Expression Analysis Of Chalcone Synthase GmCHS Gene And Isoflavone Content Changes Under Soybean Cyst Nematode Stress

Soybean Cyst Nematode(SCN)is one of the most serious diseases in soybean production in the world,causing billions of dollars in economic losses to soybean production every year.How to prevent SCNhas become the focus of research.With the continuous development of genetic engineering technology,screening resistance genes and breeding disease-resistant varieties are particularly important for the control of SCNand increasing soybean yield.Chalcone synthase(CHS)is a key site in the isoflavone formation pathway.As phytoalexins,isoflavones play an important stress-resistant role in plants under the stress of both biological and abiotic factors.The CHS gene has potential resistance to SCN.In this experiment,the resistance genes of Hui Pizhi heidou and susceptible variety Williams82 were used to detect the expression of four genes CHS5,CHS7,CHS8 and CHS9 at different periods after SCN3 stress,and determined the Danizen and Glycintein in the isoflavones component of its downstream product,as well as studied on the role of CHS gene in resistance to soybean varieties under the stress of SCN,in order to further reveal disease-resistant varieties the molecular mechanism of SCN lays the theoretical basis.The main results are as follows:1.According to bioinformatics analysis,CHS7 and CHS8 are composed of 389 amino acids,and CHS5 and CHS9 are composed of 388 amino acids.CHS5,CHS7,CHS8,and CHS9 are all stable proteins.Hydrophilicity and hydrophobicity predict that CHS7 and CHS8 genes are hydrophilic,and CHS5 and CHS9 are hydrophobic.DNAMAN analysis found that the primary structure of CHS protein in soybeans is basically the same,and CHS is more conservative in soybean varieties.For structural analysis of CHS5,CHS7,CHS8,and CHS9,random coil accounts for the largest proportion,followed by Alpha helix,and extended strand accounts for the least proportion.Subcellular localization predicts that CHS8 protein is located in the cytoplasm and chloroplast of eukaryotes,and that CHS5,CHS7 and CHS9 proteins are all located in the cytoplasm of eukaryotes.2.Acid fuchsin staining found that SCN3 had different infections in resistant and susceptible soybean varieties,and the number of nematodes in susceptible varieties wassignificantly higher than that of resistant varieties,indicating that there is a mechanism for resistant varieties to resist nematode infection.3.qRT-PCR was used to detect the expression of CHS5,CHS7,CHS8 and CHS9 genes of Williams82 and Hui Pizhi heidou under SCN3 stress.The results showed that the relative expression level of CHS gene in Hui Pizhi heidou was significantly higher than that of susceptible variety Williams82.In the disease-resistant variety Hui Pizhi heidou,4CHS genes were up-regulated at 5dpi(days post inoculation,dpi): CHS5 was up-regulated by 4.54 times,CHS7 was up-regulated by 1.99 times,CHS8 was up-regulated by 2.44 times,and CHS9 was up-regulated by 3.57.Times.4.Through protein interaction analysis,it was found that the co-expression of 4CL(4-coumarate-Co A ligase)and CHS speculates that phenylpropane metabolism is highly correlated with flavonoid biosynthesis.The co-expression of CHS-CHI-F3H-DFR in the interaction network speculates that flavonoids as related metabolites may be related to nematode infection and colonization.Compared with the transcriptome analysis of B.cinerea,CHS7 and CHS8 were up-regulated in nematode colonization and development period(5dpi and 10dpi),and downstream CHI2 A and CHI4 A were up-regulated in development and sex differentiation period(10dpi and 15dpi).5.High-performance liquid chromatography(HPLC)was used to detect the isoflavone content of Williams82 and Grifola frondosa under the stress of SCN3.The results showed that in six standard isoflavone varieties,only daidzein and glycintein were detected.Daidzein and glycintein were detected and expressed in large quantities in Hui Pizhi heidou at 1dpi after SCN3 stress.At the same time,1dpi-3dpi were higher than Wiliams82 content.Therefore,it was preliminarily speculated that the disease-resistant variety of Hui Pizhi heidou can quickly produce a stress effect after being stressed by soybean cyst nematode,and play a role in the resistance of soybean cyst nematode.