幽门螺杆菌ureB基因的草莓转化研究

Agrobacterium-mediated transformation for ureB gene from Helicobacter pylori in strawberry(Fragaria ananassa Duch.)

建立了农杆菌介导利用潮霉素抗性作为筛选标记的草莓转化系统。以草莓叶片为材料,把Helicobacter pylori的ureB基因连接到CaMV35S启动子后构成pCAMBIA13011-ureB表达载体,通过根癌农杆菌EHA105介导叶盘法转化草莓,经共培养和潮霉素抗性筛选,得到60株抗性植株。对所得植株进行PCR检测,表明阳性苗比例为42%。RT-PCR结果进一步证明目的基因在T0代转化植株中已经转录。

A system of production of UreB antigen in transgenic strawberry plants was estabhshed. Helicobacter pylori ureB gene was cloned to 5＇ end of GUS reporter gene between CaMV35S promoter and the NOS terminator in the plasmid pCAMBIA13Oll-ureB, and then transformed ureB into strawberry leaf-disc by Agrobacterium- mediated method. The recombinant pCAMB1A13011-ureB plasmid enables selection of transformants on media containing hygromycin and stable integration into nuclear chromosomal plant DNA. After twice selections of hygromycin cultures, a total of 60 regenerated plants with hygromycin resistance were obtained in the selection media, and about 50% of them turned out to be transgenic. PCR confirmed that the ureB gene has been integrated into the chromosomal DNA of these transgenic plants, and RT-PCR confinned the transcription of the genes.