The Study On The Regeneration Mechanism Of Dual-purpose Wheat XN136 At Seeding Stage After Mowing In Winter

With the improvement of people’s living standards,the demand for livestock products increase,which also requires the rapid development of livestock.However,the forage yield in north China still can’t meet the development of livestock in winter.Therefore,finding a reliable and high-quality forage is the key way to solve this problem,which has been beneficial to the development of livestock in China.Wheat is the most important food crop in northern China,and its area under cultivation is second only to maize and rice.In winter,wheat is at the tillering stage,which has abundant shoot biomass,and the crude fibre content in the stem and leaves is low,while the protein content is high,which is easily digestible by livestock.Wheat has been used as a high-quality forage grass in European and American countries.Since dual-purpose wheat could provide a large amount of fresh forage at the tillering stage and the yield does not decrease at the maturity stage,it could also be utilized in China.The use of dual-purpose wheat in China not only alleviates the shortage of winter forage grass,but also ensures food security.Regeneration ability is the key factor for the grain yield not decrease of dual-purpose wheat.After mowing,the cellular metabolism,physiological and biochemical metabolism and gene metabolism of wheat were significantly altered.However,most studies focused on the effects of the mowing frequency and the stubble height on wheat regeneration,but the regeneration mechanism of wheat after mowing is still unclear.In this study,for revealed the regeneration mechanism of wheat after mowing,the wheat variety"XN 136"was used as the experimental material,and the regeneration mechanism of wheat after mowing was preliminarily revealed by combining the cytological analysis,the physiology analysis and related gene expression pattern.The results of this study were shown as follows:1.Agronomic traits were investigated to determine the growth pattern of XN136 after mowing.The results showed that the growth pattern of wheat changed significantly after mowing.Under normal growth conditions,the growth of the wheat shoot almost stopped while the growth rate of the root accelerated.But after mowing,the shoot growth accelerated while the root growth was stunted.The results of paraffin section of tillering node（TN）showed that leaf formation rate and cell division rate in TN accelerated after mowing.The result of root paraffin section showed that the root diameter of secondary roots（SR）decreased and the root structure was disturbed after mowing.The submicrostructure showed that the cell activity of TN and SR increased at the early stage,and decreased at the late stage after mowing.Analysis of agronomic traits and cytological analysis indicated that mowing had a negative effect on the root growth,but could promote the shoot growth.2.The physiological and biochemical analysis of XN 136 showed that mowing of wheat resulted in physiological and biochemical changes in tillering node and secondary root.The results showed that the sugar content in TN and SR gradually decreased after mowing,while the sugar-related enzyme activities of sugar increased at the early stage.The expression changes of genes related to sugar metabolism were consistent with the enzyme activities.Root activity,H₂O₂content,MDA content,oxidoreductase activities increased significantly at the early stage after mowing and decreased at the later stage.The hormone content in TN and SR also changed significantly after mowing.Auxin content decreased in TN,and the content of cytokinin and gibberellin gradually increased after mowing,while these three hormones content decreased in SR.In addition,the contents of jasmonic acid,ethylene and abscisic acid increased significantly in TN and SR.qRT-PCR results showed that the expression of gene related to hormone metabolism was up-regulated.When wheat was treated with low concentrations of Me JA,ETH and ABA after mowing,which results in the wheat regeneration rate,leaf SPAD value and sugar content changed.The expression profiles of IAA,CTK and GA related genes also changed.3.Transcriptome analysis of XN136 showed that most of the DEGs in tillering node were mainly focused on biological process and molecular function after mowing,and the number of DEGs increased gradually after mowing.The DEGs which involved in PAL pathway were up-regulated,and the PAL and 4CL activities were significantly increased in TN and SR after mowing.The expressions of DEGs involved in pyruvate metabolism were also up-regulated after mowing.In the pyruvate metabolism pathway,the related amino acid metabolism genes were also up-regulated in TN after mowing.The DEGs which involved in sugar metabolism pathway were up-regulated,including starch hydrolase,sucrose synthetase,α-glucosidase and hexokinase,later results also showed that the activities ofβ-amylase andβ-glucose increased in TN and SR,indicating that the hydrolysis rate of sugar accelerated and provided energy for wheat regeneration.The DEGs in nitrogen metabolism pathways were up-regulated,and the activities of NR and GS in tiller nodes and roots were significantly increased.In amino acid metabolism,some DEGs were up-regulated after mowing.The DEGs in MDH metabolism pathway was up-regulated while in CS metabolism pathways were down-regulated.The MDH and CS enzyme activities were also significantly altered after mowing.In addition,the results of transcriptome also showed the KNOX and PAL genes were up-regulated in TN after mowing.4.The wheat genome contains 36 KNOX genes,which are unevenly distributed among the chromosomes.According to the expression pattern,the tissue expression profile of this gene family could be divided into two classes:class â…  and class â…¡.Class â…  genes were mainly expressed in stem apical and root apical meristems,whereas class â…¡ genes are expressed in all tissues.Class â…¡ genes were also up-regulated under many abiotic stresses.After mowing,the expression of some TaKNOX genes was up-regulated.When the wheat was treated with different hormones,TaKNOX gene was also altered.Functional validation in Arabidopsis indicated that overexpression of the wheat TaKNOX14-D gene in Arabidopsis could promote the leaf growth and improve the ability to recover from mechanical damage.Similarly,overexpression of the TaKNOX14-D gene in Arabidopsis could increase the cold tolerance.5.Totally 54 PAL genes were identified in wheat,and nine PAL genes were up-regulated after mowing in wheat according to qRT-PCR.In this study,the function of TaPAL50 was verified by transfer to Arabidopsis.The results showed that the drought and salt resistance of the transgenic lines were improved,and the transgenic lines could recover faster than WT after mechanical damage.In conclusion,the wheat growth pattern changes after mowing was due to the starch,soluble sugars and hormones in root was transferred into the shoot for wheat regeneration after mowing.In addition,in this processes,the hormone,starch and soluble sugar synthesized in the shoot were rapidly consumed,which accelerated the growth of the shoot after mowing.Meanwhile,,the enzymes and hormones synthesis related genes were up-regulated,which increased the activity the enzymes and the hormones content,ensuring the regeneration of XN136 after mowing in winter seedling stage.This study combined the analysis of the cytological,physiological,and biochemical and transcriptome in the wheat shoot and root after mowing,and the verification of the related gene functions,which provisionally revealed the regeneration pattern of wheat after mowing,and that would contribute to the development and improvement of the wheat regeneration capacity after mowing.