马铃薯X病毒载体介导的乙肝病毒表面抗原在烟草中的表达

Expression of Hepatitis B Virus Surface Antigen in Nicotiana benthamiana Using Potato Virus X Vector

目的:利用马铃薯X病毒(PVX)表达载体,在本生烟草中表达乙肝病毒表面抗原(HBsAg),为生产植物疫苗提供一条快速高效的新途径。方法:将HBsAg基因克隆进PVX表达载体,电转化农杆菌,侵染本生烟的叶片、茎和根。结果与结论:采用ELISA检测重组HBsAg的表达水平,SDS-PAGE确认其大小,Western印迹分析表明重组蛋白可与鼠抗HBsAg单克隆抗体发生特异性反应。HBsAg蛋白表达量在幼小叶片中远高于已伸展的叶片,在叶片中的表达量远高于茎根;表达量会随侵染后时间发生一定的变化,但因植株而异;重组蛋白在可溶性蛋白中的含量最高可达796.81 ng/mg。

Objective： Using plant viral expression vector potato virus X（PVX）, the hepatitis B virus surface antigen （HBsAg） was expressed in Nicotiana benthamiana. It would become potential prospective in producing plant vaccine. Methods： The HBsAg gene was cloned into PVX viral expression vector, and electrotransformed into Agrobacterium tumefaciens, then inoculated leaves, stems and roots. Results ＆ Conclusion： The expression level of the recombinant HBsAg protein was determined with ELISA, and its size wasconfirmed by SDS-PAGE and Western blot. The age of leaves significantly affected the expression of HBsAg protein, as that the younger leaves accumulating 267.81 ng/mL which is 2-60 times the concentration in older leaves. The recombinant protein accumulation within a given leaf declined slightly over time but was not significantly different between 11 and 21 days post-inoculation, and the highest expression level was 796.81 ng/mg total soluble protein.