中文摘要：

干扰（RNA interference,RNAi）是指在进化过程中高度保守的、由双链RNA诱发的、同源mRNA高效特异性降解的现象。本研究以稻纵卷叶螟可溶型海藻糖酶基因（Cm Tre1）为靶标,设计RNAi靶位点并将其命名为Cm Tre1＊,在其两端分别添加Bam H I与Spe I酶切位点并进行人工合成,然后经双酶切后与p1301植物表达载体连接,构建重组表达载体p1301-Cm Tre1＊。PCR、双酶切和测序鉴定结果证明重组表达载体p1301-Cm Tre1＊构建成功,并将其成功转入农杆菌LBA4404,构建基因工程菌。用含有p1301-Cm Tre1＊质粒的农杆菌LBA4404浸染中花11号水稻的愈伤组织,经过愈伤组织的抗性筛选、分化和植株再生,获得26株阳性转基因植株。用取食非转基因水稻的稻纵卷叶螟作为对照,用实时荧光定量PCR方法分株测定取食转基因水稻稻纵卷叶螟体内Cm Tre1基因的mRNA表达水平,并检测其体内海藻糖酶活性变化。结果显示,相较于对照组,取食转基因水稻稻纵卷叶螟体内Cm Tre1基因表达量下降了44.79%,海藻糖酶活力平均下降了14.94%。研究结果表明转基因水稻的RNA干扰效应相当明显,能对取食其叶片的稻纵卷叶螟Cm Tre1基因起到明显的沉默作用。

英文摘要：

RNA interference（ RNAi） refers to a biological phenomenon in which RNA molecules inhibit gene expression,typically by causing the degradation of homologous mRNA molecules. RNAi is a gene silencing process induced by double- stranded RNA molecules and is a highly evolutionarily conserved mechanism of gene regulation. In this paper,target sequence specific to soluble trehalase gene（ Cm Tre1） from Cnaphalocrocis medinalis was designed and synthesized with Bam H I and Spe I restriction enzyme cutting sites at both sides,and was named Cm Tre1＊. After double enzyme digestion,this fragment was inserted into p1301 to construct recombinant expression vector p1301- Cm Tre1＊. The results of PCR,enzyme digestion,and sequencing showed that the recombinant vector had been successfully constructed. Then this vector p1301-Cm Tre1＊was transformed into Agrobacterium tumefaciens LBA4404 to construct genetic engineering bacteria.The calli of rice Zhonghua 11 were infected by A. tumefaciens LBA4404 that contained p1301- Cm Tre1＊.Then the calli were screened by higromycin. After differentiation of the resistant calli and and plant regeneration,26 positive transgenic rice plants were obtained. Cm Tre1 mRNA expression levels in C. medinalis that fed on these transgenic rice were detected with real time quantitative PCR（ RT- q PCR） and trehalase activities in vivo were tested by the Trehalase Kit,using C. medinalis that fed on non- transgenic rice as controls.The results showed that the average expression level of Cm Tre1 in C. medinalis feeding on transgenic rice decreased by 44. 79% and the average trehalase activity in vivo fell by 14. 94%,compared to the control groups. The findings indicated that the transgenic rice plants containing the interference sequence Cm Tre1＊could strongly silence Cm Tre1 in C. medinalis that fed on these transgenic rice.