【作者】 邓力喜；

【导师】 梁满中； 陈良碧；

【作者基本信息】 湖南师范大学 ， 植物学， 2009， 硕士

【摘要】 长选3S是从温敏核不育系培矮64S群体中选育出来的一种对赤霉素敏感的长穗颈突变体,受一对隐性基因控制。本文以培矮64S为对照,采用水稻基因芯片和半定量RT-PCR技术对长选3S最上节间快速伸长期全基因组表达模式进行分析,主要结果如下:1、在具有60000个杂交点,代表36926个单一基因的的寡聚核苷酸基因芯片上,检测到545个差异表达基因,其中有362个为上调表达的转录本,占差异表达基因的66.42%;183个为下调表达的转录本,占差异表达基因的33.58%。通过半定量RT-PCR对差异基因做进一步分析,结果与基因芯片结果一致,表明芯片结果中基因表达差异的可靠性高。2、通过生物信息学分析发现,这些差异表达的转录本包括信号传导相关基因、膜及运输基因、细胞生长与凋亡相关基因、复制与修复相关基因、碳水化合物代谢相关基因,次生代谢物合成相关基因和其它功能未知基因。在信号转导相关基因以及碳水化合物和次生代谢物相关基因中发现许多与节间伸长有关的基因。eui基因可能通过影响这些相关基因的表达,实现最上节间快速伸长。3、从芯片结果中选择表达差异达10倍以上的2个基因进行生物信息学分析,表明其为未知功能基因,其启动子区存在激素响应元件,可能与激素信号转导有关。根据基因序列设计引物,通过RT-PCR扩增,获得了包含基因完整ORF的cDNA克隆。测序结果表明他们的序列与9311完全相同,而与日本晴存在碱基差异。构建了该基因的过表达载体,并通过农杆菌介导法成功将该基因转化到拟南芥中,获得了转基因植株。更多还原

【Abstract】 Changxuan 3S was obtained from thermo-sensitive genic male sterile line(TGMS) Pei’ ai64S. It is sensitive to gibberellins and is controlled by single recessive gene. In this research, the gene expression patterns in elongated uppermost internode of TGMS eui mutant Changxuan 3S through oligonucleotide microarray and semi-quantitative PCR, using Pei’ ai 64S as control. The data obtained in this research are mainly as follows:Firstly, we found 545 differentially expressed genes on the genechip with 60,000 hybridization sites representing 36,926 transcripts. Among these differentially expressed genes, 362 genes were up-regulated, and 183 genes down-regulated, which accounted for 66.42% and 33.58% separately of total differentially expressed genes. The result of further semi-quantitative RT-PCR analysis showed the reliability of the result of microarray analysis.Secondly, we found that the differentially expressed genes could be grouped into seven categories according to bioinformatics analysis. That was: signal transduction-associated genes, membrane and transport associated genes, cell growth and apoptosis associated genes, replication and repair associated genes, carbohydrate metabolism associated genes, biosynthesis of secondary metabolites associated genes and others with unknown function. In signal transduction-associated genes, carbohydrate metabolism associated genes and biosynthesis of secondary metabolites associated genes, we found lots of internode elongation related genes. eui gene may affect these gene expression to achieve rapid elongation of uppermost internode.Thirdly, we further analyzed two genes differentially expressed greater than 10-fold. It was showed that they were genes with unknown function, but there were hormone-responsive elements in their promoters. So, they might involve in hormone signal transduction. We designed specific primers according to their gene sequences, and cloned the cDNA sequences containing the whole open reading frame of the genes by RT-PCR. Their cDNA sequences are identical with 9311’ s, but different from Nipponbare’ s. Then we constructed gene over-expression vectors and delivered the genes into Arabidopsis successfully. The analysis of the transgenic Arabidopsis is in progress.更多还原