Biocontrol Rhizobium Sneb183 Regulates Soybean Isoflavone Metabolism Against Soybean Cyst Nematode

Soybean cyst nematode(SCN,Heterodera glycines)is one of the most devastating diseases to soybean production all over the world.In China,soybean cyst nematode disease is also becoming more severe,which causes heavy yield losses of soybean production about 20% to 30%,even out of production.Sneb183 was screened form soybean Rhizobium strains,which can induce soybean resistance to cysts nematode.Through iTRAQ we found the protein level of chalcone synthase was promoted by Sneb183.In order to verify the relationship between SCN resistance induced by Sneb183 and isoflavone metabolism in soybean,in this experiment,the changes of the expression of two key genes in soybean isoflavone biosynthesis pathway at protein level and transcription level were studied during the interaction between Sneb183 and soybean.Also the changes of different isoflavones content in soybean root system induced by Sneb183 were determined.The results were as follows:1.Analyses of the expression of the key enzyme of soybean isoflavone metabolism inoculated with Sneb183 showed: the susceptible Liaodou 15 was treated in four groups:inoculated with 10 mL Sneb183 fermentation for 2 d followed by inoculated with 2000 J2,2000 J2 and 10 mL Sneb183 fermentation alone and conbined,sterilized water as control.The activity of two key enzymes of soybean isoflavones metabolism was detected by ELISA.The results showed that within 48 h after Sneb183 inoculation,the contents of two key enzymes were higher than that in these two groups inoculated with nematode alone and inoculated with Sneb183 at 4 h.At 2 h and 4 h after inoculation,the content of PAL was higher than that of the other three groups,and at 24 h,it reached the maximum value of 80.42 ng/L,which was 1.44 times of that of the control,and the content of IFS at12 h and 48 h after inoculation was higher than that of the other three groups,and the content of IFS reached the maximum at 2 h after inoculation.It was 699.05 ng/L,1.29 times that of the blank control.The results show that Sneb183 can induce the expression of key enzymes of isoflavone synthesis pathway in soybean differently both time and degree.2.Regulation of the expression of two key enzyme genes(PAL,IFS)regulated by Sneb183 at transcription level: the relative expression of two key enzyme genes in soybean root after Sneb183 inoculation was measured by real-time quantitative PCR(Real-time PCR).The results showed that at 2h and 4 h after inoculation of Sneb183 alone,the expression of PAL and IFS genes in soybean root was significantly higher than that in the two groups inoculated with Sneb183 and nematodes respectively.Relative expression of PAL and IFS genes reached the maximum value at 4h,which was 3.11 times and 6.16 times of the control treatment respectively.Among them,the relative expression of the group only inoculated nematodes was significantly higher than only inoculated with Sneb183.3.Analyses of effect of Sneb183 inoculation on contents of isoflavone in soybean root system:Daidzein and Genistein in soybean roots were detected by HPLC method and the results showed that the content of Daidzein was highest in the Sneb183 treated for 35 days,which was 4.02 times of that in the control group,the lowest group.The content of Genistein in rhizobia treated with Sneb183 for 28 days was significantly higher than that in other three groups,and reached the maximum value at 28 days,which was 1.68 times of that of the control.This indicates that the rhizobia Sneb183 can induce the accumulation of Daidzein and Genistein in soybean roots after inoculation.