Omics Analysis Of Nitrogen Use Efficiency And Grain Filling Rate In Maize

As one of the most important grain crops and also a source of food,feed and fuel,maize(Zea mays L.)is cultivated broadly in the world.The maize yield has been the most important thing to ensure the national food security and the stability of the agriculture economic.Excessive nitrogen use is an important factor in the increase of corn production,but in the developed areas,people often use excessive nitrogen,this not only cause farmers' economic losses but also bring environment pollution;lack of nitrogen in the undeveloped areas will affect the yield.So,only the variety with high nitrogen use efficiency,can improve the nitrogen use efficiency of maize in low nitrogen situation and can also maintain the yield potential in high nitrogen condition.Huanghuai-hai region as one of the main areas of maize,it is full of rain and sunshine,but it has a tremendous impact on maize production for there are too much rainy day in the later part of maize growth.Previous studies have shown that if the maize grown under the adverse environment in the later part,there is no significant difference of the maize yield between the precocious and the long duration hybrids.So the maize with fast grain filling rate can make full use of light and heat resources while avoiding the adverse environment impact of late stage,has important practical significance of high and stable yield of maize.Based on this two problems,first,five maize materials with different nitrogen use efficiency were treated with normal nitrogen and low nitrogen stress,then we analyzed the changes of physiological traits and the related metabolic process;second,we studied on the dynamic changes of maize grain filling rate with three maize hybrids with different grain filling rate,from the point of proteomics and mi RNA transcriptomics,to explore the factors affecting the grain filling rate,we also did the function analysis of mi RNA159 c and its target gene who have the impact on plant growth and grain development.By analyzing the two aspects,we expect to find important regulatory factors who affecting the nitrogen use efficiency and grain filling rate of maize,and provide theoretical support for the low input and high yield of maize research.The main conclusions are as follows:1 The influence of nitrogen deficiency on tolerance mechanisms in seedlings of two maize hybrids(Xu178 × Huang-C and Xu178 × Zong3)and their parental inbred lines(Xu178,Huang-C and Zong3),which show different nitrogen use efficiency(NUE),was investigated using physiological measurements combined with global proteomics profiling.The root fresh weight and chlorophyll a/b ratio were reducedsignificantly in Huang-C(low NUE)under low nitrate treatment,whereas no significant change in these two traits was observed in Xu178(high NUE)under the same treatment compared with N-sufficient treatment.Fifty and 56 protein spots,which showed more than two-fold changes in abundance at P< 0.01 under low-N treatment compared with the control in the roots and leaves,respectively,were analyzed by protein mass spectrometry.Analysis of protein expression patterns revealed that proteins associated with carbohydrate metabolism,nucleotide metabolism,amino acid metabolism,disease/defense,and photosynthesis may be involved in N-deficiency responses.Low-N treatment led to an increased abundance of glutamine synthetasein the root to improve the efficiency of N assimilation in the inbred line with HNE,and affected photosynthetic carbon fixation and starch metabolism in the leaves and consequently seedling growth.2 The grain filling rate study among the Nongda108,Zhengdan958 and Pioneer335 showed that during the entire period of dry matter accumulation(10 to 50DAP),the three elite hybrids had significant differences in final grain dry weight and grain-filling rate.In addition,grain-filling rate dynamics was different for the three tested hybrids but with the same peak at 25 DAP.Zhengdan 958 showed a steep increase before 25 DAP and a more moderate decrease after 28 DAP in grain-filling rate,which was significantly distinguish from that of Nongda108 with distinct grain-filling rate from 25 to 40 DAP.These results implied that dry matter accumulation might mainly derive from rapid grain filling before 25 DAP and a relatively uniform decline after 25 DAP in Zhengdan958.Both Zhengdan958 and Pioneer335 have a higher grain filling rate than nongda108 in most of the grain filling stage.3 Endosperms and embryos of three elite maize hybrids(Zhengdan958,Nongda108,and Pioneer335)were sampled 17,22,25,and 28 days after pollination,for proteomic analysis to explore the regulatory factors critical for grain filling rate.In total,39 and43 protein spots that showed more than 2-fold changes in abundance at P<0.01 between any two sampling stages in the endosperm and embryo.Functional analysis revealed that metabolism was the largest category,represented by nine proteins in the endosperm and 12 proteins in the embryo,of the proteins that significantly changed in abundance.We found some important enzymes in glycolysis and TCA,they can supply energy for transporting the carbohydrate from photosynthesis organs to kernel.There are also some protein synthesis related proteins,and they may play key role in the transport protein synthesis and the accumulation of storage protein.The amount of most of these proteins changed with grain filling rate,and they are also the important reference factors of the grain filling rate.4 A deep-sequencing technique was used to research the dynamic expression patterns of mi RNAs at four distinct developmental grain filling stages in Zhengdan 958.After normalization,77 conserved mi RNAs and 74 novel mi RNAs were co-detected in these four samples.Eighty-one out of 162 targets of the conserved mi RNAs belonged to transcriptional regulation(81,50%).The result showed that mi RNA 156,393,396 and 397,with their respective targets,might play key roles in the grain filling rate by regulating maize growth,development and environment stress response.5 By comparing the results of proteomics and transcriptomics,it showd that mi RNA159 was close related to grain filling rate.We suppress the expression of mi RNA159 by the STTM method,and overexpress mi RNA159 by overexpress the Pre-mi RNA159,and we also suppress its target gene by the CRISPR method.In rice,the kernel length and thousand kernel weight of Pre159 and STTM159 are significantly higher and lower than that of the control,respectively.In Arabidopsis,the seeds of Pre159 and CRISPR-MYB33 are all larger than control.The expression amount of the mi RNA159 was increased in the Pre159 plant,and the MYB33 was decreased.About the STTM plant,the expression amount of mi RNA159 and MYB33 were decreased and increased respectively.MYB gene can suppress the synthesis and accumulation of the storage protein,in this paper,we mutant the MYB gene or suppress the MYB gene by over express the mi RNA159,to promote the the synthesis and accumulation of the storage protein and contribute to the grain filling.