Functional Analysis Of Pest-responsing Genes In Soybean

Insect herbivores are one of the major factors affecting soybean yield and quality.According to the difference of digital gene expression profiling(DGE)of Tianlong 2(a cultivated soybean that is susceptible to herbivores)and ED059(a wild soybean that is resistant to herbivores)in response to infestations by the cotton bollworm(Helicoverpa armigera),it was found that a large number of genes were specifically expressed in the leave of ED059 after being attacked by the H.armigera.In order to study the mechanism of response to pests in soybean,two genes,GsSQM1 and GmMYB010,which were significantly up-regulated in the DGE,were selected and studied accomplished by agrobacterium-mediated transformation in Arabidopsis thaliana.1.The GsSQM1 gene is predicted to be a squalene monooxygenase gene.It is known that squalene monooxygenase and its downstream oxidosqualene cyclase(OSC)are two key enzymes for the synthesis of phytosterols and triterpenes.In this study,GsSQM1 gene was overexpressed in Arabidopsis thaliana.The homozygous transgenic plants were inoculated by newly hatched larvaes of H.armigera.The results showed that the transgenic plants of GsSQM1 gene were significantly susceptible to H.armigera,which indicated that the metabolites produced by squalene monooxygenase could be beneficial to the growth and development of insects.In order to analyze the mechanism of the effects of phytosterols and triterpenoids on insects,four OSC genes(AtCAS1,AtLAS1,AtPEN3 and AtLUP5)were selected to conduct further studies.The AtCAS1 gene encodes a cycloartenol synthase and the AtLAS1 gene encodes a lanosterol synthase,both of them are key regulators in the plantsterol synthesis branch.The AtLUP5 gene encodes the lupeol synthase and the AtPEN3 gene encodes the pentacyclic triterpene synthase,both of them are enzymes catalyzing the synthesis of triterpenoid saponins.The homozygous transgenic Arabidopsis lines of the four OSC genes were inoculated by the newly hatched larvaes of H.armigera.The results showed that the AtCAS1 and AtLAS1 overexpressing lines were significantly susceptible to H.armigera,while the AtPEN3 and AtLUP5 overexpression lines showed no differences in resistance to H.armigera compared with the controls.The results suggest that overexpression of squalene monooxygenase gene(GsSQM1)might increase the synthesis of sterols and prompt the growth of H.armigera through the pathways of downstream genes,CAS1 and LAS1.2.GmMYB010 belongs to the R-R-type of the MYB-related family.In order to study its function,the function of GmMYB010 and AtMYB010 were studied based on sequence,subcellular localization,transcriptional activation activity and phenotype of transgenic Arabidopsis plants.The results show that,although GmMYB010 and AtMYB010 were orthologous genes,they had obviously different funcitons.The newly harvested larvae of H.armigera were used to analyze the resistance of homozygous GmMYB010 overexpressing transgenic lines.The results showed that the transgenic lines were significantly susceoptbile to H.armigera,indicating that GmMYB010 might be a negative regulators in response to pests in soybeans.