| In view of the increase of unusual weather,and the quality and resistance of main planting flue-cured tobacco varieties were degraded in recent years,the study used Hongda,K326,Yunyan87,KRK26,Anyan2,Yunyan202,NC297,NC71,09-11,QL-3 and so on as experimental materials with different quality,fertilizer tolerance,resistance to high temperature and drought and bacterial wilt respectively,through the combination of field experiment,field pot experiment and hydroponic culture experiment,in Wenjiang District and Xingwen County of Sichuan Province in the past 2012-2017 years.Studied on the different grafting techniques,plant morphogenesis,roots morphogenesis,dry matter accumulation and operation and production quality,as well as fertilizer requirement characteristics changes of grafted flue-cured tobacco.We also studied the effects of grafted flue-cured tobacco on drought,high temperature stress,as well as the resistance and physiological mechanism to bacterial wilt.The main results are as follows:1.The effect and efficiency of grafted flue-cured tobacco were not only related to the compatibility of rootstocks and scions,but also affected by the grafting methods,the quality of grafted seedlings and the storage environment of grafted seedlings.The results showed that the survival rate of 1/2-1 leaf scion and the relative humidity of air about 85% after grafting were higher and the cost was lower.Choosing the suitable variety combination,and the tobacco plants with 7-leaf-1-core seedling age and a certain degree of lignification of stem were used as rootstock,and the tobacco plants with 6-leaf-1-core seedling age as scion.Then used PTFE TAPE winding the rootstocks and scions.2.The indexes of grafted flue-cured tobacco leaves were affected by rootstocks and Scions to a certain extent,and depended on the variety combination of scions and rootstocks,such as root and plant morphogenesis,dry matter accumulation and yield,chemical composition and sensory quality.Except for alkaloids and total nitrogen content,scion had greater influence than rootstock on the indexes that the topographic morphology and appearance of the plants,the main quality indexes,but rootstock had greater influence than scion on the growth of plant,dry weight of root system and vigor(bleeding)of rootstock.Compared with the self-rooted seedlings of scions and rootstocks,although the yield of grafted flue-cured tobacco has no obvious advantages,and even has a reduction in yield,the quality of tobacco leaves can be improved by choosing suitable scions and rootstock combinations,and the economic benefits can be improved.Among them,high-quality varieties KRK26 and K326 can be grafted to improve inferior the quality of Anyan2.3.The plant height,stem circumference,pitch and LAI were increased with fertilization.The self-grafted seedlings of fertilizer-sensitive variety KRK26 increased the most,the self-grafted seedlings of fertilizer-tolerant variety K326 increased the least,and the grafted seedlings were between them.The yield of grafted flue-cured tobacco was also between rootstock and scion,but closer to scion seedling,so selecting high-yielding varieties as scion was an effective way to improve the yield of flue-cured tobacco,fertilization could increase the yield of flue-cured tobacco,especially low-yielding and fertilizer-tolerant varieties;Sugar content and nicotine content were improved to improve the quality and increase the proportion of superior tobacco to a certain extent by fertilization.Compared with KRK26,the NR activity of K326 was lower,and the nitrogen,phosphorus pentoxide and potassium oxide uptake and accumulation and utilization rate were lower.NR activity,nitrogen,phosphorus pentoxide and potassium oxide uptake and utilization ratio of grafted flue-cured tobacco were between rootstock and scion,and more inclined to scion.Therefore,selecting varieties with high NR activity and fertilizer utilization ratio as scion for grafting could improve the fertilizer requirement characteristics of flue-cured tobacco to a certain extent,improve its ability of absorbing and utilizing nutrients,and provide varieties basis for reducing fertilizer application.4.Under drought stress,there was decreased that the root length,surface area,pitch,stem circumference,leaf area,dry matter accumulation per plant,and the more serious the drought,the greater the decline of the above indexes.The number of roots increased under mild drought,but decreased under severe drought,showing the same behavior at different growth stages,but the above morphological indexes and dry matter accumulation of different grafting materials were significantly different under drought stress.There was a certain interaction between grafting and drought treatment.The degree of drought stress effect was KRK26/Anyan2<KRK26/K326<KRK26/KRK26,which indicated that grafting the varieties with weak drought resistance into the varieties with strong drought resistance could improve their drought resistance to a certain extent.Drought stress affected physiological indexes of grafted Flue-cured