ie-1和lef-1基因dsRNA表达元件转染及转化细胞对家蚕核型多角体病毒增殖的抑制

Inhibitory Effects of Cells Transformed and Transfected With ie-1 and lef-1 dsRNA Expression Elements on Proliferation of Bombyx mori Nucleopolyhedrovirus

通过RNAi介导可以抑制病毒增殖,将相应的RNAi元件通过转基因转入宿主有可能使宿主对病毒产生一定的抗性。根据家蚕核型多角体病毒(BmNPV)基因组中的病毒复制必需基因ie-1、lef-1设计相应的RNA干扰区段,并构建相应的转基因载体转入家蚕BmN细胞中,瞬时表达结果显示转染了转基因RNAi载体的BmN细胞均对BmNPV有一定的抑制作用。IE dsRNA在病毒滴度为10-4时有抑制作用,在病毒滴度为10-5时有比较好的作用;LEF dsRNA则在病毒滴度为10-5时有抑制作用,在病毒滴度为10-6时有较好的抑制作用。通过转基因及G418筛选后,转基因IE dsRNA细胞在病毒滴度为10-4显示出一定的抑制作用,在病毒滴度为10-5表现了明显的抑制作用;而转基因LEF dsRNA细胞在病毒滴度为10-5有一定的抑制作用,但只维持了一个时段。

Viral proliferation can be inhibited by RNA interference technology, and transgenic organisms with corresponding RNAi element may elicit viral resistance. We selected the ie-1 and lef-1 of BmNPV genome as the RNAi target region in this study. The transgenic vectors harbored the fragments producing the double strands RNA were constructed and transfected into BmN cells. Transient expression of transgenic vector in the cell resulted in its resistance to BmNPV. IE dsRNA had resistance at 10^-4 viral titre, and stronger resistance at 10^-5 viral titre. Meanwhile, LEF dsRNA vector had resistance at 10^-5 viral titre, and stronger resistance at 10^-6 viral titre. After screened by G418, transgenic cell line with IE dsRNA demonstrated the resistance at 10 ^-4 viral titre, and stronger resistance at 10^-5 viral titre. Transgenic cell line with LEF dsRNA showed different resistance at 10^-5 viral titre. But the difference of resistant ability only kept a short time.