【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。更多还原

【Abstract】 Enhancing the expression of phytoene synthase (Psy )gene and blocking the expression of lycopene cyclase (Lcy )gene, which are the main genes regulating the transformation of the lycopene,are 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 cDNA of the Arabidopsis thaliana. The phytoene desaturase 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(Dds );the Psy gene was also inserted into the same vector under the manipulation of the Pds promoter,then the expression vector was obtained and could enhance the expression of Psy gene and interfere the expression of Lcy gene at the same time by transforming it into the plant.更多还原