| Soybean [Glycine max(L.)Merr.] originated in China,is an important food and oil crop in the world.Sulfur is an essential element for all organisms and the fourth largest nutrient element after nitrogen,phosphorus and potassium.In recent years,with the improvement of multiple cropping index and the large-scale planting of high-yield crops,the sulfur in soil has been largely consumed and cannot be supplemented.The phenomenon of soil sulfur deficiency in most parts of the world has become increasingly prominent.Low sulfur content in soil will lead to the normal physiological activities of plants blocked,metabolic disorders,reduce the tolerance of plants to stress and resistance to diseases and insect pests,and then adversely affect the yield and quality of crops.It is a feasible way to cultivate soybean varieties with high sulfur utilization efficiency or low sulfur tolerance.The main contents of this study are as follows:(1)165 soybean mutants were selected as the research objects to carry out the hydroponic experiment with low sulfur at seedling stage,investigate the phenotypic traits related to low sulfur tolerance,evaluate the low sulfur tolerance of each line based on phenotypic data,and screen low sulfur tolerant materials and low sulfur sensitive materials.Then,the phenotypic data were combined with SNP molecular markers for genome-wide association analysis to obtain the associated sites or block of low sulfur tolerance,and on this basis,the candidate genes were further screened.(2)Based on previous transcriptome data,a low sulfur tolerance candidate gene Gm LST1 was screened,and its function was preliminarily verified by hairy root transient transformation system to further analyze the role of Gm LST1 in soybean low sulfur tolerance.The main results are as follows:1.10 traits of 165 soybean mutants were measured under three environmental repetitions.The results showed that sulfur deficiency had significant effects on all phenotypic traits of the whole population.The chlorophyll of new leaf,chlorophyll of old leaf,plant height,fresh weight of shoot,fresh weight of root and dry weight of shoot were significantly decreased,while the root length,fresh root shoot ratio and dry root shoot ratio were significantly increased under sulfur deficiency.Heritability analysis showed that all traits had high heritability.The low sulfur tolerance parameters of each character basically showed normal or near normal distribution,which proved that the low sulfur tolerance of soybean was a quantitative character controlled by multiple genes.2.The mobility of sulfur in plants is poor,and the symptoms of sulfur deficiency are usually the earliest in young leaves.Therefore,SPAD(N)(SPAD value of new leaves)and DWS(dry weight of shoot)which have practical significance in production were selected as evaluation indexes to screen low sulfur tolerant resources.The-S/+S ratio of the above two traits was defined as the low sulfur tolerance parameter,and the low sulfur tolerant materials and low sulfur sensitive materials were selected according to the order of the low sulfur tolerance parameters.NJAU＿M061、NJAU＿M065、NJAU＿M099、NJAU＿M115、NJAU＿M128、NJAU＿M134 showed tolerance to low sulfur treatment in both indexes,and NJAU＿M041、NJAU＿M056、NJAU＿M101、NJAU＿M125 was sensitive to low sulfur treatment among the two traits,.GLM model in tassel software and Restricted Two-stage Multi-locus genome-wide association analysis(RTM-GWAS)were used to analyze all phenotypic data.GLM model analysis showed that multiple traits in different environments were associated with 98 significant loci,and the locus AX-93704456 significantly associated with SPAD(O)was repeatedly located in E1 and E2 environments.The results of RTM-GWAS showed that the number of association sites and blocks increased significantly,but only BLK＿GM07＿13083495＿13183280 was found in the same trait and multiple environments.According to the genetic attenuation distance of the mutant population and gene annotation,a number of candidate genes were screened out near AX-93704456 and BLK＿GM07＿13083495＿13183280.3.Based on previous transcriptional data,a candidate gene Gm LST1 related to low sulfur tolerance was screened,and its possible role in low sulfur stress was preliminarily identified by hairy root transient transformation system.We measured the chlorophyll content,plant height,fresh weigh of shoot and root,dry weight of shoot and root of Gm LST1 transgenic soybean chimera.In terms of plant height,compared with respective empty plasmid control,there was no significant difference between Gm LST1 overexpression and interference plants under +S condition,but the plant height of interference plants under-S condition was significantly higher than that of control.In terms of biomass,there was no significant difference between the chimeric lines under +S condition and their empty control,but under-S condition,the fresh weight and dry weight of hairy roots of the over expressed chimeric lines were significantly lower than those of the empty control lines.On the contrary,the four biomass indexes of the chimeric lines were significantly higher than those of the empty control lines,which indicated that the gene interference reduced the biomass loss of soybean under low sulfur stress.These results indicated that Gm LST1 negatively regulated the low sulfur tolerance of soybean. |
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