一个新的水稻显性大粒基因的图位克隆、功能分析和育种利用

Grain weight is one of the most important components of rice yield. To date, more than ten genes related to grain weight were cloned, which indicated the complexity of grain weight controlling. Furthermore, most grain weight related genes are functinal loss in their large-grain alleles, which restricts the immediate application on hybrid-rice breeding. We have previously identified a large-grain rice line with thousand kernel weight of 64g, and proved the large grain trait was controlled by a new major dominant QTL,GS2. By using of the F2, BC2F2, BC3F2 populations, the gene was mapped to a 10 Kb interval and the candidate was supposed. We also developed NILs for several restorer lines with different grain size and proved GS2 improved both grain length and width, result in a significant improvement of the grain weight, by incresing cell size of the spikelet hulls. In the present study, we use the BC4F2 population and the Fixed Recombinants to verify the result of fine-mapping, and aim to clone and functionally anlyize GS2 gene by functional complement, over-expression and down-regulation of GS2 expression. We also want to disclose its expression pattern by using the method of GUS staining and YFP-labeling. Besides, we aim to evaluate the potential of GS2 in high-yield breeding of hybrid-rice,by investigating the field performance of NILs and the corresponding F1 hybrid-rice. Our result will bring new insight for grain weight controlling and provide immediate implications for rice yield improvement.

粒重是水稻单产的重要构成因素，迄今已有十余个相关基因被克隆，显示出粒重控制机理的复杂性。同时，多数基因的大粒等位变异表现为功能缺失，限制了其在杂交水稻中的直接应用。我们发现了一个水稻特大粒资源材料，千粒重约64克。遗传分析表明该大粒性状主要受一个主效显性QTL控制，利用F2和多代回交群体将该基因定位于第2染色体10Kb范围，为一个新的粒重控制基因，暂定名为GS2。对不同粒重恢复系构建近等基因系发现，该基因可以显著提高各恢复系的千粒重。细胞观察发现该基因主要通过增加颖壳细胞大小同时增加粒长和粒宽，进而提高粒重。本研究拟在进一步定位验证的基础上，克隆候选基因，构建互补、过量和下调表达载体，验证该基因的功能；并通过GUS染色和YFP融合蛋白方法研究该基因的表达模式。同时，创制主流恢复系的近等基因系并探讨其在杂交稻育种中的应用。本研究将为粒重控制机理提供新的视点，并可能促进杂交稻产量的进一步提升。

粒重是重要的水稻产量构成因子，主要由籽粒大小决定，一般由数量性状位点(QTLs)控制。尽管目前已有很多调控粒重的QTL被鉴定克隆，但控制水稻籽粒大小的调控网络仍不十分清楚。在本研究中，我们图位克隆了一个主效的粒重QTL， Grain Length and Width 2 (GLW2)，发现其主要通过同时增加籽粒长度和宽度正向调节粒重。GLW2位点编码OsGRF4 (Growth-Regulating Factor 4)，在水稻体内受到microRNA miR396c的直接调控。OsGRF4中的突变扰乱了OsmiR396对OsGRF4靶标调控，从而使水稻粒型显著增加，产量提高。此外，我们还证明了OsGIF1 (GRF-interacting factors 1)能与OsGRF4发生直接相互作用，过量表达OsGIF1可以显著提高籽粒大小。上述结果表明，miR396c-OsGRF4-OsGIF1调控模块在调控水稻粒型方面起着重要作用，在水稻产量提升方面具有明确应用价值。