Functional Analysis Of Part Of The New Secreted Protein Family OG5₁54821 Members From Phytophthora Sojae And Gene ADA2 In Plant Immunity

Soybean root rot is one of the most disastrous diseases in the soybean production,part of this disease caused by Phytophthora sojae and is a typical soil-bone disease that can infect and kill soybean plants throughout all the growing season.The evaluated economic loses of annual soybean production caused by P.sojae alone is about 1-2 billion dollars around the world..sojae belongs to the Oomycetes which are classified in the Kingdom Protoctista and are related to heterokont,biflagaellate,folden-brown alage while fungi from a separate kingdom and are evolutionarily related animals.Bioinformatics analysis indentifyied almost 400 candidate RxLR effector genes in the genomes of P.sojae.The growing number of research results of effectors recover that they play the essential role during the pathogen infection and can use various methods to affect different plants defense layers.But many of the mechanisms about how these effectors disturb plant immune system are still unknown.Functional identification of the new secreted proteins family OG5₁54821 from P.sojae.There are many P.sojae strains’ whole genome sequencing have been completed,which will provide grate help us to find which proteins play the essential roles during the infection.Now there are many different families are under researching like RXLR,CRN and so on.In this part,I used the OrthoMCL to cluster of the P.sojae.Strain P6497 proteins.Comparing with the secreted protein database in my library to find the families which contain the secreted proteins.Next I screened the families with over 20 menbers which show high level transcription.Finally I chose the OG5₁54821 family as my target for further research.I found only the Ps₁28701 have the function to induce cell death by Overexpression in the Nicotiana benthamiana leaves.Then I used the crispr technology to knock out it from the P.sojae.Genome and picked out the transformants to testify their pathogenicity and growth rate.There are no difference comparing with the WT and CK.These results showed that though Ps₁28701 can induce cell death,it did not exert important function during the P.sojae Infection.P.sojae effector Avh23 can disturb the plant immunity by interacting with GmADA2-l and disturb plant immunity.In this part,I cloned the gene AtADA2 from Arabidopsis thaliana and found that it also can interact with PsAvh23 by using the yeast two hybridization,which recoverd the interaction between PsAvh23 and ADA2 was genereally exist in different plants.However the key area of ADA2 to interacting with PsAvh23 and the mechanism of how it exerts its function are still unclear.I designed different mutants of PsAvh23 and used the Y2H to testify the interaction between the mutants and GmADA2-1.I found the second structure of PsAvh23 protein had slight effect to their interaction.According to the bioinformatic analysis of its protein sequence,I found there are two internal repeated sequence（named IR1,IR2）,and they are the key areas of PsAvh23 for its interaction with GmADA2-1.In addition,I overexpress the GmADA2-1 in the tobacco leaves,and found it can suppress the infection of Phytophthora.capsici and silenced the ADA2 homologous of N.benthamiana can promote the infection of P.capsici.All these results shows that gene ADA2 plays as a positive regulator during the plant immunity process.