The Analysis Of Mo And Cd Accumulation In Rice Core Collection And Functional Verification Of Two Related Candidate Genes

As one of the most important crops of the world,the production of Rice(Oryza sativa L.)has important effects on the development of the global economy.The growth and development of rice requires the participation of various essential mineral elements,along with the absorption of various non essential mineral elements.The absorption level of those mineral elements directly impacts the yield and quality of rice.Therefore,studying the mechanism absorbing and accumulating mineral elements in rice would be helpful to cultivate green rice with high yield and good quality.Molybdenum(Mo)is an essential micronutrient for plants,serving as a cofactor for enzymes involved in nitrate assimilation,sulfite detoxification,abscisic acid biosynthesis,and purine degradation.Cadmium(Cd)is not a necessary mineral element for plants,and its accumulation forms a health risk for most organisms.In this study,the content of Mo and Cd in different stage,such as the straw of heading period,the grain and straw of mature period,of 533 cultivated rice core germplasm with wide genetic diversity was analyzed.Based on the study,the variation and distribution characteristics of these two elements in natural populations were preliminarily understood.In addition,two candidate genes named Os MOT1 and Os CDT1,respectively were found by GWAS and their roles in uptake or translocation of Mo and Cd were characterized.The results are summarized as follows:We analyzed the concentration of Mo in the germplasm materials,and found that the concentration of Mo in the straw of heading period and the grain of mature period are higher than that of the straw of mature period,suggesting that most of Mo are moved to the seeds in the mature period.Through GWAS scanning of the Mo concentration,a locus with significant repeatability in chromosome 8 was identified.Moreover the lead SNP of this locus is sf0800085089,and it was located in the promoter of Os MOT1.As we all known,At MOT1 is the molybdenum transporter in Arabidopsis thaliana,and Os MOT1 is the homologous protein of At MOT1,so we concluded that Os MOT1 is likely a candidate gene for this significant site.Because this site was found only in indica,especially in Ind I,we conclude that the function of Os MOT1 may mainly occur in Ind I.With bioinformatics tools and sequencing data analyzed all SNPs and INDEL information in the coding region and promoter region of the Os MOT1.A SNP closely linked with the lead SNP sf0800085089 and resulting in the change of an amino acid in the coding region was found.The SNP may the only mutation locus in the coding region of Os MOT1 in natural rice populations.The expression level analysis of Ind I in different materials indicated that Os MOT1 mainly expressed in roots,and the expression of Os MOT1 in high Mo materials is higher than that in low Mo material.Subcellular localization results showed that Os MOT1 encoded a protein localized in the cell membrane.In short,Os MOT1 is mainly expressed in roots and encode a protein localized in the cell membrane,and the variation in promoter affects the accumulation of Mo in Ind I.The concentration of Cd in the grain of mature period was found higher than the straw of heading period and mature period by analyzing the concentration of Cd in the germplasm materials.This suggests that a large number of Cd is limited to the seeds in the mature period.Additionally,the Cd concentration of different subsets was analyzed.The results showed that the accumulation of Cd in brown rice in indica is much higher than that in japonica.Through GWAS scanning of the Cd concentration,we identified a significant locus in chromosome 3,and the lead SNP is sf0325616618.On the other hand,Os CDT1,the most adjacent gene of the SNP,was found to encode a cadmium transporter,so we concluded that Os CDT1 is likely a candidate gene for this significant site.Because this site was only found in japonica,especially in Te J,we conclude that the function of Os CDT1 may mainly occur in Te J.We analyzed all SNP and INDEL information in the coding region and promoter region of the Os CDT1.The results showed that there is no multiple SNPs closely linked to the lead SNP sf0325616618 in the coding region,but it was found in the promoter region,indicating that the function of Os CDT1 is mainly in the promoter region.Under cadmium stress,the expression of Os CDT1 was inhibited in root,but it can be induced in shoot,indicating that the difference in accumulation of Cd is closely related to Os CDT1 in rice.Moreover,yeast functional complementation experiments also showed that Os CDT1 had cadmium transport activity.