Genome Wide Association Analysis Of Alkali Resistance In Soybean During Germination Stage

Soybean is the main source of vegetable oil and protein for mankind,and there is a huge demand for soybean in China.Soil salinization has become a global problem,affecting global plant growth.In order to increase soybean production in China,the utilization of alkaline land is one of the ways.Germination stage is one of the important stages of soybean growth and development,and it is also a relatively sensitive stage to alkali stress.Screening soybean resources with high alkali tolerance in germination stage can provide excellent resources for alkali tolerance soybean breeding.At the same time,mapping and marker development of target genes through genome wide association analysis is of great significance for promoting the process of soybean alkali tolerance breeding.In this study,350 soybean germplasm resources mainly from Northeast China were used as experimental materials.During the seed germination period,simulated alkali stress was conducted with a 20 mol/L Na2CO3 alkali solution.The germination experiment was conducted in a constant temperature incubator.The germination potential was measured on the third day of cultivation,and the germination rate was measured on the seventh day of cultivation.The alkali resistance level was evaluated based on the relative germination potential and the relative germination rate,Therefore,phenotypic comprehensive identification of alkali resistance was conducted on 350 soybean germplasm resources.At the same time,a 50 K beadchip was used to genotype 350 soybean accessions,and TASSEL5.2.60 was used for quality control of the genotype data and analyze the genetic relationship.Structure 2.3.4 was used for population structure analysis.GAPIT,a R-based software package was used for genome-wide association to detect SNPs significantly associated with soybean alkali tolerance.Gene annotations were searched through soybean genome data and candidate genes were preliminarily predicted.The main research results are as follows:According to the alkali damage index calculated based on the relative germination rate,there are 24 soybean resources with high alkali tolerance,63 with moderate tolerance,96 with moderate tolerance,92 with sensitivity,and 62 with high sensitivity.According to the alkali damage index calculated based on the relative germination potential,there are 43 soybean resources with high alkali tolerance,93 with moderate tolerance,80 with moderate tolerance,63 with sensitivity,and 58 with high sensitivity.2.The differences in alkali tolerance indicators ST-GP and ST-GR among 350 varieties are significant,consistent with normal distribution,and can be used for genome-wide association analysis.The relative germination potential is positively correlated with the relative germination rate,with a correlation coefficient of 0.683.There was no significant correlation between relative germination potential and relative germination percentage and agronomic traits and other stress resistance traits.The proportion of highly alkali resistant resources in bred varieties(lines)is higher than that of local varieties,while the proportion of alkali resistant resources in bred varieties(lines)is lower than that of local varieties.However,the t-test found that there was no significant difference in the relative germination potential and relative germination percentage between local and bred varieties and local varieties(P=0.363).Through comprehensive evaluation,six soybean varieties,including Gong79,Baimoshidou Jilin 12,Jilin Jinyuan 30,Honghuangdou,and Hefeng 14,were selected for alkali tolerance breeding due to their alkali tolerance resources.5.After quality control,a total of 30607 high-quality SNP sites were for subsequent analysis.The results of cluster analysis showed that 350 test materials could be divided into 3groups,and each group including 2 subgroups;The results of population structure analysis showed that 350 materials were divided into 6 subgroups,and the generated Q-value matrix could be used as a covariate for genome-wide association analysis.The LD decay distance of the soybean population was estimated as about 109 kb.6.At the threshold of-log10(P)=4.49,no SNPs significantly associated with relative germination potential were detected.Twenty-one SNP loci significantly associated with relative germination rate were detected,located on chromosomes 4,6,8,and 17,respectively.Among them,ss715628044 on chromosome 17 was the most significant,followed by ss715594644 on chromosome 6.7.Based on the LD attenuation distance,we searched for genes within the 218 kbp interval centered on ss715628044 and ss715594644,including 30 genes(Glyma.17g069100-Glyma.17g072000)and 32 genes(Glyma.06G281400-Glyma.06G284500),respectively.By searching for gene annotations and combining relevant literature reports,it is predicted that Glyma.17g070500 located on chromosome 17 is an important candidate gene,possibly related to alkaline tolerance in soybean.Glyma.06G283200 on chromosome 6 may also be a functionally related gene in response to alkali stress.