Functional Analysis Of Phosphate Transporter GmPHT2 Family Members In Soybean

Soybean is an important grain and oil crop in China.Adequate phosphorus nutrition can significantly increase the dry matter accumulation and yield of soybean.However,there is less available phosphorus in the soil that can be used by plants.The application of phosphate fertilizer in the later period not only increased crop yields,but also caused environmental pollution and eutrophication of water bodies,and the phosphate rock,which is the main raw material of phosphate fertilizer,became scarce with the year-by-year mining.Therefore,it is very important to improve soybean’s utilization efficiency of phosphorus and increase soybean yield.The members of the plant phosphate transporter(PHT)family are responsible for absorbing phosphorus nutrients from the soil,redistributing and using them.The PHT2 family was found in higher plants such as tobacco and rice,and the growth of plants lacking the PHT2 family is significantly worse.Therefore,the study of soybean PHT2(Gm PHT2)family members is of great significance for further revealing the biological functions of Gm PHT2 and improving soybean phosphorus utilization.In this experiment,two Gm PHT2 family members were selected from soybeans,and made predictions through bioinformatics methods.Verification of bioinformatics results by subcellular localization and yeast complementation experiment.Promoter element analysis was performed on the promoter regions of the two genes,and soybeans were treated with different light,low phosphorus and 4℃.Then q PCR was used to detect the expression levels of the two genes.Construct an annulus and overexpression vector of the Gm PHT2 genes,and transform it into Arabidopsis to study the functions of the two genes.The specific research results are as follows:1.Gm PHT2;1 and Gm PHT2;2 were cloned from soybean.According to bioinformatics prediction.Gm PHT2;1 and Gm PHT2;2 proteins have the typical PHO4 superfamily domain of PHT2 family,and they may be phosphate transporters located on the chloroplast.2.Using the Arabidopsis protoplast transformation method,the subcellular localization test confirmed that Gm PHT2;1 and Gm PHT2;2 are located on the chloroplast.In the yeast replenishment test,both genes can replenish the phosphorus utilization ability of the defective yeast.3.The Gm PHT2;1 and Gm PHT2;2 genes have the ability to respond to low phosphorus stress,and the gene expression at a phosphorus concentration of 0.01 m M is significantly higher than the 0.5 m M level.Gm PHT2;1 and Gm PHT2;2 have a circadian rhythm in response to light.Compared with white light,gene expression changes under red and blue light conditions were consistent.4.After being stressed by low phosphorus,the expression of Gm PHT2;2 was significantly higher than that of Gm PHT2;1,showing the same trend throughout the growth period.After Gm PHT2;1 and Gm PHT2;2 were supplemented to the Arabidopsis mutant atpht2;1,the chloroplast and the whole plant phosphorus content of Gm PHT2;2 plants were significantly higher than those of Gm PHT2;1 plants.It shows that under low phosphorus stress,Gm PHT2;2may have a more important effect on plant phosphorus metabolism homeostasis than Gm PHT2;1.5.According to the analysis results of the promoter regions of Gm PHT2;1 and Gm PHT2;2,soybeans were treated at 4°C.It was found that the expression level of Gm PHT2;1 was up-regulated,and was significantly higher than the expression level of Gm PHT2;2 during the treatment period 3-12 h.It indicates that Gm PHT2;1 may also play a certain function in response to low temperature stress in addition to participating in the process of phosphate transport in soybeans.