Key Laboratory of Integrated Pest Management in Crops, Ministry of Agriculture. Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100081, P. R. China.
Int J Biol Sci. 2013 Apr 23;9(4):370-81. doi: 10.7150/ijbs.5929. Print 2013.
RNA interference (RNAi) caused by exogenous double-stranded RNA (dsRNA) has developed into a powerful technique in functional genomics, and to date it is widely used to down-regulate crucial physiology-related genes to control pest insects. A molt-regulating transcription factor gene, HaHR3, of cotton bollworm (Helicoverpa armigera) was selected as the target gene. Four different fragments covering the coding sequence (CDS) of HaHR3 were cloned into vector L4440 to express dsRNAs in Escherichia coli. The most effective silencing fragment was then cloned into a plant over-expression vector to express a hairpin RNA (hpRNA) in transgenic tobacco (Nicotiana tabacum). When H. armigera larvae were fed the E. coli or transgenic plants, the HaHR3 mRNA and protein levels dramatically decreased, resulting developmental deformity and larval lethality. The results demonstrate that both recombinant bacteria and transgenic plants could induce HaHR3 silence to disrupt H. armigera development, transgenic plant-mediated RNAi is emerging as a powerful approach for controlling insect pests.
外源双链 RNA(dsRNA)引起的 RNA 干扰(RNAi)已发展成为功能基因组学的一种强大技术,迄今为止,它被广泛用于下调关键生理相关基因以控制害虫。选择棉铃虫(Helicoverpa armigera)的蜕皮调节转录因子基因 HaHR3 作为靶基因。克隆了四个包含 HaHR3 编码序列(CDS)的不同片段到载体 L4440 中,在大肠杆菌中表达 dsRNA。然后将最有效的沉默片段克隆到植物过表达载体中,在转基因烟草(Nicotiana tabacum)中表达发夹 RNA(hpRNA)。当棉铃虫幼虫取食大肠杆菌或转基因植物时,HaHR3 mRNA 和蛋白水平显著降低,导致发育畸形和幼虫死亡。结果表明,重组细菌和转基因植物都可以诱导 HaHR3 沉默,从而破坏棉铃虫的发育,转基因植物介导的 RNAi 正成为控制昆虫的一种有力方法。