Development Of Rice SNP Marker Based On KASP Technology And Its Application In Breeding

The rapid development of rice functional genomics not only provides a powerful tool for rice genetic research,but also provides reliable information support for breeders to improve rice varieties through molecular marker orientation.Viable and efficient gene orientation improvement requires a thorough understanding of the genetic basis of target traits,development of DNA molecular markers,screening of molecular loci associated with target traits,and accurate design of breeding programs.However,the lack of a high density,high accuracy,easy detection and low cost molecular marker technology system hinders the practical application of molecular markers in breeding.With the deepening of the rice genome project,SNP(single nucleotide polymorphism)as the core of the new generation of molecular markers will greatly improve the efficiency of molecular marker breeding.In the SNP genotyping technique,KASP(competitive allele PC R)detection method supports low,medium and high throughput SNP detection,and has low cost,high accuracy and good application prospect.In this study,the international rice genome database and literature resources were retrieved,and the available SNPs were obtained.A total of 481 rice germplasms widely distributed at home and abroad were used as materials to design and develop the whole ge nome of rice based on KASP genotyping SNP labeling system.We conducted an association studybetween molecular markers and traits using the grain yield and quality of rice as the target traits.The main findings are as follows:1.It compares the Sanger sequencing and K ASP test results with Wx_Intron1_K,Badh2-E7,ALK and Pita markers to verify the accuracy of KASP test results.On the basis of this,596 loci were collected from the SNP database and tested for 565 pairs of primers.And 481 rice germplasms were detected using KASP with these primers.There were 558SNPs with fluorescence signals,of which 467 were polymorphic,which were transformed into KASP-SNP markers.The KASP-SNP marker has an avera ge marker density of 1.24SNP per Mb in the rice genome,and 244 markers associated with important traits(yield,quality,resistance,root development,plant development and fertility)have been reported.2.The genetic diversity of 481 rice germplasms was analyzed by K ASP-SNP markers.The results showed that the mean values of frequency,gene diversity,heterozygosity and polymorphism were 0.80,0.28,0.01,and 0.24,respectively.The 40 rare alleles(gene frequencies below 1%)were detected in 50 rice plants at 40 sites.C luster analysis and population structure showed that 481 rice germplasms could be divided into three subgroups.The genetic diversity and polymorphism information content of the small subgroups in the three subgroups were 0.19 to 0.50 and 0.16 to 0.37,respectively.And they had a high genetic diversity.Using the five functional markers of the K ASP-SNP marker system,97,18 and 3 genes with multiple germplasms were screened among the three subgroups.3.The length,width,aspect ratio,whiteness,thicknessof milled rice,100-milled rice weight,grain thickness,100-grain weight,the differential value of thickness and weight between grain and milled rice were investigated in 323 materials of 481 rice germplasms.The results showed that the ten agronomic traits were consistent with the nor mal distribution curve.And there was no significant correlation between the other traits and the traits except the grain length and the partial traits.It had a certain degree of LD for 14,325combinations between 413 markers.The average value of the D'and r~2was 0.076 and 0.438respectively.The LD attenuation distance was about 100kb in this population.O n the basis of this,37 markers were significantly associated with grain shape or weight by whole genome correlation analysis using general linear model GLM.In these markers,1 of which were significantly correlated with three traits,and 10 of which was significantly correlated with two traits.In this study,the whole genome KASP-SNP marker was first developed in rice,and the contribution of some SNP mark ers to grain traits was preliminarily clarified,which laid the foundation for further breeding of SNP molecular markers.