| Maize is one of the world's most valuable crops and genetic model organisms.Through the combination of traditional breeding methods and modern molecular biology technique,the yield of maize has achieved remarkable improvments in the past centuries.Under the rapid change of climate,shrinking arable land and growthing population,genetic improvement of maize still facing huge challenges.Expanding the available genetic variation gene pool and understanding the genetic regulation mechanism of maize cell life activities are the basis of modern maize genetic improvement.This study explored the complexity of maize genetic variation and genetic regulation from different perspectives by using different maize genetic materials and genomics sequencing data.Based on the genome resequencing data of all subspecies of Zea,this study constructed a genetic variation map of Zea and analyzed the evolutionary history of Zea,further explored the environmental adaptation mechanism of Zea.In addition,based on the transcriptome data of cultivated maize,this study also analyzed the molecular characteristics and formation mechanism of maize circular RNAs,and extended the transcriptional regulation ways.Finally,based on the maize single cell DNA methylation sequencing data,this study analyzed the methylation pattern of maize monospore,and further analyzed the reasons for the heterogeneity in gametes developmental cells.The main findings are: 1.Genomic variation and adaptive evolution of Zea genusIn order to expand the modern maize genetic improvement variation pool and explore the molecular mechanisms of different taxa of Zea genus while adapting to different environments,genome sequencing data of all wild taxa of Zea genus(teosinte)and cultivated maize were studied.By comparing the differences between 690 genomes of teosintes and maize,we identified over 65 M single nucleotide polymorphisms(SNPs)as well as dozens of novel inversions.Our analysis of maize and teosinte genomes revealed the divergence time and effective population size change under the evolutionary of Zea genus.It reveals evidence of selection within taxa displaying novel adaptations including perenniality and a high tolerance to waterlogging by studying the positive selection genes of different taxa of Zea genus.In addition,based on genome-wide selection analysis,1,387 candidate sites related to highland adaptation and 2,057 sites related to high latitude adaptation were identified,and indicated the key role of hormone pathway in highland adaptation.Finally,by conducting environment genome-wide association analysis on 34 soil properties under 8 depth and the living altitude,we complement the candidate regions in plant-soil interaction.36 quantitative trait control sites that interact with soil and one site directly related to altitude were found.In the altitude related site,Zm WOX11 was regarded as the candidate gene.Functional analysis of Zm WOX11 indicate it can regulate root growth by responding to the plasma levels in the soil,and further affecting the high-altitude adaptability of teosinte.This work not only provides a comprehensive genetic variation map of Zea genus,but also gains new understanding of Zea evolution and adaptation.The study also identified numbers of valuable adaptation-related variation sites by combing genome-wide selection and association analysis,revealing the importance of hormone pathways in the teosinte highland adaptation and provides a theoretical basis and experimental evidence for the cultivation of new maize varieties with environmental adaptability through the improvement of hormone pathway related genes and soil interaction factors.2.Identification of maize circular RNAs and their function analysisIn order to deepen the understanding of maize functional genome and analyze the molecular mechanism of the maize circular RNA on phenotype variation,this study used the circular RNA specific and ?900 public access maize transcriptomic sequencing data to carry out large-scale identification and systematic analysis of maize circular RNA.By developing the bioinformatics analysis pipeline for genome-wide identification of circular RNAs,2,804 high-confidence maize circular RNAs were uncovered which show distinct genomic features.Further analyses demonstrated that sequences related to LINE/L1 elements(LLE)and their Reverse Complementary Pairs(LLERCPs)are significantly enriched in the flanking regions of circular RNAs.Interestingly,as the number of LLERCPs increase,the accumulation of circ RNAs varies,whereas that of linear transcripts decrease.Furthermore,genes with LLERCP-mediated circular RNAs are enriched among loci that associated with phenotypic variation.Finally,we showed that circ1690 was related to the variation of ear height.Our study indicates circular RNAs are widespread in maize genome and the biogenesis of maize circ RNAs are related to specific types of transposon.The study also uncovers a potential new way for transposons to modulate transcriptomic and phenotypic variations by mediating the production of circ RNAs.3.Establishment of the methylation map of single cell tetrad of maize and analysis of the heterogeneous sites generated by reprogrammingIn order to analyze the epigenetic changes during maize meiosis,this study sequenced the single cell microspore of maize tetrad based on a newly single-cell DNA metylation sequencing method in our laboratory.BRIF-MAP,a method to improve the utilization rate of single cell methylation sequenceing data was first developed.Based on BRIF-MAP,we uncovered the genome-wide DNA methylation landscape of maize tetrad microspore.Through the systematic analysis of the maize single-cell methylation map,we found the methylation rates among the four microspores in one maize tetrad were similar but significantly vairies among tetrads,indicating a obvious cell heterogeneity.By comparing the DNA methylation level between each single tetrad microspore and it's particular parent,we found the heterogenetity between tetrads were related to reprograming process during meiosis.Compared with the parents,averagely 1.33% methylation sites in the tetrad underwent reprogramming,among which 93.8% sites gained DNA methylation.The heterogeneous loci generated by reprogramming have certain preference on genomic components but remain similar under different genetic backgrounds.Through the utilization of population genetic data,this study not only identified large numbers of new genetic variations,but also expanded the available gene pool for modern maize improvement and provided a theoretical basis and experimental evidence for the improvement direction of adaptive maize new varieties.This study also used transcriptome and epigenome data to study the maize circular RNAs and single cell DNA methylation,further enhance our understanding of the regulatory complexity in maize cell activities. |
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