Dissection Of Genetic Basis Of Tiller Angle And Flag Leaf Angle In Rice(Oryza Sativa)by Genome-Wide Association Study

As a major cereal crop,the grain yield of rice needs to be improved continuously with the increasing human population.Plant architecture is an important trait to determine the rice grain yield,and increasing the planting density is an effective way to increase yield.As two important agronomic traits for determining the rice ideal plant architecture,tiller angle and leaf angle have abundant natural variations in cultivated rice,so understanding their molecular mechanisms and exploring the favorable alleles will be helpful to improve rice varieties.Genome-wide association studies(GWAS)can quickly and efficiently identify loci closely associated with phenotypic variation using germplasms,which will be helpful to better understand the genetic basis of the complex agronomic traits.The rice research team of National Key Laboratory of Crop Genetic Improvement(NKLCGI)established a GWAS platform by sequencing a diverse global collection of 529 Oryza sativa accessions.In this study,we investigated the tiller angle and flag leaf angle at the heading stage of this germplasm collection,and then performed GWAS using linear regression and linear mixed model in the full population,and indica and japonica subpopulations,seperately.The major results are as follows:1.Although both tiller angle and flag leaf angle are important traits for plant architectrue and grain yield in rice,they did not show significant correlation according to the performance of 529 Asia cultivars.Rice tiller angle was dominantly controlled by genetic factor although it was also influenced by environment.The interaction between genotype and environment accounted for more partial of tiller angle variance in japonica subpopulation than in indica supopulation.The range of phenotypic variation on tiller angle of indica was wider than that of japonica.The average value of tiller angle in indica subpopulaitons was larger than that in japonica subpopulation.Accordingly,flag leaf angle was also mainly determined by genetic factor in rice,although environment and the interaction between genotype and environment influenced its performance.The interaction between genotype and environment played a relatively large role in the indica subpopulation.The flag leaf angle of cultivated rice showed a wide phenotypic distribution compared with tiller angle,and both the distribution and average value of the flag leaf angle of japonica were larger than those of indica.2.We detected a total of 30 QTLs controlling the natural variation of tiller angle of cultivated rice by GWAS,and 7 of the associations were identified both at Hainan and Wuhan.TAC1 located in q TA9 c region and q TA8 a were detected in the full population and indica subpopulation that were not influenced by the environment.While there was no locus commonly detected in japonica subpopulation in two environments;the rest 10 and 13 loci were detected only at Hainan and Wuhan,respectively.TAC3,the candidate gene of q TA3 on chromosome 3,was cloned by mutant analysis in this study.It encoded a conserved hypothetical protein and preferentially expressed in the tiller base.D2/CYPD90D2,known as a gene controlling rice leaf angle and plant height,was proved to be the candidate gene of q TA1 b by the mutant of D2 exhibited a decreased tiller angle.3.A nucleotide diversity analysis revealed TAC1,TAC3 and D2 had been subjected to selection during japonica domestication and genetic improvement.The genotypes at these three loci were fixed with favorable alleles in japonica subpopulation,but they were segregated with several genotypes in indica subpopulation.4.A total of 62 flag leaf angle-related associations were identified by GWAS,and only 8 of these loci were commonly detected both at Hainan and Wuhan.Four genes,reported controlling rice leaf angle in previous studies,were co-localized with the associated sites in our research.Four genes of b HLHs subfamily 16 were fallen into the QTL region of q FLA2 f,q FLA3 b and q FLA10 c.Then,three novel genes,Os153,Os173 and Os174,were confirmed to be leaf angle-related genes by overexpressing them in wild type rice.Transgenic plants overexpressing the orthologous gene in the subfamily Style2.1 of Solanum pennellii showed an increased leaf angle,too.We concluded that the members of b HLH subfamily 16 had a conserved function on controlling leaf angle in rice.5.We constructed the haplotypes of flag leaf angle-related genes detected in our study in two subpopulations,respectively,and compared significant difference on the flag leaf angle among haplotypes.We found there were significant differences on the flag leaf angle among haplotypes of each gene in japonica subpopulation,but there were no significant differences among haplotypes of every gene but Os BRI1 in indica.6.According to the associations detected for tiller angle and flag leaf angle,only the confidential region of flag leaf QTL,q FLA8 f,was partially overlapped with the confidential regions of tiller angle QTLs,q TA8 a and q TA8 b.In conclusion,tiller angle and flag leaf angle are two important plant architecture traits that affect the rice grain yield,but the genetic basis of tiller angle is very different from that of flag leaf angle;the variations of tiller angle and flag leaf angle are controlled by different genetic factors between indica and japonica subpopulations.It is the novel gene TAC3 together with TAC1,D2 and other newly identified genes that control the natural variation of tiller angle in cultivated rice.The members of b HLH subfamily 16 have the conserved function on regulating rice leaf angle.The favorable alleles identified in this study are good resources for breeding varieties with idea plant architecture.