水稻胚乳特异性表达基因启动子的鉴定

Identification of rice endosperm-specific gene promoters

水稻胚乳既是生产重组蛋白的理想部位,也是食物营养价值改进的重要目标组分。缺少胚乳特异性表达强启动子是限制从胚乳中获得重组蛋白和进行营养成分改良的主要因素之一。本论文采用RNA-Seq从全基因组水平筛选获得1040个胚乳特异性表达基因。经分析,所得胚乳特异性表达基因启动子大都含有GCN4、AACA和Prolamin-box等胚乳特异性基序以及与胚乳中贮藏相关蛋白调控表达有关的O2-sites。从1040个胚乳特异性表达基因中随机选取8个进行实时荧光定量PCR(qRT-PCR)分析,发现这些基因都在胚乳中特异性表达。^(pro)moter:GFP实验结果表明OsEnS100^(pro)、OsEnS93^(pro)、OsEnS80^(pro)和OsEnS50^(pro)均可驱动GFP基因在胚乳中特异性表达,且OsEnS100^(pro)和OsEnS93^(pro)启动子活性更高。综合以上,本论文筛选获得的潜在胚乳特异性表达基因将为植物基因工程提供潜在的胚乳特异性强启动子,同时为水稻胚乳发育的分子机理研究提供基因资源。

Rice endosperm is an ideal biological synthesizer for recombinant protein production and also an important target organ for improving the nutritional value of rice.The lack of endosperm-specific(OsEnS)gene promoter is a major factor limiting recombinant protein production and nutrient improvement of rice endosperm.In this study,we used RNA-Seq to screen endosperm-specific genes genome-wide,which revealed 1040 potential endosperm-specific genes.Sequence analysis of these gene promoters showed that most of the promoters contain the GCN4,AACA and Prolamin-box endosperm-specific motifs,and O2-site,which may be involved in controlling the expression of storage-related proteins in endosperm.We randomly selected eight genes for further research.Quantitative real-time PCR(qRT-PCR)results confirmed that all the selected genes were specifically expressed in endosperm.Furthermore,promoter:GFP reporter gene experiment revealed that OsEnS100^(pro),OsEnS93^(pro),OsEnS80^(pro) and OsEnS50^(pro) could induce the expression of GFP specifically in endosperm,and the promoter activities were much higher in OsEnS100^(pro):GFP and OsEnS93^(pro):GFP.Taken together,the study identified potential endosperm-specific genes,which may provide potential endosperm-specific promoters for gene engineering with rice endosperm.These genes also provide candidates for further research of the molecular mechanisms underlying rice endosperm development.