Genetic Analysis And Fine Mapping Of A Dwarf And Fragile Mutant Dwfl In Rice(Oryza Sativa L. Ssp. Indica)

Plant mechanical strength is one of the most important agronomic traits in rice, which reflects the physical properties of cell walls. Cell wall is the main form of biomass on the earth and it is the necessary fiber materials and raw bioenergy of human life. Fiber, one of the main contents of cell wall, is the most abundant biological macromolecules and the most important renewable resources. Therefore, the studying of the structure and biosynthesis regulation of cell walls has important theoretical significance and practical value. Rice is an important grain crop and also regaedes as one of the model plant of the grass family and monocotyledon. The research of brittle mutants and genes has great significance to demonstrate the regulation of grass family and monocotyledon, and is also useful for high yield, lodging resistance, resistance to diseases and insect pests and stress resistance.A dwarf and fragile mutant designated as dwfl was screened out in the progeny of an excellent indica restorer line Jinhui10with seeds treated by ethyl methane sulfonate (EMS). In this study, dwfl was identified, phenotypically and genetically analyzed, fine mapped and the candidate gene was cloned. The main results were as follows:1. Agronomic traits analysis and phenotypic identificationCompared with the wild type, the roots, stems, leaves and grains of dwfl were extremely brittle during its whole growth period. Agronomic traits such as plant height, panicle length, seed setting rate, internode length and1000-grain weight was lower at different degrees than those of the wild type.2. Mechanical strength analysis of mutant dwflThe tension test showed that tension stress and bending strength of dwfl decreased obviously compared with that of wild type. In addition, the fracture surface of dwfl was smooth. 3. Content analysis of cell wall components in dwflCompared with the wild type, cellulose and lignin content of the cell wall in dwfl decreased, while hemicellulose content increased sharply and silicate content had no significant difference.4. Cytological analysis of dwflAnatomical analysis of the second internodes and leaves showed that the stem surface of wild type was smoother but dwfl has a lot of serration. In wild type, the size and arrange of parenchymal cells were regular with two larger lumens in the middle and other two smaller lumens in two sides. However, in dwfl, the parenchymal cells were smaller, and even in the same level the size varies and arrange were inconsistent and disorganized, four lumens had the same size. Observed from the longitudinal section, the cells of wild type arranged regularly and became bigger step by step from outer to the inner. However, in dwfl, cells were irregular and arranged randomly, also its cell length shortened slightly.5. Genetic analysisReciprocal hybrid between Xinong1A and the dwfl was done for genetic analysis. The plant height and mechanical strength of F1progenies represented normal. In the F2progenies, the segregation ratio of normal plant (2347) to mutant plant (756) fitted the expected ratio of3:1(x2=1.67<x20.05、1=3.84), which suggested that the traits of dwf1were controlled by a single recessive nuclear gene.6. Fine mapping of gene DWF1Based on the BSA method, DWF1was mapped on chromosome9between maker Indl (autonomous synthetic) and RM201, with genetic distances3.35cM and4.34cM respectively. Within the region between InDell and RM201, we developed more polymorphic makers. Eventually, DWF1was located between Ind6and Ind4, and the numbers of recombinants were4and12, with genetic distances0.30cM and0.91cM, respectively.7. Analysis of candidate genesWe selected several genes that related to brittleness in the region for sequencing, such as cellulose synthase-CESA9, cellulose synthase family C-CSLC2, and xylanase inhibitors. The results showed that compared with Jinhui10, the2208th base of cellulose synthase-CESA9(LOC_Os09g25490) changed, and a single base missense mutation arised on the seventh extron in LOC_Os09g25490in dwfl, resulting in an amino acid change from cysteine to arginin because of the code change from TGC to GGC. LOC_Os09g25490was tentatively designated as the candidate gene of dwfl. Homologous comparison revealed that this mutation occurred in the highly conserved region of LOC_Os09g25490. These results would contribute to mutational mechanism research of rice dwarf and brittle mutants.