Study On Quality Change And Mechanism Of Tobacco Seeds During Development,storage And Priming

Tobacco seed is a special product in tobacco industry as it is the carrier of elite varieties.Generating high-quality seeds is the most cost-effective measure for increasing the production of tobacco leaf.This study aims to establish an integrated quality control system for seed production,processing,storage and pre-planting treatment of tobacco.Therefore,two varieties HonghuaDajinyuan(HD)and Yunyan 97(Y97)were analyzed for vigour,physiology and biochemistry as well as their molecular regulation in the process of maturation,storage and priming before sowing.Five major findings were achieved as follows.1.The relationship between internal substances and seed quality during seed maturation was examined and the optimal seed harvest time of HD and Y97 was determined.The viability and vigor of seeds reached their maximum at 31 and 33 days after pollination(DAP)for HD and Y97 respectively,and both declined at 35 DAP.The moisture content decreased sharply from 90%to 30%from 7 DAP to 35 DAP,and ABA content declined by more than 2000 ng g-1 in seeds for both HD and Y97.From 7 to 23 DAP,oil and protein accumulated in seeds,accompanied by the consumption of soluble sugar and starch.During the same period,crude fiber and arginine increased while ash and lysine decreased.In general,germination percentage in HD and Y97 was correlated with amount of oil,moisture,soluble sugar,protein,starch,ABA,lysine,arginine,seed weight and seed coat color.However,only moisture,oil and protein had a significant correlation with vigor index.Seed viability could be used to estimate the relationship between internal substances and seed quality and to predict the optimal seed harvest time to tobacco.2.A method of nondestructive determination by near infrared spectroscopy(NIRS)was established to estimate content of seed oil,sugar and crude figure during seed development.This method can facilitate making full use of tobacco seed biomass which are potential feedstock for biofuels.Four pre-processing methods were performed to optimize the original spectra before establishment of calibration models.Subsequently,two methods were utilized to develop the calibration models,and the LS-SVM model was found to have a better performance than the PLS model.The best LS-SVM models of oil,sugar,and crude fiber were established after pre-processed by MSC-SG-1st D.These results indicate that NIRS can be used to rapidly and accurately measure composition of tobacco seeds.3.The optimal seed packaging methods and storage conditions were identified and Ithe effect of lipoxygenase(LOX)on the deterioration of tobacco seeds was revealed.The preservation of seed viability and quality during storage is important for both commercial use and germplasm conservation.To reveal potential mechanisms of seed deterioration in tobacco,seeds that were packed by cloth bag(C)or vacuum bag(V)and stored under room temperature(RT)and low temperature(LT,18?)were sampled periodically for laboratory testing.Seeds stored in low temperature with vacuum bag(LT/V)exhibited the highest seed vigor after 25 months of storage,while seeds stored at room temperature with cloth bag(RT/C)had the lowest seed vigor and germination ability after 20 months'storage.Seeds stored in RT/C had increased content of endogenous hydrogen peroxide(H2O2)and malondialdehyde(MDA),lipoxygenase(LOX)activity,and NtLOX3 expression.In addition,regression analysis indicated that LOX activity had a strong negative correlation with seed vigor.To determine the role of LOX in seed vigor,two inhibitors of LOX activity,caffeic acid and catechin,were applied to the tobacco seeds during pre-treatment followed by artificial accelerated aging.Seeds pre-treated with inhibitors exhibited lower levels of LOX activity,MDA and H2O2 contents as well as higher seed vigor and seed quality.These results indicate that LOX accelerates seed aging and suppression of LOX activity enhances seed vigor and viability in accelerated aging seeds.4.Priming conditions were established for HD and Y97,including time,temperature,and solution pH.Twenty combinations were applied according to the RSA design,and seed germination were measured for each treatment.In addition,seedling height,dry weight,fresh weight and vigor index were measured and analyzed by RSA and PCA.The optimal priming conditions for HD were priming time 49 h,priming temperature 30? and pH 5;for Y97 are priming time 45 h,priming temperature 21? and pH 6,respectively.5.This potential contributions of reactive oxygen species(ROS)and gibberellin acid(GA)to seed germination were established.Diphenylene iodonium chloride(DPI)and uniconazole(Uni),biosynthesis inhibitors of H2O2 and GA respectively,were applied to tobacco seeds during seed priming.Priming with DPI or Uni decreased germination percentage as compared to priming with H2O;and priming with DPI+Uni further decreased the germination.H2O2 and GA completely reversed the inhibition induced by DPI and Uni.The germination percentage of H2O2+Uni or GA+DPI combinations was the same as the H2O treatment.Furthermore,GA or H2O2 increased GA content and decreased ABA content through regulating expression of genes involved in their homeostasis and signal transduction.Furthermore,GA or H2O2 induced the activation of storage reverse mobilization as well as enhancement of soluble sugar content and isocitrate lyase(ICL)activity.These results strongly suggest that H2O2 and GA are essential for seed gemination in tobacco and induce each other in regulating seed germination.Furthermore,LCL activity was regulated by both ABA/GA balance and ROS concentration.