板栗开花相关基因的克隆和功能初步分析
其他题名Isolation and characterization of related flowering-time gene from chestnut
刘涛
学位类型博士
导师周浙昆
2007-05-25
学位授予单位中国科学院昆明植物研究所
学位授予地点昆明植物研究所
学位专业植物学
关键词板栗 Mads-box基因 Lfy同源基因
摘要本研究以板栗为研究材料,研究了内源激素和可溶性糖代谢在板栗花发育中的作用;同时从板栗的花序中分离得到了四个与花序发生和发育有关的三个MADS-box同源基因和一个LFY同源基因。并分析了它们的序列结构、系统发育关系和表达式样,探讨了板栗花序发育的分子机理。主要研究结果包括: 1. 分析了激素和可溶性糖代谢在异常开花板栗株和正常株一年期间的含量对比变化。研究结果表明:它们的ZR(玉米素)、ABA(脱落酸)和GA(赤霉素)含量变化展现不同的式样,而IAA(生长素)和可溶性糖表现相似的变化趋势。推测ZR在板栗开花中可能扮演着抑制开花的角色;内源IAA、ABA和GA扮演着促进板栗开花的角色;可溶性糖可能为板栗花芽的早期发育提供所需能量;并对内源激素和糖代谢在异常株异常开花过程中可能的作用进行了讨论。 2. 以板栗叶与花序为材料,通过cDNA-AFLP方法,采用64对引物组合,共得到2131条多态片断。其中在花序中特异表达的条带数为466条,在叶中特异表达的条带数为484条。它们的共有条带数为1181条。将110条进行克隆测序,结果在GeneBank中比对,发现有28条序列归属于已知的同源序列,其中包括参与转运﹑代谢﹑和能量产生相关的蛋白。 3. 从板栗花序中分离克隆出了三种不同亚族的MADS-box基因:CmMADS2、CmMADS3和CmMADS4。氨基酸序列分析结果证明它们都是MIKCc型MADS-box基因。系统发育分析结果表明CmMADS2与AG类基因聚在一起,CmMADS3与SEP1/2/3/4类基因聚在一起,CmMADS4与AP1/SQUA类基因聚在一起。同时用RT-PCR的方法分析了CmMADS2、CmMADS3和CmMADS4在板栗不同组织中的表达式样,对它们在控制板栗花发育中的作用进行了讨论。 4. 从板栗花序中分离出了控制板栗花序发育的CmLFY基因,用RT-PCR的方法分析了CmLFY在板栗不同组织中的表达式样。拟南芥转基因实验结果显示:表达CaMV 35S::CmLFY的转基因T1代拟南芥表现出提前开花的行为,表明CmLFY确实是具有LFY功能的同源调控基因。
其他摘要In our experiment, the role of the endogenesis hormone and soluble sugar in the flower development of chestnut was examined. Three MADS-box homologous gene and one LFY homologous gene that are potentially important for the initiation and development were isolated from chestnut inflorescence. Furthermore, the sequence structure, phylogenetic relationship and expression patterns of three MADS-box homologous genes and the LFY homologous gene were characterized respectively. Based on these results, their genetic and molecular mechanisms controlling chestnut flowering were discussed. The main results are shown as follows: 1. Examination was done about the contents change of endogenesis hormone and soluble sugar between abnormal and normal flowering chestnut monthly for a year. The results show that the contents change of zeatin riboside (ZR), abscisic acid (ABA) and gibberellins (GA) are significantly different between the normal and abnormal trees. The contents change of indoleacetic acid (IAA) and soluble sugars show a similar pattern between them. We speculated that ZR might repress flowering of chestnut, while IAA, ABA and GA might acceletate flowering. Soluble sugar might provide energe for early development of flower. The roles of endogenesis hormone and soluble sugar in the abnormal flowering were discussed. 2. The expression profiles between inflorescences and leaves of chestnut were detected by cDNA-AFLP, and 2,131 fragments were found using 64 combinations of AFLP primers in all. Among them, 484 and 466 are specific to leaves and inflorescences respectively, 1181 are common to leaves and inflorescences. 101 of the fragments were choised for cloning and sequencing and the results of sequnce blast show that 28 fragments are homologous of the function known entries in the GenBank, including transport facilitation, metabolism and energy-related proteins. 3. Three MADS-box homologouses, named CmMADS2、CmMADS3 and CmMADS4 belonging to different sub-families were isolated from chestnut inflorescence. The analysis of amino acid sequences revealed that they are all MIKCc type MADS-box genes. Phylogenetic analysis indicated that CmMADS2, CmMADS3 and CmMADS4 are clustered with AG-like genes, SEP1/2/3-like genes and AP1/SQUA-like genes respectively. Besides, the expression patterns of these genes during different cultures were studied by RT-PCR. The functions of these genes in the development of flower development were discussed. 4. CmLFY, the LFY homologous gene that is important for the initiation and development was isolated from chestnut inflorescences. Moreover, its expression pattern in different cultures was investigated by RT-PCR. In order to verify its function further, the 35S::CmLFY was translated into Arabidopsis. The transgenetic plant T1 flower ahead of time. This result strongly support that CmLFY is a LFY homolog.
页数125
语种中文
文献类型学位论文
条目标识符http://ir.kib.ac.cn/handle/151853/274
专题昆明植物所硕博研究生毕业学位论文
推荐引用方式
GB/T 7714
刘涛. 板栗开花相关基因的克隆和功能初步分析[D]. 昆明植物研究所. 中国科学院昆明植物研究所,2007.
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