摘要： 构建CaMv35S启动子驱动人白细胞介素12基因(hIL-12)的植物表达载体pBI121-hIL-12,通过三亲杂交法将重组载体导入根癌农杆菌菌株EHA105,农杆菌介导法转化马铃薯茎段,经卡那霉素抗性筛选,获得28株抗性植株。通过PCR检测,22株为阳性,从PCR阳性植株中随机选取5株,ELISA法检测茎和叶中hIL-12的表达量,其中2株与对照组相比有显著性差异(p<0.001),hIL-12的表达量分别为(3 714.0±48.8)pg/g和(3 465.0±185.0)pg/g,初步表明外源基因hIL-12已经整合进马铃薯基因组中并得到了表达,为进一步研究重组hIL-12的分离纯化和生物学活性奠定了试验基础。

关键词: 马铃薯, 人白细胞介素12, 遗传转化, 根癌农杆菌

Abstract: hlL-12 DNA was cloned into plant expression vector pB Ⅱ21 under the control of 35 S pronroter,and thenpBⅡ21-h1L-12 was transferred into Agrobacterium tumefaciens strain EHA105 through the assistant plasmid pRK2013 bytriparental cross. The recombinant plasmid was transformed into potato stems induced by Agrobacterium tumefaciensEHA105.28 transformed plants in all transformed plants were selected by kanamycin selective medium. The hIL-12 genewas found from 22 transformed plants by PCR. ELISA assay showed that the protein expression of the stems and leaves of 2 transgenic potatoes in 5 transgenic potatoes selected at random in the 22 transformed plants was obviously different fromthat of control getup (p<0. 001),and the content of hlf,-12 in the tc}o transgenic potatoes was (3 714. 0士8. 8)pg/ g and(3 465. 0士185. 0)pg/ g respectively. It was sure that the foreign gene was integrated into the genome and expressed inpotato,resulting in two transgenic plants. It lays an experimental foundation for further research of extraction and purifica-tion of the hIL-12 and bioactivity detection.

Key words: Rotato, Human interleukin 12, Cenetic transformation, Agrobacterium tumefaciens

中图分类号: 

• Q789