本氏烟抗细胞凋亡基因NbDAD1克隆及遗传转化研究

[目的]克隆本氏烟的抗细胞凋亡基因NbDAD1,并对该基因进行遗传转化研究。[方法]通过RT-PCR技术分离本氏烟NbDAD1基因,构建基因超表达载体,以获得转入NbDAD1基因和携带HAtag标签的NbDAD1基因抗性植株。[结果]试验克隆出了全长351 bp的本氏烟NbDAD1基因;成功构建了重组质粒pCAMBIA1301-NbDAD1和pCAMBIA1301-NbDAD1HAtag;共获得3个基因型50株T0代抗Hyg本氏烟植株,其中23株检测PCR呈阳性。[结论]该试验为研究NbDAD1基因在本氏烟中的具体功能及深入探讨DAD1蛋白在植物细胞程序性死亡中可能的作用机制奠定了基础。

Cloning and Gentic Transformation of NbDAD1 Gene from Nicotiana benthamiana

[Objective] The aim was to clone NbDAD1 gene from Nicotiana benthami- ana and study its genetic transformation. [Method] NbDAD1 gene was isolated from N. benthamiana by using RT-PCR technology and over-expression vectors were con- structed to obtain NbDADl-overexpression resistant plants and NbDADl-overexpres- sion resistant plants carrying HA tag. [Result] The 351 bp long NbDAD1 gene was cloned from N. benthamiana; recombinant plasmids pCAMBIA1301-NbDAD1 and pCAMBIA1301-NbDAD1HAtag were constructed successfully; 50T0-generation N. ben- thamiana Hyg-resistant transgenic lines of three genotypes were obtained, including 23 positive transgenic plants. [Conclusion] This study laid the foundation for investi- gating the specific functions of NbDAD1 gene in N. benthamiana and exploring the possible functional mechanism of DAD1 protein in programmed cell death of plants.