重组棉铃虫病毒的外源基因向其他生物的转移

Transfer of the AaIT Gene of a Recombinant Helicoverpa armigera Nucleopolyhedrovirus to its Surrounding Organisms

本文报道了基因工程棉铃虫核多角体病毒(HaSNPV-AaIT)的外源蝎子毒素基因(AaIT)是否会向环境中的植物病原微生物或捕食性天敌转移的实验结果。首先,于实验室内将重组病毒HaSNPV-AaIT与棉花黄萎病病菌(VerticilliumdahliaeLleb.)进行了长达90d的混合培养,在混合培养30d,60d和90d后分别提取棉花黄萎病病菌的基因组DNA,用AaIT基因作探针进行点杂交,结果显示无阳性信号。另外,从多次施用过重组病毒的棉花田中采集了120只龟纹瓢虫和七星瓢虫,利用健康蚜虫饲养3-4d,用碱解液处理瓢虫体表后,从处理液中可以检测到病毒DNA;但瓢虫体表经碱解液和Dnase处理后,从瓢虫体内提取的基因组DNA,用PCR和斑点杂交的方法,都没有检测到AaIT的序列存在。本研究的实验结果说明,基因工程病毒的外源基因向其它生物转移的可能性极低。

To test if the foreign gene of a recombinant baculovirus can be transferred to organisms in the same ecological niche, two experiments were performed. First, the fungus Verticillium dahliae Lleb. was cultivated for up to 90 days in laboratory in the presence of BV,ODV and DNA of recombinant Helicoverpa armigera single nucleocapsid nucleopolyhedrovirus which contains an insect-selective toxin （AaIT） （HaSNPV-AaIT）. At 30, 60 and 90 days, the genomic DNA of Verticillium dahliae was isolated and analyzed for the presence of AaIT sequences by dot-blot hybridization. The results show that no positive signal was .detected. Second, ladybeetles （Pro pylaea japonica thunberg） were collected from cotton fields which had been treated several times with a HaSNPV-AaIT formulation. After rearing on healthy aphids （Rhopalosiphurn pseudobrassicae Davis） for 3-4 days, DNA samples were extracted from the surface of ladybeetle bodies treated or untreated with alkaline solution and Dnase. PCR products specific for HaSNPV-AaIT were found in several samples from the untreated ladybeetle bodies, whereas no such products were found from the bodies after the alkaline and Dnase-treatment by both PCR and dot blot hybridization. From the results presented here, the likelihood of the AaIT gene of the recombinant,HaSNPV transferred to surrounding organisms is very low.