【作者】 韩丹萍；

【导师】 张耀伟；

【作者基本信息】 东北农业大学 ， 蔬菜学， 2011， 硕士

【摘要】 大白菜(Brassica rapa L. ssp. pekinensis)的先期抽薹,是在大白菜的营养体还未充分长成之前,在不该抽薹时提前抽薹的现象。抽薹后的大白菜会失去食用价值,大大降低经济效益。因此,研究其抽薹开花相关的分子机制,对白菜育种具有重要意义。大白菜在春化后会抽薹开花,在抽薹前表现出一个叶色转变过程,叶片颜色从鲜亮的绿色转为没有光泽的暗灰色后花茎就会伸长并开始现蕾抽薹,叶色转变似乎是大白菜抽薹的一个生理信号,在弱冬性品种上的表现尤为突出,因此推测叶色转变与抽薹开花有关,叶色转变前后基因和蛋白表达必将发生变化。本试验从抽薹开花相关基因表达和蛋白质表达两个方面研究大白菜生殖发育转变过程中的分子机理。以四个冬性不同的大白菜品种为材料,分别采取春化前、春化后、花芽分化、花茎分化、叶色转变、花茎伸长六个时期的植株生长点部位新生幼嫩叶片,来测定叶片激素含量,并进行FT和LFY两个开花相关基因的RT-PCR表达分析和蛋白质双向电泳分析;另外在每个品种的叶色转变期取植株的根、短缩茎、茎端生长点幼嫩叶片、近端叶片、远端叶片五个部位,进行FT和LFY基因的RT-PCR表达分析。主要试验结果如下:1.通过测定激素含量的变化,得知它与大白菜抽薹开花过程中叶色的转变有一定关系,大白菜抽薹过程中, IAA和GA3含量均在叶色转变期出现峰值,叶色转变后花茎开始伸长,叶色转变是大白菜抽薹的一个生理信号。2.FT基因在春化前和春化后也即整个的营养生长阶段不表达或微量表达,在花芽分化期以后表达强度递增,在叶色转变期和花茎伸长期表达强烈,另外还发现FT基因在弱冬性品种中要比在强冬性品种中表达强烈。FT基因在根和短缩茎中不表达,在生长点嫩叶中和近端叶片、远端叶片表达,在弱冬性品种中,FT在近端叶片中强烈表达,在远端叶片中表达极微弱,而在强冬性品种中,FT在近端叶片与远端叶片中表达均较强。3.LFY基因在春化前不表达,花芽分化期和叶色转变期和花茎伸长期表达较强,在生长点嫩叶中表达最强烈,在近端叶片和远端叶片中也有较强表达,在短缩茎中有极微弱表达,在根中不表达。4.蛋白质表达图谱显示叶色转变后有18种新的特异蛋白质产生,在花茎分化期表达的17种蛋白质在叶色转变期停止表达;有29种蛋白质在叶色转变后表达上调,还有21种蛋白质表达下调。5.对17个蛋白点进行质谱鉴定,成功鉴定出其中的16个点,这些蛋白1个与转录调控有关,2个与信号传递和能量传递有关,6个与细胞生长及代谢有关,还有1个是结构蛋白,此外还有1个未知蛋白,5个功能未知的假设性蛋白质。更多还原

【Abstract】 Premature bolting of Chinese cabbage (Brassica rapa L. ssp. pekinensis) is a phenomenon that bolting at a improper time before its vegetative plants ripen, then the vegetable will lose value,It causes a great loss in production. It has great significance to study the Physiological and Molecular mechanism connected with bolting of Brassica.rapa,especially the genes associated with bolting and flowering in vernalization pathway.Chinese cabbage will bolting and flowering after vernalization , the leaf color changes before the bolting, stems elongate and the plant start to bud and bolt after the leaf color changes from bright green to dark grey without being shiny or lustrous .the changes of the leaf color seems to be a physiological signal of the bolting for Chinese cabbage ,the phenomenon is particularly acute for the weak winterness cultivars, so we speculate that the changes of leaf color is relevant with the bolting and flowering process. the genes and protein expression varies before and after leaf color changes. In present study, we used four cultivars of Chinese cabbage that were different in winter to study the molecular mechanism of reproductive development by testing the related genes and proteins expression. We took samples of fresh young leaves in six development stages for hormone levels Determination and 2-D electrophoresis,which are before and after vernalization, floral bud differentiation period,stem differentiation period,the period of leaf color transition and the stem elongation period. Then we took samples at root,dwarf stem, fresh leaves at growing point, proximal leaves, distal leaves in the period of leaf color transition for FT and LFY RT-PCR all together with the samples mentioned above. Main results were as follows:1. It turns out that the hormone content is associated with the leaf color changes in the bolting process, different winterness varieties get content peaks of IAA and GA3 in the period of leaf color transition respectively in bolting process. The stems begin to elongate after this stage.The changes of leaf color is a physiological signal in cabbage bolting.2 . FT gene expresses stronger in weak winterness cultivars than in strong winterness cultivars; It doesn’t express or express very weak in the whole vegetative growth stage, it expresses stronger and stronger after the bud differentiation stage, and expresses very strongly in the leaf color transition period and stems elongation period .FT gene doesn’t express in roots and dwarf stems, it expresses in the fresh leaves at growing point, proximal leaves and distal leaves. FT expresses strongly in proximal leaves and very weak in distal leaves in weak winterness cultivars, however, in strong winterness cultivars,FT expresses both strongly in proximal leaves and distal leaves .3. LFY gene doesn’t express before vernalization,expresses strongly in the bud differentiation period,the leaf color transition period and stems elongation period,most strongly in the fresh leaves at growing point.it expresses strongly in proximal leaves and distal leaves,it doesn’t express in roots ,and express very weak in dwarf stems.4. We found 18 new proteins espresses after leaf color changes,and 17 proteins don’t espresses anymore after this transition, There are also 21 proteins are up-regulation of experssion and 29 are down-regulation of experssion according to analysis and comparation.5. 17 protein spots were identified using MS.and 16 of them are successfully identified. 1 of them is related with transcriptional regulation, 2 of them are related with signal or energy transmission,6 proteins participate in cell growth and metabolism,and the last one is structural protein.更多还原