控制水稻籽粒充实和产量的基因GIF2的图位克隆及功能研究

Rice in an important food crop of China, filling characteristics of rice grain and metabolism in the filling process are the key that affects rice yield. The materials which were used in our study are a rice imcomplete filling mutant A183 and its wild type. The fine mapping work of this mutated gene has been finished, after comparison of the analysis of candidate gene and gene sequencing, candidate gene ORF4 was found which first exon had two base deletion. The gene map-based cloning work and gene function analysis will be carried out, including functional complementary experiment and functional verification, gene expression, subcellular localization, and also the comparison of accumulation of dry matter, enzyme activities and the parameter of physiology and biochemistry of the mutant and its wild type during rice grain-filling process. The results of the gene function analysis and its expression in space and time characteristic can help us to discover its regulate mechanism. Understanding the factors that regulate grain filling is important for improving rice yield potential．

水稻在我国粮食生产中具有重要地位，水稻产量的提高，离不开水稻籽粒充实过程的研究。本项目利用灌浆不饱满突变体A183为研究材料，已完成该基因的精细定位工作，通过对候选区域基因预测及测序对比分析，获得候选基因ORF4（命名为GIF2）的第一外显子发生了2个碱基缺失，接下来拟对该基因进行图位克隆，围绕该基因功能开展相关研究，通过对突变基因的功能互补、组织表达、编码蛋白的亚细胞定位、突变体与野生型基因酶活比较、籽粒充实过程中各生理生化参数比较、胚乳干物质积累动态变化比较等方法来研究该基因的功能，明确GIF2基因对籽粒充实过程和粒重影响的分子机理，对于进一步揭示水稻结实机理，促进籽粒的灌浆和充实，最大限度地发挥品种生物学潜力，有十分重要的理论及实践意义。

本项目以水稻灌浆不饱满突变体gif2 为材料，通过 SSR, InDel 等分子标记精细定位并克隆该突变体基因，测序分析表明ORF1 的第一外显子发生了2 个碱基的缺失，造成移码，翻译提前终止。进一步研究表明该突变是编码焦磷酸化酶大亚基2 的基因发生突变导致的突变表型，突变体灌浆过程中焦磷酸化酶活性远远低于野生型。葡萄糖焦磷酸化酶是淀粉合成的关键酶，该基因突变，蔗糖向淀粉的转化受阻，影响淀粉的生物合成，可溶性糖含量测定结果显示，灌浆过程大量的蔗糖未完成向淀粉的转化，仍然以自由糖的形式存在于成熟胚乳中。GIF2 编码蛋白主要定位于细胞质中，同时GIF2 突变导致了突变体中淀粉合成相关基因表达量显著上升，这可能和GIF2 的反馈调节有关。本项目结果丰富了水稻籽粒灌浆调控的机理研究，并从分子水平上阐明籽粒灌浆遗传调控机制。