加强Psy基因表达和通过RNAi抑制Lcy基因表达的载体构建  

Construction of Expression Vector for Enhancing Psy Gene Expression and Blocking Lcy Gene Expression through RNAi

作　　者：万群[1] 张兴国[1] 宋明[1] 苏承刚[1] 

机构地区：[1]西南大学园艺园林学院,重庆市蔬菜学重点实验室,重庆400716

出　　处：《农业生物技术学报》2007年第3期477-481,共5页Journal of Agricultural Biotechnology

摘　　要：根据GenBank中拟南芥的八氢番茄红素(Psy)基因序列从拟南芥(Arabidopsis thaliana)的cDNA中分离了一段长1296bp的Psy基因;根据GenBank中番茄的番茄红素β-环化酶(Lcy)序列和八氢番茄红素去饱和酶基因(Pds)启动子序列分别从番茄(Lycopersicon esculentumMill.)DNA中扩增出了Lcy基因的高度保守的含302bp的DNA片段和含1790bp的Pds启动子片段。根据RNAi的原理,将Lcy基因的DNA片段以正反两个方向通过一段内含子序列连接在一起形成RNAi片段,将该片段与Psy基因一起插入到抗Kan的pVCT2020的表达载体中,并通过具有果实特异性特点的启动子Pds启动子调控这两个片段的表达,从而构建成了能同时加强Psy基因的表达和抑制Lcy基因表达的果实特异性植物表达载体,通过酶切鉴定证明所构建的载体正确。Enhancing the expression of phytocne syrthasc （Psy）gene and blocking the expression of lycopcne cyclase （Lcy） gene, which arc the main genes regulating the transformation of the lycopene, arc the convenient way to increase the content of lycopene. The primers were designed according to the gene sequence NM-121729, U46919 and X86452 in GenBank. The Psy gene was isolated from the eDNA of the Arabidopsis thaliana. The phytocne desaturasc gene （Pds） promoter and the DNA segments of the Lcy gene were obtained from the genome DNA of tomato （Lycopersicon esculentum Mill.）, The 3＇end of Lcy DNA segment was connected together with an intron to inform the RNA interference （RNAi） segment and inserted into the expression vector with the fruit-specific promoter（Pals ）;the Psy gene was also inserted into the same vector under the manipulation of the Pals promoter,then the expression vector was obtained and could enhance the exnression of Psy gene and interfere the expression of Lcy gene at the same time by transforming it into the plant.

关 键 词：番茄 Psy基因 Lcy基因 Pds启动子 RNAI 

分 类 号：S188[农业科学—农业基础科学]