Identification Of The RNAi Mutants Of Brachypodium Distachyon BdWRKY38,BdWRKY78 And Functional Analysis Of Their Homologous In The Interaction Of Wheat And Puccinia Striiformis F.sp.tritici

Wheat stripe rust is a widely distributed and fast-spread disease of wheat in global,which caused by Puccinia striiformis f.sp.tritici.Because of the frequent virulence variability of stripe rust fungus,resistant wheat cultivar in agriculture was not used for a long time,which increased the threateness to food security.In this way,illuminating the molecular mechanism of wheat resistance against rust fungus become very meaningful and important.However,how to quickly obtain key genes is still a great challenge to study functional genes based on the known genomic information.We utilized Brachypodium distachyon ecotype Bd21-3 of gramineous model plant for researching functional genes of wheat,which can accelerated the analysis of wheat resitance mechanism.Therefore,this paper aims to WRKY transcription factor,which playes an important role in plant defense,and tries to study the function of WRKY in the interaction between wheat-pst.Our research lay a foundation for revealing the mechanism of wheat resistance to wheat stripe rust.The main study results were listed as follows:(1)We detected positive plants of BdWRKY38 and BdWRKY78 of the candidate T4 RNAi mutants.We irrigated root with 20 μM/L dexamethasone to induce gene silencing and inoculated Puccinia brachypodii H-Ki.We found the hyphal length and colony area of RNAi plants in 24 h and 72 h significantly reduced compared to wild type;hydrogen peroxide of RNAi plants in 48 h area remarkably increased.According to the above results,we speculated that BdWRKY38 and BdWRKY78 play a negative regulatory role in the interaction of rust-infected Brachypodium distachyon.(2)Blast analysis showed that TaWRKY45 and TaWRKY40 were homologous with BdWRKY38 and BdWRKY78 respectively.TaWRKY45 and TaWRKY40 belong to WRKY group Ⅲ and WRKY group Ⅰ respectively.They contained the unique zinc finger structure and WRKY domain of the WRKY family;Yeast one-hybrid showed that TaWRKY40 had self-activation ability,but TaWRKY45 didn’t;The nicotiana benthamiana subcellular localization experiment showed that the two genes were both located in the nucleus.(3)TaWRKY40 was induced in the combination interaction of wheat-pst,and low abundance expression in the uncombination interaction of wheat-pst,but the change was not significant compared with the control group.The expression trend of TaWRKY45 was both similar in the combination and uncombination interaction of wheat-pst.After inoculating 48 h,the gene expression level was 1.2 times compared with the control group.(4)Transient silencing of TaWRKY40 and TaWRKY45 by VIGS technology,we found that in the compatible interaction,spores in silent plants leave was reduced.We speculated TaWRKY40 and TaWRKY45 play as a negative regulator in the interaction of wheat-pst.