根癌农杆菌介导大豆转Bt-cryIA抗虫基因

摘要

The transgenic plants were obtained by Agrobacterium - mediated transformation of soybean (DongNong 50) cotyledon nodes. The transformation of cryIA was confirmed by PCR,RT - PCR and Southern blot. The resistance to soybean pod borer of transgenic plants was detected by bioassay method. 114 PPT - resistant plants were obtained and 4, 3and 9 positive plants from T0, T1 and T2 generations respectively were tested by PCR. A 1 800bp target fragment was detected by RT - PCR from 3 T1 transgenic plants and the hybridization bands could be detected by Southern - blot assay in two T1 transgenic plants. The gene of cryIA was successfully integrated into the genome of transgenic plants and could be transcripted normally in T1 generation. Additionally, the results of bioassay indicated that the transgenic plants displayed high insect - resistance to soybean pod borer. Further investigations on the genetic stability of cryIA in different transgenic lines and generations are to be carried out.%通过根癌农杆菌介导法,以子叶节为外植体将抗虫基因cryIA转入东农50大豆品种中,对获得的T0、T1和T2植株采用PCR、RT-PCR及Southern杂交法进行分子检测,并对转基因大豆进行食心虫室内抗性鉴定.经PCR鉴定,T0植株有4株为阳性植株,T1有3株为阳性植株,T2有9株为阳性植株.对3株T1阳性植株进行RT-PCR检测,均扩增得到1 800bp目标片段,Southern杂交分析显示,有2株出现杂交信号,分别为单拷贝和双拷贝.室内抗虫鉴定表明转基因植株食心虫抗性明显优于对照品种,在蛋白质及脂肪酸含量等品质性状上,与对照没有显著差异.证明cryIA基因已整合到受体大豆基因组中,该基因在大豆T1转基因植株中能够正常转录,抗虫基因cryIA的导入可以提高东农50对大豆食心虫的抗性,其后代的遗传稳定性还有待于进一步的研究.