目的 获得转金属硫蛋白2(metallothionein 2,MT2)红花植株,为MT药用蛋白的生产奠定基础。方法 采用Nco I/Hind III酶切pEASY-oleosin-MT获得oleosin-MT融合基因,将其插入到植物油体高效表达载体pOP上,构建重组质粒pOP-oleosin-MT,进行PCR和双酶切鉴定。采用冻融法将重组质粒pOP-oleosin-MT转入根瘤农杆菌EHA105中,通过农杆菌介导法转化红花,对转MT基因红花植株进行PCR检测及Southern blotting检测。结果 成功构建了MT基因的植物油体表达载体pOP-oleosin-MT,获得了3株PCR检测呈阳性的转MT基因红花植株。结论 建立了完善的红花再生体系,获得了转MT基因的红花植株。

Objective To obtain the transgenic safflower plants which expressed Arabidopsis thaliana metallothionein 2 (MT2) gene, and lay a foundation for development of MT products. Methods The oleosin-MT gene was obtained from pEASY-oleosin-MT by Nco I/Hind III, then was inserted into plant expression vector pOP. The recombinant plasmid named pOP-oleosin-MT was transferred into Agrobacterium tumefaciens EHA105. The oleosin-MT gene was introduced into safflowers via Agrobacterium-mediated method and positive transgenic plants were determined by PCR analysis. Results The recombinant plasmid pOP-oleosin-MT was successfully constructed. PCR and Southern blotting analysis confirmed that MT gene was integrated into the genome of safflower plant and three transgenic plants were obtained. Conclusion The safflower regeneration system is constructed successfully and MT gene is successfully transformed into safflower plant.

国家高技术研究发展计划(863)项目(2011AA100606);国家自然科学基金资助项目(31201237);吉林省教育厅“十三五”科学技术研究项目重点项目(吉教科合字[2016]179号);吉林农业大学大学生创新创业项目(201610193046,201410193045);吉林省科技厅中青年领军人才及优秀创新团队项目(20111815);教育部博士点基金-青年教师基金项目(20122223120002)