烟草基因瞬时表达体系的建立与优化研究

Abstract

本研究以GUS基因和RIN基因作为报告基因，利用农杆菌(Agrobacterium tumefaciens) GV3101对烟草叶片注射侵染，探究了农杆菌侵染浓度对本氏烟草(Nicotiana benthamiana)叶片瞬时表达效果的影响，建立并优化了烟草中的瞬时表达体系。结果表明，β-葡萄糖苷酶(GUS β-glucuronidase)基因与RIN基因在OD600 = 1.0的侵染浓度时表达效果最佳。农杆菌介导的烟草叶片瞬时表达方法简单高效，结果准确可靠，从种子播种到收获蛋白只需25天左右。该体系的建立与优化为基因表达和蛋白功能性研究等方面提供了一定支持。 To establish and optimize a transient expression system in Nicotiana benthamiana, the method was developed with the β-glucuronidase (GUS) and Ripening Inhibitor (RIN) as marker genes. Using the agrobacterium-mediated transformation method, GV3101 was used for the effects of different bacteria concentration on the efficiency of protein transient expression in Nicotiana benthamiana. Observed by the results, a higher transient expression level of GUS & RIN gene could be obtained as OD600 value of A. tumefaciens for intiltration 1.0. The entire process only took 25 days from sowing seed to protein analysis. Therefore, this method is simple and rapid. It has a potential application in dissecting gene expression and function in Brassica napus.