State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China.
Int J Biol Sci. 2014 Nov 7;10(10):1171-80. doi: 10.7150/ijbs.9598. eCollection 2014.
RNA interference (RNAi) has great potential for use in insect pest control. However, some significant challenges must be overcome before RNAi-based pest control can become a reality. One challenge is the proper selection of a good target gene for RNAi. Here, we report that the insect ecdysone receptor (EcR) is a good potential target for RNAi-based pest control in the brown planthopper Nilaparvata lugens, a serious insect pest of rice plants. We demonstrated that the use of a 360 bp fragment (NlEcR-c) that is common between NlEcR-A and NlEcR-B for feeding RNAi experiments significantly decreased the relative mRNA expression levels of NlEcR compared with those in the dsGFP control. Feeding RNAi also resulted in a significant reduction in the number of offspring per pair of N. lugens. Consequently, a transgenic rice line expressing NlEcR dsRNA was constructed by Agrobacterium- mediated transformation. The results of qRT-PCR showed that the total copy number of the target gene in all transgenic rice lines was 2. Northern blot analysis showed that the small RNA of the hairpin dsNlEcR-c was successfully expressed in the transgenic rice lines. After newly hatched nymphs of N. lugens fed on the transgenic rice lines, effective RNAi was observed. The NlEcR expression levels in all lines examined were decreased significantly compared with the control. In all lines, the survival rate of the nymphs was nearly 90%, and the average number of offspring per pair in the treated groups was significantly less than that observed in the control, with a decrease of 44.18-66.27%. These findings support an RNAi-based pest control strategy and are also important for the management of rice insect pests.
RNA 干扰 (RNAi) 在害虫防治中有很大的应用潜力。然而,在 RNAi 为基础的害虫防治成为现实之前,必须克服一些重大挑战。一个挑战是正确选择 RNAi 的良好靶标基因。在这里,我们报告昆虫蜕皮激素受体 (EcR) 是褐飞虱 Nilaparvata lugens 的一个很好的潜在 RNAi 为基础的害虫防治靶标,褐飞虱是水稻的严重害虫。我们证明,使用 NlEcR-A 和 NlEcR-B 之间共有的 360bp 片段 (NlEcR-c) 进行喂食 RNAi 实验,与 dsGFP 对照相比,NlEcR 的相对 mRNA 表达水平显著降低。喂食 RNAi 还导致每对 N. lugens 的后代数量显著减少。因此,通过农杆菌介导的转化构建了表达 NlEcR dsRNA 的转基因水稻品系。qRT-PCR 的结果表明,所有转基因水稻品系的靶基因总拷贝数为 2。Northern blot 分析表明,发夹 dsNlEcR-c 的小 RNA 在转基因水稻品系中成功表达。N. lugens 的刚孵化的若虫在喂食转基因水稻品系后,观察到有效的 RNAi。与对照相比,所有检测品系的 NlEcR 表达水平均显著降低。在所有品系中,若虫的存活率几乎为 90%,处理组的每对后代数量明显少于对照组,减少了 44.18-66.27%。这些发现支持基于 RNAi 的害虫防治策略,对于管理水稻害虫也很重要。