| The stem non-structural carbohydrates(NSC)are the main assimilate source for grain filling and yield formation of rice.Increasing stem NSC accumulation and translocation,and improving grain filling of inferior spikelets are important approaches to high grain yield potential in rice.There are significant genotypic differences in NSC accumulation and translocation,low NSC translocation of stems was observed in some varieties or under high nitrogen condition,which leading to poor grain filling.On the other hand,poor grain filling in inferior spikelets is commonly considered as a main limiting factor for high grain potential.Therefore,our study investigated the accumulation and translocation of stem NSC,grain filling processes of superior and inferior spikelets and their relationships with sucrose-starch conversion,and also investigated grain assimilate phloem unloading processes,with objectives to elucidate the underlying physiological and molecular mechanism for stem NSC accumulation and translocation,differences in grain filling between superior and inferior spikelets,and phloem unloading in rice,which provides theoretical basis for high-yield cultivation and breeding.Based on those purposes,the researches of this study are as follows:(1)gene expression and activity of leaf sucrose phosphate synthase and their relationships with plant assimilates accumulation and yield formation;(2)activities of enzymes involved in sucrose-starch conversion in stems and their relationships with stem NSC accumulation and translocation;(3)genotypic differences in characteristics of large and small vascular bundles of peduncle and their relationship with stem NSC translocation;(4)genotypic differences in stem NSC translocation and phloem unloading and its physiological and molecular mechanisms;(5)the differences in grain filling of superior and inferior spikelets and their physiological and molecular mechanisms.The main results are as follows:1.The results indicated that expressions of leaf SPS genes(OsSPS1,OsSPS2,OsSPS6,OsSPS8 and OsSPS11)decreased at different developmental stages from panicle initiation to heading date in three rice varieties,Liangyoupeijiu,Shanyou63,and Huanghuazhan.Changes in SPS activities were consistent with that of SPS genes expression.Low nitrogen(LN)treatment increased SPS gene expressions and SPS activities when compared to high nitrogen(HN)treatment.Correlation analysis showed that the gene expressions and activities of SPS were significantly and positively correlated with leaf NSC concentration;OsSPS1 expression and SPS activation were significantly and positively correlated with spikelets per panicle and grain yield;expressions of OsSPS2,OsSPS6 and OsSPS8 were significantly and positively correlated with grain filling percentage and grain weight.These results suggest that enhanced expressions and activities of leaf SPS were beneficial to NSC accumulation of plant and yield formation via calculated reductions in nitrogen application rate.2.Genotypic differences in stem NSC accumulation and translocation of rice were observed,which was influenced by the nitrogen application.LN increased stem starch and NSC concentration before heading,and enhanced the hydrolysis of starch and NSC translocation in stems after heading.LN treatment shortened active grain filling period and increased grain filling rate of superior grains.In addition,higher stem NSC concentration before heading and NSC translocation were observed in SY63 than those in LYPJ.During the period between panicle initiation and heading,the activities of adenosine diphosphate-glucose pyrophosphorylase(AGP),starch synthase(StS),and starch branching enzyme(SBE)involved in starch synthesis increased under the LN treatment and positively correlated with increases in stem NSC.During grain filling,the activities of enzymes involved in starch-to-sucrose conversion(?-amylase,?-amylase,and sucrose phosphate synthase)increased under the LN treatment and positively correlated with stem NSC remobilization.Overall,the investigated enzymes exhibited higher activities in SY63 than in LYP9.These results suggest that low nitrogen increases the activities of AGP,StS,SBE,?-amylase,?