Genetic Evolution Of CKX Genes And Their Association With Agronomic Traits In Maize

Cytokinin oxidase(CKX,cytokinin oxidase)plays an important role in many aspects of plant growth and development by degrading cytokinin hormone,which is one of the key factors regulating plant growth and development.CKX proteins are encoded by cytokinin oxidase genes(CKX)and are often found in plant genomes in the form of a multi-gene family.There are 13 CKX genes in maize genome,which are crucial for the morphogenesis of maize.However,little is known about the genetic evolution of CKX genes and how their genetic variation affect the agronomic traits of maize.In this study,32 teosintes,68 maize landraces,and 348 maize inbred lines were used as experimental materials.The CKX genes were resequenced by BGI Life Tech Co.,Ltd.using the target sequence capture technology on the NimbleGen platform.The sequence polymorphism of CKX genes in three populations and the patterns of evolution during the domestication process were analyzed.In addition,the data of 12 maize agronomic traits measured in three environments were used to mine the favorable alleles significantly associated with maize agronomic traits based on candidate gene association.The main results are as follows:1.10 members of the maize CKX gene family were successfully resequenced in 448 lines.After multiple sequence alignment,sequence variation analysis was performed.It was found that the nucleotide polymorphism of CKX genes in downstream,intron,and downstream was higher,while the CDS was relatively lower.During the domestication process,the nucleotide diversity in each region of CKX genes was decreased from teosintes,landraces to inbred lines which indicated that the selection pressure had impact on the full length of the CKX genes' sequence.The LD pattern analysis showed that CKX2,CKX3 and CKX9 may be significantly selected.2.Statistical analysis of 12 maize agronomic traits in 348 maize inbred lines showed maize inbred lines existed extensive phenotypic variation.The coefficient of variation of tassel branch number was the largest(43.81%),kernel thickness was the lowest(5.32%).Most traits were correlated with each other to some extent,and the correlation coefficient between ear weight and ear grain weight was the highest,reaching 0.57.Leaf angle,tassel branch number,crown root number were negatively correlated with most other traits or the correlation coefficient was relatively small.3.Candidate gene association analysis detected a total of 42 variants of 6 CKX genes were significantly associated with 8 traits.Variants in CKX4 had significant effects on stem diameter and leaf width,and variants in CKX6 on leaf width.Among them,CKX4_SNP-1423,CKX4_SNP-1434 were significantly associated with stem diameter and leaf width.There are 9 variants in CKX6 are associated with leaf width,of which 5 variants are in strong LD.The contribution,R2,of CXX6_InDel-1511 in the promoter region to leaf width reaches 8.63%.And CKX6_InDel1072,which shared a strong LD with CKX6_InDel-1511,caused a nonsynonymous mutation,and the contribution reaches 7.04%.4.In the three populations,the allele frequency of significant variants detected by association analysis were compared.It was found that the SNP-1423,SNP-1434 and SNP-1449 of CKX4 might be selected during the domestication process.Whereas 7 of the 9 significant variants in CKX6 were detected having artificial selection signal and the nonsynonymous mutation site CKX6_InDel1072 with strongly selection signal and may be a critical variant affecting the target phenotype.Results from this research will provide a better understanding about sequence variation and genetic evolution of the CKX genes during maize domestication.The identified favorable alleles can be used through molecular marker-assisted selection for germplasm improvement.