| Platycodon grandiflorum is a kind of homologous plant resources widely distributed in hilly areas in china.The artificial cultivation of Platycodon grandiflorum can obtain high economic benefit.Both artificial cultivation and natural resources need to breed in seed.The small seeds(The weight of 1000 seeds<lg 1000)of Platycodon grandiflorum were sown in 2cm soil layer usually.The effect of drought stress on seed germination and seedling stage was easily affected,causing production decline.The increase in production and the protection of wild resources is restricted.This paper studied the characteristics of seed germination and seedlings growth under drought stress.The changes of physiological and Photosynthetic Characteristics and so on were studied.Effects of exogenous NO on seeds Germination and seedling growth were investigated.We have Cloned the key genes in response signaling network for root elongation and lateral root development of Platycodon grandiflorum,and analysis of genes expression of seedling under drought stress and exogenous NO treatment.The method to improving seed seedling rate was looked for under drought.1.Some indexes such as germination ratio,germination vigor,germination index,vigour index on seed of platycodon,free proline content,soluble sugar content,activity of antioxidant enzyme,and the content of MDA in leaf,etc.were determined and compared by treating with PEG-6000,and the effects of drought stress on seed germination and seedling growth were studied.The result shows that initial germination time of seeds was delayed,germination ratio,germination vigor,germination index,vigour index on seed,radicle length,embryonic bud length,root/shoot ratio,and fresh weight of seedling were lower at drought stress than those at natural water.With increasing drought stress degree,those index of being mentioned significant also reduced.drought stress could more inhibit to the growth of radicle than the growth of embry onic bud.The content of free proline at 10%PEG-6000 stress was higher,and its at other PEG-6000 stress were lower that of the contrast.The content of soluble sugar was decreased by low content PEG-6000(≤5%)stress,and its were improved by high content PEG-6000(>5%)stress.To compare with the contrast,activity of SOD in leaf on seedling were observable increased at different drought stress degree,activity of POD were significant decreased,and activity of CAT was slim enhanced by low content PEG-6000 stress,whereas its were observable reduced by serious drought stress.and content of MDA were higher than that of the contrast.The research found that drought stress could inhibit platycodon seed germination and seedling growth to a certain extent.One of reasons on low seedling ratio of platycodon seed in field was that the growth to radicle was inhibited by drought stress.2.With PEG simulation method,the effects of the drought stress on stomatal morphology of the seedling leaf,chloroplast ultrastructure,photo synthetic characteristics,product distribution and accumulation were studied,to provide the basis for germplasm innovation and breeding varieties to resistant drought of Platycodon grandiflorum.Under normal water condition in leaf chloroplast of Platycodon grandiflorum were single-sided convex lens shaped or fusiform,clear outer membrane,grana layer along the longitudinal axis of the chloroplast stacking closely and neatly,matrix lamellae clear and orderly.The average contains 3.2 starch grains in the chloroplast.10%PEG after 7d treatment,Platycodon grandiflorum seedling leaf chloroplast envelope boundary blur,reducing the number of grana.After 20%PEG treatment,most of the chloroplast outer membrane dissolved,grana lamellae stacking structure disappeared,leaving only the fracture of the stroma lamellae remnants.With the increase of drought stress,the chlorophyll content decreased gradually,with chlorosis of leaf color and stomatal closure.The value of Fo,NPQ and QP increased respectively.The numerical values of Fm,Fv,Fs,Fv/Fm,Fm/Fo,Fv/Fo,Fv’/Fm’,ETR,ΦPSⅡ,Pn,the activity of RuBPCase and the total substance accumulation decreased respectively.The mass of root and the root/shoot ratio were first increased and then decreased.The change of Pn,CE and WUE showed double Peak curve during the day,the change curve of Gs and LUE showed an irregular "V" curve,and the change of Tr showed a single peak curve under drought stress during the day.The change of Ci showed an irregular "W" curve under 10%PEG stress,and "V" curve under 20%PEG stress with a phenomenon of midday depression appeared.3.Controlled experiments were conducted to investigate the effect of temperature,drought and their interaction on seeds germination and seedling growth of platycodon,to aim at better understanding how they adapt to the seasonally drought and cold in spring at seed germination stage.The results showed that seed germination was optimal at the fluctuating night/day temperature regime of 10/20℃ and constant temperature range from 15℃ to 25℃,with more than 75.3%of seeds germinating in distilled water.Seed germination was significantly inhibited and their initiatory time was delayed at 10℃ germination percentage was increase and then decreased with the increasing of drought stress.While both of germination percentage and vigor index was always declined at other temperature.Root length,shoot length and fresh weight were declined with the decreasing of temperature,and root was more sensitive than shoot of seedling to changes of temperature.The results of a two-way ANOVA showed that temperature,drought and their interaction significantly affected the seed germination and seedling growth.4.Some indexes such as germination starting time delay,germination rate and germination potential decreased seed in the Drought Stress.The germination time of seeds in 50μmol/LSNP solution was earlier one day than that of the control,and the germination rate was obviously accelerated,and the seed germination was obviously promoted.the germination rate,the germination potential was higher than CK2(15%PEG)increased by 35 and 37.3 percentage points,germination ability recovered to the level before the drought stress.NO synthesis inhibitor L-NAME and NaN3 had no significant effect on Seed Germination under drought stress,root length,bud length,root length and shoot length were slightly lower than that of CK2.Based on the treatment of 15%PEG+ 0.1 nM L-NAME or 0.1mM NaN3,50μmol/LSNP has a significant role in promoting seed and root elongation on Seed Germination.5.The ORF of Nial/Nia2 gene and PgAB14 gene was found through amplification cDNA from seedling by RT-PCR and was sequenced.The length of ORFs were 2742bp and 1242bp,respectively.The numbers of amino acid were 913 and 413 in the polypeptide chain.The relative molecular weight of protein were 102.456KD and 44.038KD,the theoretical isoelectric points were 6.26 and 5.43,respectively.PgNial/Nia2 proteins was hydrophobic and stable proteins,with 3 conserved domains,including metal ions,ligand binding and catalytic activity.PgAB14 protein has 4 transmembrane helices,with signal peptide composed of 29 amino acids.It is a hydrophilic and stable transmembrane protein.The expression of PgNial/Nia2 gene increased slightly,while the expression of PgAB 14 gene increased significantly under 15%PEG stress.The expression of PgNial/Nia2 gene and PgABI4 gene in seedlings increased significantly When exogenous NO was applied.The results indicated that PgNial/Nia2 gene and PgAB 14 gene play an important role in response to drought stress and NO.6.The optimum formulation of Platycodon grandiflorum’s seeds was 55%meadow soil,the 43%layer soil,the 0.5%carboxymethyl cellulose,the 1%hydrogen phosphate amine,the 0.5%water retaining agent,and the 50 mol/L SNP.The cracking time of pellet was 70.3s.The containing kernel ration in pellets was 87%,the single seed rate was about 68.3%,and the number of seeds in per pellet was 2.61.The seed germination rate decreased significantly,and germination potential decreased slightly under normal water.The seed germination rate and germination potential were respectively 84.7%and 68.7%under 15%PEG stress.They were significantly higher than the control CK2.The seeds of Platycodon grandiflorum should be planted deeply after granulation.The seedling rate was 72.7%when the sowing depth was 5cm,This was significantly higher than the other treatments.Seed pelleting is an effective measure to solve the difficulty of seedling formation in the field. |
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