Genetic Basis Of Important Agronomic Traits And Gene Expression Changes During Modern Maize Breeding

Maize(Zea mays ssp.mays L.)is the largest crop in China.During the process of modern maize breeding,maize yield has increased by more than 7 times which was marked by the development of maize breeding technology in the first half of the 20 th century.Both cultivars with increased tolerance to high-density planting and maize architecture contributed significantly to the increased yield per unit land area.Systematically identifying the key agronomic traits and the associated genomic changes during modern maize breeding remains a significant challenge due to the complexity of genetic regulation and interaction of the various agronomic traits,and most of them are often controlled by numerous small-effect quantitative trait loci.Previous studies have investigated the genome-wide changes that occurred during maize breeding,but the mining of expression regulatory variation and key regulatory genes during modern maize breeding is not systematic and in-depth enough.Therefore,in order to better study the genetic basis of important agronomic traits during modern maize breeding and identify the key regulatory genes regulating phenotypic variation and favorable haplotypes,we collected a total of 137 inbred lines(ILs)representing multiple breeding periods in China(CN1960&70s,CN1980&90s,and CN2000&10s).The phenotypic data of important agronomic traits were collected,and the improvement rules of important agronomic traits during modern maize breeding were systematically analyzed.Transcriptome sequencing analysis of three distinct tissues/developmental stages(seedlings,leaves,and ears)to identify the expression patterns of expressed genes across the historical breeding eras.In combination with weighted gene co-expression network analysis(WGCNA),we identified the key regulatory genes regulating important traits and constructed the coexpression regulatory networks.By combining expression of quantitative trait loci(eQTL)and spliced quantitative trait loci(sQTL)association analysis,correlation coefficient analysis,and population genetics analysis,we identified the key candidate genes that might have been under selection and contributed to the genetic improvement of various agronomic traits during modern maize breeding.The main results are as follows:1.The statistical analysis of 9 yield-related traits phenotypic data based on 137 ILs showed that four traits(ear weight,hundred kernel weight,kernel length,and kernel volume weight)were significantly improved at different breeding periods in China.2.Gene cluster analysis based on three different tissues(seedlings,leaves,and ears)of 137 ILs were identified four groups of expressed genes with distinct trends of expression pattern change across the historical breeding eras(genes that exhibited continuously decreased expression across the breeding eras,genes that exhibited fluctuating expression across the breeding eras,with lower expression in era III than in era I,genes that exhibited continuously increased expression across the breeding eras,genes that exhibited fluctuating expression across the breeding eras,with higher expression in era III than in era I).Gene enrichment analysis revealed that the expression-changed genes in different tissues showed tissue specificity.These genes might play important roles in the development of seedlings,leaves,and ears during maize breeding improvement.3.WGCNA analyses based on 20 traits phenotypic data and expressed genes identified dozens of modules that were either positively or negatively correlated with various plant architecture-and yield-related traits.We also identified several new candidate regulatory genes potentially involved in maize inflorescence development,leaf angle and leaf development,stalk development and plant height,photomorphogenesis and flowering time,and ear development.4.eQTL analyses were performed based on the transcriptome data and structural variation maps of 137 maize inbred lines.We identified 12,920,10,624,and 7091 ciseQTLs corresponding to 8181,6919,and 6117 genes to were detected in seedlings,leaves,and ears,respectively.Overlapping analysis between the genes regulating ciseQTLs and the genes with continuously expression showed that 669,352,and 406 candidate genes regulated by cis-eQTLs for agronomic traits adapted to high-density planting were detected in seedlings,leaves,and ears,respectively.5.By combination with cis-eQTL analyses,WGCNA analysis,structural variation analysis of several complete maize genomes,population genetics,and molecular studies,we identified a set of candidate genes that might have been under selection and contributed to the genetic improvement of various agronomic traits during modern maize breeding,including a number of known key regulators of plant architecture,flowering time and yield-related traits.We also validated the functional variations in GL15,ZmPHYB2,and ZmPYL10 for the regulation of kernel row number,flowering time,plant height and ear height,respectively.6.Through the analysis of expressed transcripts of historical maize breeding eras,it was found that the proportion of alternative splicing of genes increased by about 20%during maize breeding.By combining cis-eQTL analyses identified 2056,511,and1473 cis-sQTLs corresponding to 1745,859,and 2381 genes in seedlings,leaf,and ear,respectively.Through analysis of cis-eQTLs and cis-sQTLs identified 8,162,and 504 SNPs both regulate gene expression and alternative splicing changes in seedlings,leaves,and ears,respectively.Functional variation classification analysis showed that cis-sQTLs were mainly enriched in the intron regions,while cis-eQTLs were mainly enriched in the intergenic regions,indicated that they were regulated by different ciselements.Overall,based on the phenotypic,re-sequencing,and transcriptome data of maize inbred lines during modern maize breeding in China,we analyzed the expression trend of expressed genes,constructed the regulatory network of key candidate genes regulating important agronomic traits,and identified the key candidate regulatory genes and functional variations regulating the improvement of important agronomic traits during modern maize breeding.This study provides important theoretical guidance and valuable genetic resources for the molecular design breeding of densitytolerant maize varieties.