膜区之间的162bp片段,构建了原核表达载体pGEX-MxIrt1,经原核表达、亲和层析、获得GST-MxIRT1融合蛋白,以融合蛋白为抗原,制备多克隆抗体,ELISA方法检测抗体效价阳性,蛋白质印迹检测植物体内总蛋白,获得与预期大小一致的特异性条带。上述结果表明,表达的目的蛋白可用于免疫组织化学、蛋白质印迹检测。
MxIrt1 is a divalent cation transport membrane protein gene cloned from the genus Malus. To further investigate the function of this gene, a prokaryotic expression vector pGEX-MxIrt1 was constructed by using a 162 bp fragment of MxIrt1 gene located between the third and the fourth transmembrane regions. After prokaryotic expression and affinity chromatography, the GST-MxIRT1 fusion protein The fusion protein was used as antigen to prepare polyclonal antibody. The antibody titer was detected by ELISA and the total protein in plant was detected by Western blotting. The specific band with the expected size was obtained. The above results show that the expressed protein can be used for immunohistochemistry, Western blot detection.