【作者】 杨翅春；

【导师】 朱登云；

【作者基本信息】 中国农业大学 ， 遗传学， 2005， 硕士

【摘要】 根据油菜花瓣特异性表达基因(GeneBank序号BD082011)的序列设计引物,利用PCR的方法扩增出该基因的启动子序列,大小为355bp,命名为XY355。将克隆得到的XY355启动子分别与玉米花青素调节基因Lc及改造过的黑色素基因连接,构建植物表达载体pXY60、pYC26。 用农杆菌介导的方法将pYC26、pXY60、pQT56(35S启动子驱动花青素调节基因Lc)转化烟草,将pYC26、pXY60转化矮牵牛。对再生植株进行了PCR检测:转基因烟草中,pYC26阳性植株有23棵,pXY60阳性植株有20棵,pQT56阳性植株有18棵。转基因矮牵牛中,pYC26阳性植株有15棵,pXY60阳性植株有16棵。 对转基因植株的花色表型进行观察。对照组(转pBI121)的矮牵牛的花筒为紫色,花冠为白色;而转pXY60的矮牵牛中,16棵阳性植株有8棵矮牵牛的表型发生变化,花冠变成了浅紫色,花筒颜色没有明显的变化;转pYC26(XY355启动子驱动Lc黑色素基因)的矮牵牛花筒和花冠的颜色均没有发生明显的变化,但是在转基因PCR阳性植株发现一棵由于花形发生变化而导致的花色发生了变化 相对于对照组(转pBI121)的烟草,转pXY60(XY355启动子驱动Lc)的烟草,20棵PCR阳性植株有10棵表型发生变化,花色变成了红色;转pQT56(35s启动子驱动Lc)的烟草中,18棵PCR阳性植株中有15棵表型发生变化,花色变成了深红色。转pYC26(XY355启动子驱动黑色素基因)的烟草花色没有发生明显的变化。更多还原

【Abstract】 The primers were designed according to the sequence of Brassica napus’ (rapes’) petal-specific promoter (GeneBank No. BD082011), and the promoter was amplified by PCR which was named XY355. The XY355 promoter was connected with maize’s anthocyanin regulatory gene Lc, and with a modified melanin gene respectively to construct expression vectors pXY60 and pYC26.The expression vectors pYC26, pXY60, and pQT56 (the expression vector provided by our laboratory was containing CaMV35 promoter and Lc gene) were introduced to the Agrobacterium tumefaciences LBA4404.Then the leaf discs of petunias and tobaccos were transformed by Agrobacterium tumefaciences LBA4404.Some regenerated plants had been chosed at random and checked by PCR. Among these paints, there were 61 positive tobaccos including 23 plants of pYC26, 20 plants of pXY60, and 18 plants of pQT56, and there were 31 positive petunias including 15 plants of pYC26 and 16 plants of pXY60.Transgenic plants’ flowers color had been observed. The negative control petunia lines (transformed by pBI121) had purple flower tubes and white.corollas. 8 plants of 16 pXY60 PCR-positive petunias had light purple corollas. Most pYC26 positive petunias had not evident alteration of flower color. However, one of pYC26 PCR-positive petunias had changed its flower color caused by a alteration of the flower shape.The negative control petunia lines (transformed by pBI121) had pink petals. 10 plants of the 20 pXY60 PCR-positive tobaccos had red petals and anther filaments. 15 plants of 18 pQT56 PCR-positive tobaccos had intense red petals and anther filaments. As for the tobaccos transformed by pYC2, no obvious changes in flower color had been observed.更多还原