-amylase,and SPS,leading to increased accumulation and remobilization of stem starch and NSC in SY63.We conclude that calculated reductions in nitrogen application and the choice of an appropriate cultivar may improve rice grain yields via enhanced stem NSC accumulation and translocation.3.Genotypic differences in numbers and total cross areas of large and small vascular bundles were observed using 4 recombinant inbred lines,R46,R94,R118,and R232,derived from the combination of Zhenshan97 and Minghui63.No significant differences in average cross sectional area and the ratios of phloem area to cross sectional area of large vascular bundle and to small vascular bundle were observed in different rice lines.Nitrogen treatments had no influence on the numbers,average cross sectional areas,total cross sectional areas and total phloem areas of large and small vascular bundles.The number of vascular bundles was positive correlation with stem NSC translocation,grain filling percentage,1000-grain weight and grain weight,and small vascular bundle showed tight association with stem NSC translocation according to the correlation coefficient.Functional differences in phloem of large and small vascular bundles may contribute to their differences in contribution to yield formation.Therefore,varieties with more vascular bundles(especially small vascular bundles)and cultivation techniques for increasing vascular bundles should be considered for high stem NSC translocation and high grain yield.4.Three recombinant inbred lines,R91,R156 and R201,derived from the cross between Zhenshan 97 and Minghui 63 were used in this study.Significant differences in stem NSC translocation have been observed among R91,R156 and R201.The results indicated that grain filling percentage,harvest index,and grain yield were lower in R201 when compared with R91 and R156,and stem NSC translocation was lower in R201 as well.Source-flow-sink related traits were determined,the results showed that the three lines had similar performances in specific leaf weight,leaf area index,stem NSC concentration at heading,spikelets per panicle,panicle/m~2,remobilization capability of NSC in stems,sink capacity and activity,biomass,number and cross sectional area of small vascular bundles.Meanwhile,greater number and cross sectional area of large vascular bundles,and higher SPAD were observed in R201.These results suggest that source,flow,and sink were not the limiting factors for low stem NSC translocation and grain filling percentage in R201.However,stem and rachis NSC concentration at maturity were significant higher in R201 than those in R91 and R156.Based on those results,we concluded that low NSC translocation,grain filling and yield formation in R201 might attributed to poor unloading of filling grains.5.The results showed that the densities of plasmodesmata between various cell types in phloem of dorsal vascular bundle of grains in R201 were lower than that in R91 and R156.The simulation of phloem symplastic pathway using carboxyfluorescein diacetate(CFDA)dye showed that the symplastic unloading was poor in grains of R201.A combination of western blotting,immunohistochemical localization and gene expression assay showed that expressions of sucrose transporter(SUT)and cell wall invertase(CWI)protein was lower in R201 than those in R91 and R156,and lower activity of CWI was observed in R201.These results indicated that the apoplasmic unloading was lower in R201 than that in R91 and R156.In conclusion,phloem unloading barriers was the main cause for low stem NSC translocation and sequentially low grain filling percentage,harvest index and grain yield of R201.6.Significant difference in grain filling was observed between superior and inferior spikelets in LYPJ and SY63.Inferior spikelets had lower grain weight and grain filling percentage than that of superior spikelets,which attributed to low grain filling rate of inferior grains.Nitrogen treatments had no effect on grain weight and the grain filling percentage of superior and inferior spikelets.However,high nitrogen application extended the active grain filling period and reduced the grain filling rate of superior and inferior spikelets.Firstly,the cross sectional area,total vascular area and total phloem area were higher in rachis of superior spikelets than those in inferior spikelets.On the other hand,the densities of plasmodesmata between various cell types in phloem of dorsal vascular bundle,the fluorescence intensity of CFDA,gene and protein expressions of SUT and CWI,and CWI activity in superior spikelets were higher than those in inferior spikelets,suggesting that superior spikelets have higher phloem unloading.Additionally,genes related to grain size,grain weight,grain filling,sucrose-starch conversion,and abscisic acid synthesis in superior were higher in superior spikelets than those in inferior spikelets;moreover,genes involved in ethylene synthesis were higher in superior spikelets than those in inferior spikelets.In conclusion,the superior spikelets have the advantage in the transport of assimilates,phloem unloading,and sucrose-starch conversion,which attribute to the better grain filling in superior spikelets. |
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