Genome-wide Association Study And Candidate Gene Mining Of Major Agronomic Traits In Northeast Spring Wheat

It is always one of the important objectives of wheat breeding to improve the yield of wheat by improving the main agronomic characters and aggregating the excellent characters,but it is more and more difficult to improve wheat yield by traditional breeding methods.The genetic diversity of spring wheat is very rich in Northeast China,but the utilization of germplasm diversity is not enough.With the development of molecular marker technology,the use of Genome-wide association study（GWAS）to explore the favorable allelic variation of major agronomic traits has become one of the important means to improve the efficiency of molecular breeding.In order to excavate the significant association loci of the main agronomic traits of spring wheat and develop the linkage markers,improve breeding efficiency through molecular means,in this study,251 natural populations of spring wheat varieties were constructed and genotyped by 55K SNP chip in Northeast China,based on the phenotypic data,GWAS analysis was carried out to find the main trait association loci and candidate genes,and to develop closely linked molecular markers,which can provide reference for Marker-assisted selection（MAS）.Based on the characters,the investigated characters were divided into two parts,one was directly related to yield,named as yield and spike trait,the other was yield-related morphological characters,name it a morphological trait.The main results are as following:1.Statistical analysis showed that the distribution of yield,spike traits and morphological traits showed a normal distribution trend.ANOVA showed that all traits were significantly affected by genotype,environment,genotype and environment interaction.The correlation coefficient between yield and spike traits was significant,but the correlation between thousand-kernel weight and spikelet number、thousand-kernel weight and kernel number per spiker were not significant.The correlation coefficient between other traits were significant or extremely significant.Only the uppermost internode length was negatively correlated with the flag leaf width,the correlation between other traits was not significant.Population structure and principal component analysis showed that 251 spring wheat samples were divided into three subgroups according to their genotypes.The subgroups were ranked according to geography and breeding time,and there were significant phenotypic differences among the subgroups.2.There are 52503 high-quality SNPs were obtained by genotyping with 55K SNP chip.The results of genome-wide association analysis showed that there were 38 loci associated with yield and spike related traits.Among them,8 loci are significantly associated with SNU,6 loci are associated with SN,19 loci are most associated with SL,7 loci are associated with KNS,4loci are associated with TKW,13 loci are associated with grain yield,and each can explain7.6%-16.7%of phenotypic variations.Among these,10 loci were previously reported and 28 loci were possibly newly discovered.Based on these loci,69 haplotypes were obtained.Of these,there were 30 loci associated with morphological traits,9,7,7 and 7 loci were significantly associated with flag leaf length,flag leaf width,uppermost internode length and plant morphology,respectively,and each can explain 5.9%-17.8%of phenotypic variations.Among these,13 loci were coincident with those reported in previous studies,and the other 17 were newly found association loci.Based on these loci,68 haplotypes were obtained.3.For yield and spike associated loci,six candidate genes were identified,encoding cytokinin nucleoside,E3 ubiquity transferase,F-box protein,serine kinase and trehalose6-phosphatase,respectively.Seven candidate genes were identified by the morphological trait loci,encoding E3 obliquity transferase,cytokinins indican hydrogenase 5’-phosphoric acid,F-box family protein,leucine rich repeat receptor protein kinase and C₂H₂zinc finger protein,respectively.The functional verification of candidate genes was conducted by qRT-PCR analysis using the test materials with phenotypic differences.The qRT-PCR results indicated that all the13 candidate genes involved in the corresponding pathway and significantly difference among extreme accessions.4.For the main association loci,two KASP markers,K＿AX-109181055 and K＿AX-110090611,were successfully developed,which were associated with spikelet number and spike length,respectively.The validity of KASP marker was verified by using 78 northeast spring wheat lines.The results showed that 78 lines could be divided into three types:AA,GG and AG by marker K＿AX-109181055,and there were significant differences in the number of spikelets among the homozygous genotypes.The 78 lines could be divided into three types:AA,GG and AG by marker K＿AX-110090611.There were significant differences in spike length among homozygous genotypes.The above results shows that the KASP markers are effective and can be used to further marker-assisted selection（MAS）in breeding work.In summary,GWAS of spring wheat in Northeast China identified a number of loci associated with yield,spike traits and morphological traits,and materials abounding in favorable alleles in populations could be used to promote wheat breeding process.The association loci,candidate genes identified and KASP markers developed can effectively promote the progress of wheat MAS breeding.