Genetic Dissection Of Major QTL Controlling Grain Size In Oryza Rufipogon Griff

Grain size is one of the important agronomic traits that cause grain weight implies a significant effect on grain quality and yield in rice crop.However,the mechanisms underlying grain size control remain vague.To disclose the genetic mechanism of grain size and weight,we constructed an introgression Line BC₄F₆ population derived from a cross between a large grain ZH8015?a famous Chinese restorer line?and contrasting parent Oryza rufipogon GRIFF.Commonly known as?common wild rice CWR?,evaluated by two different methods such as marker-assisted selection and Genome-Wide Association analysis in two different environments at Hangzhou and Hainan for four seasons.We also performed RNA-Seq analysis of young panicle to detect grain size mechanism in a young panicle at 3 and 5 cm.In the first experiment,QTL mapping was conducted by using bulk sergeant analysis method to construct linkage map and performed QTL analysis for detection of Quantitative trait loci for grain size traits in Hainan and Hangzhou Environments.A bunch of six QTL existing in the adjacent of Pericentromeric region on chromosome 3 was identified tightly associated with grain size characters.Four QTL alleles qGL3.1,qLWR3.1,qTGW3,qTGW3.1 responsible for grain length,length-width ratio and thousand-grain weight derived from the large grain parent ZH8015 explained 80.91%,63.43%,24.40%,41.23%phenotypic variance respectively.Here qGL3.1 and qLWR3.1 displayed hierarchical effects on grain length,and length-width ratio resultantly increased in grain weight predicted that GS3,qGL3,and qGL3.2 might be the candidate region.While two QTL qLWR3 and qGW3.1a increasing grain weight within the population explained 12.91%and 20.16%derived from the small grain parent Oryza rufipogon Griff exist within the identified region of major QTL GW3,and GW3.1 might be the candidate region for QTL qLWR3 and qGW3.1a.In the second experiment,we performed Genome-Wide association Study GWAS analysis of grain shape related traits,such as grain length?GL?,grain length-width ratio,?LWR?1000-grain weight?TGW?and grain width?GW?to detect the candidate gene disturbing grain size traits within the population.We carried out an association analysis to identify QTL affecting rice grain shape and size.Genome-wide association analysis using28,193 SNPs Through general linear model?GLM?,a total of 56 significant SNPs on different loci associated with these four traits identified.The range of 100 kb upstream and downstream of the significant loci defined as a candidate interval,which found two known GS3 and qGL3 grain size-associated dominant QTL/genes including seven new loci with pleiotropic effects on phenotype affecting the grain shape and size characters.Also,Haplotype analysis,Gene expression analysis,DNA sequencing analysis,and combined gene-based association and functional annotation allowed us to shortlist two significant QTL Os03g0407400 and Os03g0646900 regions disturbing the grain and size.The most likely candidate gene?s?for each vital QTL also discussed.This research established a greater influence to pick out candidate genes affecting complex phenotypes by the strategy of combined GWAS,The identified candidate genes and loci provided valuable sources for future functional characterization and genetic improvement of rice grain shape and size.These findings can provide useful information for the understanding of the genetic control of grain shape and marker assistant selection MAS breeding in rice.After identification of candidate gene GS3 a major cloned QTL,we conduct our third experiment for characterization of GS3 transcriptional activity;we performed RNA sequencing using young panicle initiation?YPI?at 3 and 5 cm stages in the view of our previous study that GS3 highly expressed in 3 and 5 cm young panicle?YP?.We report comparative,transcriptomes profiling/analysis of YPI precise transcripts,their isoforms and regulatory mechanisms of biological,molecular,and cellular events.Comparison between transcriptomes of YPI at 3 and 5 cm of Niponbare?wild type?and induced mutant of GS3 exploiting RNA-Seq analysis generated 76.028 million cleaned high paired-end sequence reads it aligned with Nippnbare reference genome,a total of37,829 genes identified.During the transcriptomic investigation of YP,1303,and 1931,stage-specific genes/transcripts solely differentially expressed genes?DEGs?identified at both stages.Gene ontology analysis revealed 572 enriched DEGs in both YPI stages.Cluster analysis of differential genes displayed 20 similar gene expression profiles.We detect 653161 point mutations with 429226 transitions and 222379 transversions throughout the genome.Results revealed 10119 new variable splicing events.We identified three new genes with 21novel transcriptions.This research presents one of the massive transcriptome datasets together with related bioinformatics of rice young panicle initiation may serve as a blueprint for grain size regulation network.These findings magnify our knowledge for the complex biological,molecular,and cellular mechanism of grain size.