Association Analysis Of Starch Traits With SSR Markers Of Rice And The Identification Of Excellent Haplotypes In OsAGPL3 Gene

Rice is an annual grass crop that has a long history of cultivation and is one of the three most important food crops in the world.With the development of living standards,people are not only concerning with eating enough but also paying more attention to quality.The study of quality-related traits has become the focus of rice research in recent years.Milling quality,appearance quality,cooking and eating quality,nutritional quality are four main indicators for evaluating rice quality.Among all the indicators,cooking and eating quality is regarded as a very important and complex index that attracting a lot of geneticists and breeders to focus on.Amylose content(AAC),amylopectin content(APC),gel consistency(GC)and gelatinization temperature(GT)are the main factors that affecting rice cooking and eating quality.Therefore,to explore the variable regulation and to identify marker loci controlling relative traits in different rice accessions are important for improving rice cooking and eating quality.Based on this,154 pairs of SSR markers distributed on 12 chromosomes of rice evenly were used to do genome-wide scanning with the natural population constructed by 269 japonica rice varieties,and four starch-related traits including amylose content,amylopectin content,gel consistency and gelatinization temperature were measured to do association analysis.A wide range of representative 77 varieties were selected as materials to do association analysis between OsAGPL3 gene identified by SSR association analysis and starch-related traits.The followings are the results:(1)The starch-related traits of 269 japonica rice cultivars showed that the coefficient of variation of gelatinization temperature was relatively low,while the coefficient of variation of amylose content,amylopectin content,and gel consistency were relatively high.The variation tendency is as following: Gelatinization temperature <amylose content <amylopectin content <gel consistency.(2)Correlation analysis of starch-related traits showed that the amylose content had a negative correlation with amylopectin content significantly,a negative correlation with gel consistency significantly,and a positive correlation with gelatinization temperature significantly;The amylopectin content had a positive correlation with gel consistency significantly,a negative correlation with the gelatinization temperature significantly.The gel consistency had a negative correlation with gelatinization temperature significantly.(3)Software Structure 2.3.4 was used to analyze the population structure,269 materials were divided into 3 subgroups(P1,P2,P3)according to K=3.P1 contained 123 materials,P2 contained 74 materials,P3 contained 15 materials,and the remaining 57 materials were allocated into a mixed group.(4)The GLM model of software TASSEL 3.0 was used to do association analysis.A total of 43 SSR markers were identified to be associated with starch-related traits significantly,and the contribution rate ranged from 4.93% to 16.38%;The MLM model was also used to do the association analysis.A total of 29 markers were detected to be associated with starch-related traits significantly.The traits were significantly related,and the contribution rate ranged from 1.94% to 9.81%.The SSR marker RM19170 was associated with AAC,APC,CC,and GT significantly in GLM model,was associated with AAC and APC significantly in MLM model.Rice Genome Annotation Project website was used to search the position of RM19170,it was found that the position of the marker site overlapped with the OsAGPL3 gene.(5)Among the 269 rice materials,77 materials were selected with a wide representative,and gene OsAGPL3 in 77 samples were sequenced.Polymorphism analysis showed that 200 SNPs and 44 InDels were found in the coding and non-coding regions of OsAGPL3,and the distance of LD decay was 1260 bp(R2=0.1).Using Structure 2.2 software for population structure analysis,the 77 rice populations were divided into 3 sub-groups according to K=3.The first sub-group contained 44 materials,and the second sub-group contained 16 materials.There are 17 species in the three sub-groups.(6)Software Structure 2.3.4 was used to do population structure analysis,the 77 rice materials were divided into 3 subgroups according to K=3.The first subgroup contained 44 materials,the second subgroup contained 16 materials,and the third subgroup contained 17 materials.(7)The general linear model of software TASSEL 5.0 was used to do the association analysis,it was found that 11 polymorphic loci in OsAGPL3 were associated with starch-related traits significantly.Five polymorphic sites resulted in amino acid variations,and the tested materials were divided into 12 haplotypes.