中文摘要：

RNA干扰（RNAi）技术是研究基因功能的一种常用方法,为了快捷高效地构建水稻（简易RNAi表达载体,本研究采用简易RNAi构建法,设计3＇末端9个碱基互补的正向和反向寡聚核苷酸引物,通过聚合酶延伸成为具有反向重复序列的小干扰片段.以pCAMBIA 1301-Gt 13a为表达载体,将小干扰片段与表达载体经过一次双酶切与连接得到RNAi载体.用农杆菌（Agrobacterium tumefaciens）介导法得到水稻（Oryza sativa ssp.japonica）转基因株系后,用叶片GUS染色和qRT-PCR法,对RNAi转基因水稻和野生型日本晴叶片GUS组织活性和籽粒中目的基因表达量进行检测.水稻叶片GUS染色结果显示,有10株转基因水稻叶片切口处呈现蓝色,而野生型无显色反应,说明RNAi载体T-DNA区域的基因已整合到水稻基因组上,并表达.在mRNA水平上转基因水稻籽粒中目的基因相对表达量降低至18％～55％,差异极显著（P＜0.01）,表明整合在水稻基因组上的小干扰片段能降低目的基因的表达水平.籽粒灌浆成熟后对10株转基因水稻及野生型日本晴籽粒的单粒重、粒长、粒宽和厚度分别进行测量.与野生型日本晴相比,转基因水稻籽粒单粒重发生显著性降低（P＜0.05）,籽粒变小,长和宽都发生显著性变化（P＜0.05）,说明简易RNAi载体对水稻目的基因的表达起到很好的沉默效果.本研究不仅为利用简易RNAi构建法研究水稻功能提供了依据,也为快捷高效地研究水稻基因功能提供了基础资料.

英文摘要：

RNA interference （RNAi） technique has been used on gene function research frequently.In order to construct a kind of simple RNAi expression vector which is adapted to rice（Oryza sativa ssp.japonica） rapidly and efficiently,a pair of forward and reverse primers oligonucleotides with 9 bp of complementary nucleosides at 3＇ end was designed,and a small interfering segment with inverted repeated sequence of target gene by polymerase extending was formed,which was digested and ligased into the basic expression vector pCAMBIA1301 to construct RNAi vector with an inverted repeated sequence.After Agribacterium tumefaciens transformation,both wild type and transgenic lines of rice were validated by leaf GUS staining and qRT-PCR of target genes in grains.The GUS staining result showed that the cut edges of transgenic rice leaves were blue,and there was none in the wild type,suggesting that the genes within T-DNA region of RNAi vector had already been integrated into the rice genome and expressed,qRT-PCR result showed that the rnRNA relative expression quantity of the target gene decreased to 18％～55％（P＜0.01）,showing that the small interfering segment within rice genome could decrease the expression level of target gene.Comparing with wild type rice,the single kernel weight of 10 transgenic rice plants decreased significantly （P＜0.05）,and the size of transgenic rice kernel was smaller,both length and width changed significantly（P＜0.05）.This showed that simplified RNAi vector could effectively interfer target genes expression.This research provides a simple RNAi construction for quick and efficient rice gene function research.