碳离子束辐照拟南芥介导外源基因转移的研究

Study on Transferring Extraneous Gene to Arabidopsis Thaliana by Carbon Ion Beam Irradiation

采用700keV或4.0MeV碳离子束辐照拟南芥种子,通过对样品的显微摄影,发现随着辐照剂量的增加,碳离子束对种子表面的损伤逐渐加剧,特别是在4.0MeV碳离子束辐照下,当剂量达到1×1014ions·cm-2后,种皮局部逐渐被刻蚀殆尽,甚至造成种皮局部破裂。对拟南芥种子进行台盼蓝染色后的显微观测显示,碳离子束辐照可以导致拟南芥种皮细胞着染,在剂量较大的情况下,部分皮下细胞也可着染,表明碳离子束可作用到皮下细胞,为外源基因提供导入的通道。GUS基因导入后的组织化学检测表明:质粒pCAMBIA1301能够进入4.0MeV碳离子束辐照后的拟南芥种子,并在种子和幼芽中获得瞬间表达。

Arabidopsis thaliana seeds of wild type were irradiated by C ion beams with the energy of 700keV or 4.0MeV. Microscopy of the irradiated intact samples showed that the damage of seed surfaces became more intensive with the increasing of radiation doses of C ion beams with different energies. Especially, local seed coats were gradually itched after the implantation doses were higher than 1×1014ions·cm-2 when the seeds were irradiated with C ion beams with the energy of 4.0 MeV, even that local seed coats were broken down. Microscopic investigation of the seeds dyed with vital dye of Trypan blue showed that the seed coat cells can be stained no matter they were irradiated with 700keV or 4.0 MeV C ion beams. Partial cells beneath seed coats could also be tinted if the doses were high enough. The results indicated that C ion beams with the two energies could act on the seed cells beneath the coats and made channels for extraneous genes. Histochemical analysis of GUS reporter gene showed that 4.0 MeV C ions implantation could mediate extraneous gene instantaneous expression in Arabidopsis thaliana seeds and plumelets.