梁晓 陈青 伍春玲 方永军

摘  要：转录因子Nrf2对脊椎动物抗氧化酶的调控起关键作用，然而当前Nrf2对害虫（螨）抗氧化酶调控功能的相关研究报道较少，本研究拟克隆六点始叶螨转录因子EsNrf2，并分析采用RNAi沉默EsNrf2对六点始叶螨取食感螨橡胶树种质‘IAN2904和抗螨橡胶树种质‘IRCI12后抗氧化酶EsSOD、EsCAT基因表达水平和超氧化物歧化酶（SOD）、过氧化氢酶（CAT）活性的影响。结果表明，采用RACE-PCR从六点始叶螨体内克隆得到大小为1719 bp的序列，并经同源分析和功能域注释确证其为六点始叶螨转录因子EsNrf2。饲喂浓度为50 ng/?L的dsEsNrf2 48 h后对EsNrf2基因的沉默效率达到90.6%，进一步研究发现，与EsNrf2沉默前相比，EsNrf2沉默后六点始叶螨取食抗、感螨橡胶树种质后体内EsSOD和EsCAT的基因表达量、SOD和CAT酶的活性均显著降低（P<0.05）。上述结果从分子水平证明EsNrf2能够调控六点始叶螨抗氧化酶SOD和CAT的表达。

关键词：RNAi;六点始叶螨;转录因子Nrf2;抗氧化酶;调控功能

Abstract： The transcription factor Nrf2 （Nuclear factor erythroid 2-related factor） plays a key role in regulating antioxidant enzymes in vertebrates. However， there are few reports regarding the Nrf2 function of insects （mites）. This study aimed to clone the Nrf2 gene of Eotetranychus sexmaculatus （EsNrf2）， furthermore， after EsNrf2 was silenced via RNA interference （RNAi）， the transcription of EsSOD and EsCAT and enzyme activity of SOD and CAT was analyzed while E. sexmaculatus feeding on mite-susceptible rubber tree germplasm ‘IAN2904 and mite-resistant rubber tree germplasm ‘IRCI12， respectively. The results showed a sequence 1719 bp long was cloned using RACE-PCR， and then it was identified as transcription factor EsNrf2 by homology analysis and conserved domain. In addition， it was found that EsNrf2 silencing efficiency could reach 90.6% after feeding 50 ng/?L of dsEsNrf2 for 48 hours. Further analysis showed that， compared with those before silencing， the transcription of EsSOD and EsCAT as well as the activity of SOD and CAT were significantly reduced after RNAi of EsNrf2 （P < 0.05）. The above results showed that EsNrf2 could regulate the expression of SOD and CAT in E. sexmaculatus.

Keywords： RNAi; Eotetranychus sexmaculatus; transcription factor Nrf2; antioxidant enzymes; regulation function

天然橡膠是我国重要的战略物资，在国防和国民经济建设中具有不可替代的作用。我国是世界上第一大天然橡胶消费国和进口国，对外依存度已超过80%，天然橡胶安全问题的严峻性十分突出[1-2]。六点始叶螨（Eotetranychus sexmaculatus）是危害我国橡胶树最严重的一种世界危险性害螨，2008年以来六点始叶螨在海南、云南多地的橡胶树上暴发成灾，导致大量叶子变黄脱落、开割树停割，胶乳产量急剧下降[3]。当前，各橡胶产区对于该螨的防治仍依赖于化学药剂防治，但橡胶树高大，药剂难以靶标，防治难度很大，特别是药剂防治过程中因施药技术落后所致农药的有效利用率不足、使用频率与剂量不断加大和“3R”等问题严重[4]。……