Abstract:

Dicer enzyme is RNase Ⅲ-like endonuclease that can specifically cut double-stranded RNA into about 20 nt small RNA, and plays a very important role in gene-silencing pathways. In this research, DCL1 gene promoter was isolated, including 2 kb segments upstream of transcription start site and 1 kb segments downstream of transcription start site, and inserted into upstream of GUS reporter gene to construct a plant expression vector. This expression cassette was introduced into wild Arabidopsis thaliana by Agrobacterium-mediated transformation and the transgenic plants were used for GUS staining and fluorescent quantitative analysis. The results indicated that GUS gene was mainly expressed in mature leaves, higher in young leaves and shoot tip, lower in stem and only a little bit in roots.

Key words: Arabidopsis thaliana, gene silencing, DCL1, small RNA, promoter, GUS staining

摘要：

Dicer酶是类似于RNase Ⅲ的核酸内切酶,能够特异性地将双链RNA剪切成约20 nt的小RNA,在基因沉默途径中起到非常重要的作用。从拟南芥中分离出DCL1基因上游启动子序列2 kb及DCL1基因的5′端1 kb序列,构建了含有该启动子和GUS报告基因的植物表达载体,通过农杆菌介导法转化拟南芥,并对转基因植株进行GUS 组织化学染色及荧光定量分析,结果表明,在DCL1基因启动子的驱动下,报告基因GUS主要在拟南芥的叶片中表达,在幼嫩的叶片及茎尖中表达量也比较高,茎中的表达量较低,在根中只有微量的表达。

关键词: 拟南芥;基因沉默;DCL1;小RNA;启动子;GUS染色