【作者】 张莉；

【导师】 李润植； 韩渊怀；

【作者基本信息】 山西农业大学 ， 作物遗传育种， 2004， 硕士

【摘要】 早在1988 年,Kooter 博士研究小组就发现在矮牵牛中转入反义的chs (chalcone synthase) 基因(编码苯基苯乙烯合成酶),可以抑制花色素的表达。同年,Dr. Grierson 博士研究小组也在西红柿中转入反义的PG (Polygalacuronase) 基因(编码多聚半乳糖醛酸酶) ,发现在转基因的西红柿中,内源的PG 基因表达受到明显抑制。两年后,Kooter 等和Jorgensen 等同时报道了在植物中转入正义的基因同样可以使该基因及其内源的同源基因沉默。人们将正义和反义转基因引起的基因沉默现象分别称为共抑制和反义抑制。共抑制与反义抑制有如下相似的特点:1.转基因编码区与受体细胞基因组间具有同源性;2.来源于内源基因的mRNA 高度降解;3. 产生与目标mRNA 序列类似的异常转录物;4.具有与病毒基因同源的正义或反义转基因植物,对该病毒都有抗性。然而,Jorgensen 等(1998)在对chs 转基因矮牵牛的进一步研究发现,正义转基因与反义转基因矮牵牛花的表型特征有明显差异。表现为正义转基因矮牵牛和反义转基因矮牵牛的花色素表达被抑制的时间,空间和程度上不同。这又暗示着共抑制与反义抑制可能以不同机理使目标基因发生沉默。因而,有必要深入探讨这两种基因沉默现象的机理。为此本实验试回答以下两个问题:一.共抑制与反义抑制有无联系?二.反义抑制是否也存在像共抑制一样可以传导的基因沉默信号? Baulcombe 等(2000)利用中子诱变技术获得了沉默缺失的突变体sde(silencing defective)-1, 2, 3。这些突变体均含有35S-GFP 转基因和35S-PVX-GFP(Potato Virus X-GFP)转基因。我们构建了由CMV-35S 强启动子控制的反义GFP 表达结构,通过农杆菌介导将其转入上述突变体中,以期观察该反义转基因能否激活上述沉默突变体重新发生基因沉默。迄今,我们更多还原

【Abstract】 Sense and antisense transgenes have been used to down-regulate genes in plants. Although there is considerable variation in both sense and antisense transformants, have been noted several striking similarities between down-regulation of gene expression by sense and antisense transgenes: (1) Both are remarkably effective and specific for the target gene; (2) Both cause a dramatic reduction in the amount of mRNA that accumulates from the endogenous gene; (3) Both lead to the production of similar fragments of endogenous mRNA (4) Both resist to the virus which has some homologous nucleotide sequences with sense and antisense transgene. It seems that sense transgene silencing and antisense transgene silencing share similar mechanism, however, there are some evidence showing that they may be different in some aspects, such as the different phenotype they cause in petunia. We therefore, proposed to compare the mechanism of sense and antisense silencing. We carry out two different experiments: the first one is to test if antisense GFP can cause release of silencing of existing GFP in gene silencing mutants sde-1, 2, 3. the second one is to check whether or not silencing signal can transfer from antisense induced Aco1 gene silencing stock to Aco1 over expression scion with the reference that systemic silencing is a feature of sense transgene induced gene silencing.更多还原