Fine Mapping Of A Female-sterility Gene In Rice

Female sterility is a common phenomenon in higher plant.Rice(Oryza sativa L.)is a model plant of monocotyledons,and one of the most important food crop in the world.It will be helpful for us to understand of plant developmental biology and provide a theoretical basis for its application in breeding when deeply study of female-sterility in rice.As an important structural of rice,female gametophyte produces eggs and then develops into embryo after fertilization.Furthermore,it mediates a lot of reproductive process,including guiding the pollen tube development,fertilization and the seed development.Comparing with the numerous of male gametophyte,there is always one female gametophyte in a spikelet.In rice breeding,partially sterile pollen might not affect the seed rate,but even if partial sterile female gametophyte will directly affect the seed rate.With the development of science and technology in recent years,such as T-DNA insertion,transgene technology and chemical mutagenesis,many of scholars have isolated a series of genes affected the female gametophyte development in Arabidopsis.However,relatively fewer female gamete-related genes have been reported in rice due to the increasing difficulty of reproduction,hybridization and sampling of complete sterility type or low fertility type mutants,therefore,semi-sterility mutants are the ideal materiails for the clonging of female gametophyte relatived genes and revelation of the development mechanisms of female gametophyte in rice.In this study,we identified a rice semi-sterility mutant.We made a detailed cytological observation of the floral organs external phenotype and the internal structure of the ovary.Genetic mapping of the mutant gene was conducted by using a F2 mapping population of 93-11/mutant.The results established the foundation for further isolating the female-sterility gene and understanding the molecular mechanism of female gametophyte development.The main results were as follows:1.Based on the phenotype observation and cytological analyses,there were no obvious differences between the mutant and wild type including plant architecture,heading date and floral organ shape.At mature stage,however,the fertility of spikelet of the mutant was only 41.55±3.68%,it was significant reduced compared to wild type(95.87%±1.70%).Cytological observations showed part of mutant's pollens were abortive at early bicellular pollen stage,and the reason may be that the microspore failed to undergo mitosis to form reproductive nuclear.Further research on the germination of pollen on the sigma and its elongation,we found the quantity of pollens attached on the stigma of mutant were less than that of wild-type.However,the pollen tube of mutant could finally extend into the embryo sac as well.At the same time,we found that the setting rate of mutant still could not restore normal even the mutant was used as female paremale nt to be pollinated with the wild-type pollen,we speculated that the abortive embryo sac might lead to the low fertility of spikelet,so we used confocal laser scanning microscope(CLSM)to observe the mature embryo sac development of wild-type and mutant.The results indicated that 96.7%embryo sac of wild-type was normal,however,the normal embryo sacs of mutant were only 55.2%.Further studied of the embryo sac development process,the result indicated that the embryo sac of mutant was abortive at functional megaspore formation stage.In this stage,the megaspore could not develop into functional megaspore,finally resulted in the abortion of embryo sac.In summary,the abortion of embryo sac was the main reason for the sterile mutant.2.Genetic analysis showed that the semi-sterility trait of the mutant was controlled by one pair of recessive nuclear genes.Then,we conducted a F2 population by crossing the mutant with 93-11.From the F2 population,57 individuals which have the same fertility of spikelet as the mutant were selected to do linkage analysis,and the female-sterility gene was located between the SSR marker N9-27 and RM189 on the long arm of chromosome 9 preliminary.Via expanding the F2 population,we finally fine mapped the gene in the region between the InDel marker M-1 and SNP marker M-9.The physical distance between M-1 and M-9 is 257-kb,there are 47 ORFs(Open Reading Frames)in this region.The cloning of the female semi-sterility is underway.