tobacco,resulting in the decrease of SPAD value,the increase of Pro content in the leaves of rosette stage,vigorous stage and mature stage,and the increase of SOD,CAT and POD activities in the leaves of rosette stage,but the activity of these three enzymes decreased in the vigorous and mature stage.In general,the decrease of SPAD value and SOD,CAT and POD activity under drought stress was the smallest in KRK26/Anyan2,and the largest in KRK26/KRK26,especially under severe drought conditions,while the increase of Pro content was the opposite,which was an important physiological basis for the improvement of drought resistance of grafted flue-cured tobacco.5.The high temperature stress significantly affected the growth of flue-cured tobacco,resulting in shorter root length,lower surface area,fewer roots,shorter plant height,shorter pitch,shorter stem circumference,and lower LAI.However,there were significant differences among different materials.Except for individual growth stages,the above-mentioned growth indexes were Anyan2KRK26/Anyan2KRK26/KRK26 on the whole under high temperature stress.Grafting KRK26(as scion)with weak heat-tolerance into Anyan2(as rootstock)could improve its heat-tolerance to a certain extent.High temperature stress also affected physiological indexes of flue-cured tobacco leaves,reduced SPAD value and POD activity at different growth stages,increased CAT and SOD activity at rosette stage,and decreased CAT and SOD activity at vigorous and mature stages.In general,the SPAD value,POD,CAT and SOD activity of grafted flue-cured tobacco KRK26/Anyan2 were higher than those of KRK26/KRK26 or reduction lower than those of KRK26/KRK26 under high temperature stress,which was the physiological basis of improving heat tolerance of grafted flue-cured tobacco.High temperature stress increased the content of nicotine in rosette stage,but decreased the content of nicotine in mature stage and soluble sugar in different growth stages.The contents of soluble sugar and nicotine in different growth stages were Anyan2>KRK26/Anyan2>KRK26/KRK26.6.After inoculation with ralstonia solanacearum,the incidence and disease index of all treatments were Y87/Y87<Y87/HD<HD/Y87<HD/HD,and the activities of PAL,PPO,CAT and POD of the tested materials leaves were increased in varying degrees at 3,6 and 9 days after inoculation,and CAT was the most sensitive,which was the physiological mechanism of resistance to bacteria.The activities of PAL?PPO?CAT ad POD were Y87/Y87>Y87/HD> HD/Y87>HD/HD,contrary to their incidence and disease index,which indicated that the increase of these enzymes activities was the physiological basis for the enhancement of disease resistance of grafted flue-cured tobacco.Grafting with strong varieties can improve its disease resistance.Paraffin section analysis showed that the diameter ratio of vascular bundle to pulp and fusion layer to pulp of inoculation treatment was smaller than that of non-inoculation treatment,while the diameter ratio of cortical cells to pulp was larger than that of non-inoculation treatment.The extent of the damage to the grafted flue-cured tobacco cells that inoculation of bacterial wilt was generally HD/HD>HD/Y87>Y87/HD>Y87/Y87.7.The research results showed that tobacco grafted seedlings participated in the response of grafted seedlings to bacterial wilt through MAPK signaling pathway signal transduction pathway.Nitab4.5?0001961 g0030.1 encoding ribosome 40 s protein and transcription factors Nitab4.5?0002587g0020.1,Nitab4.5?0000005g0300.1,Nitab4.5?0001740g0070.1 and Nitab4.5?0008146g0050.1 encoding ribosome 40 s protein were found through the analysis of gene expression pattern and enrichment of MEtan module GO.The expression levels of these genes were Y87/Y87>Y87/HD>HD /Y87>HD/HD,which were similar to the rules of the incidence,disease index,PPO,CAT,POD and PAL activities of different grafting combinations.After Inoculated Ralstonia solanacearum,Nitab4.5?0007851g0010.1 encoding a Pathogenesis-related protein was first up-regulated and then down-regulated in resistant grafted seedlings Y87/Y87 and Y87/HD,but was inhibited after inoculation in sensitive grafted seedlings.TAZ-type Zinc finger Nitab4.5?0001812 g0040.1 was enriched in the aspect of temporal regulation,which was involved in the regulation of temporal and spatial expression of grafted seedlings.These results indicated that these genes were involved in the resistance of grafted flue-cured tobacco to bacterial wilt,which might be the molecular mechanism of grafted flue-cured tobacco to bacterial wilt.The Nitab4.5?0003324g0110.1 encoded Nitrogen regulatory protein P-II was highly expressed in HD/HD and low in Y87/Y87,but the Nitab4.5?0000363g0020.1encoded Nitrogen regulatory protein P-II was highly expressed in Y87/Y87 and low in HD/HD.The expression genes of HD/Y87 and Y87/HD were higher than that in the self-grafting combinations with low expression,indicating that the heterotropic combinations could combine the dominant characters of scions and rootstocks. |
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