人表皮生长因子在烟草中的表达及其重组蛋白的初步鉴定

目的:运用Dot-ELISA方法对转基因烟草中所表达的重组hEGF蛋白进行初步检测,以确定是否表达出具有免疫功能的蛋白质.方法:将所合成的hEGF基因置于CAMV35S启动子的驱动下,用农杆菌介导的方法转入烟草中,经过抗性筛选和PCR鉴定出阳性转化子后,提取转化子的总蛋白.总蛋白提取物经透析后,取适当浓度蛋白质点在尼龙膜上,用Dot-ELISA方法对所提取的蛋白质进行测定.结果:结果表明,转基因烟草中的重组蛋白能与抗体发生特异性结合,Dot-ELISA反应后的膜上显示出清晰的棕色斑点.结论:通过实验证明Dot-ELISA方法是一种简便快捷的检测方法,适用于植物基因工程中大规模重组蛋白生产的初步检测.

Expression of human epidermal growth factor in tobacco plants and primary identification of the recombinant protein

Objective: To identify the immunity activity of recombinant hEGF with Dot ELISA method in hEGF gene transgenic tobacco plants.Methods: In this study,the synthetic hEGF gene was controlled under CAMV35S promoter and was transformed into tobacco plants via agrobacterium-mediated procedure.Total protein was extracted from the transforms identified by PCR,and extraction was used to test the immunity activity of recombinant hEGF by Dot-ELISA method.Results: According to the result,the synthetic hEGF gene was expressed in transgenic tobacco plants and the recombinant protein could link to the antibody especially.Conclusion: It is an explanation that the Dot-ELISA method is simple and rapid,this method can be used to identify recombinant protein in plant transgenic engineering.