Functional Research Of Three Potato Ralstonia Solanacearum Effectors

Bacterial wilt caused by Ralstonia solanacearum is responsible for substantial losses in culivated potato(Solanum tuberosum)crops worldwide.Most bacterial pathogens manipulate plant immune response by secreting effector proteins into plant cells.Effectors is an important pathogenic factors of the Ralstonia solanacearum,so this project aims to explore the pathogenesis of potato bacterial wilt by studying the basic function of the effectors.In this study,microtuber of potato “ E3 ” was used as the receptor,the stable overexpression transgenic lines of Ralstonia solanacearum effector Rip5,Rip6 and Rip13 were obtained by agrobacterium mediated potato transformation system.In order to verify the influence of the effectors on the immune pathway,we further inoculated the transgenic lines.At the same time,transcriptome sequencing of Rip6 transgenic lines were carried out,and the differentially expressed genes related to plant resistance were found through data analysis.In order to further study the function of Rip5,the candidate interaction proteins of Rip5 were preliminarily screened by yeast two hybrid.The results of the yeast two hybrid were confirmed by Coimmunoprecipitation.Above results laid a good foundation for the further study of the effectors on the potato immune response mechanism.The main results are as follows:1,The transgenic lines of Rip5,Rip6 and Rip13 were obtained by agrobacterium mediated potato genetic transformation system.Finally,there are 9,13,and 30 of the transgenic lines of Rip5,Rip6 and Rip13 respectively.Through the positive test,we selected Rip5 transgenic lines 4,5 and 7,Rip6 transgenic lines 1,3,8 and 11 and Rip13 transgenic lines 11,14 and 25 for following experiments.2,Transgenic lines of Rip5,Rip6 and Rip13 were inoculated with UW551 strain.Compared with control E3,the inoculation phenotype of 3 effectors transgenic lines was significantly susceptible than control.3,Expression of PTI pathway marker genes in transgenic lines were detected.It was found that although the resistance of transgenic lines were decreased,the PTI pathway was not affected.It indicated that the introduction of effector genes may affected other immune pathways.4,We analyzed the transcriptome data of the Rip6 transgenic lines and found that there were a large number of calmodulins,some heat shock proteins and WRKY transcription factors.They were of differential expression compared to the control.It was speculated that the decrease of the resistance of transgenic lines was closely related to the expression of these genes.5,The candidate of interaction protein of Rip5 was preliminarily screened by yeast two hybridization experiment.Finally,the interaction proteins of Rip5 were verified by Coimmunoprecipitation.