Study On The Function Of TaCDPK6 In The Interaction Between Wheat And Puccinia Triticina

Puccinia triticina is a highly specialized polysporous transparasitic fungus,which usually propagates in the form of summer spore generation,and is highly infectious and destructive to wheat.Wheat resistant to Puccinia triticinacan limit the infection by producing anaphylaxis in mesophyll cells.Some studies have shown that Ca2+signaling plays important roles in inducing HR production in wheat mesophyll cells.TaCDPKs are calcium-dependent protein kinases,as aspecial Ca2+signal decoding protein,can directly convert Ca2+signal into phosphorylation signal and transmit it to the downstream components,thus causing corresponding biochemical reactionsin wheat mesophyll cells.In this study,RNA-Seq data of wheat Thatcher and TcLr26 infected with Puccinia triticinaphysiological race 260 were analyzed.Based on transcriptome data,a gene Unigene19634_All was identified in wheat in response to leaf rust infection was screened in Ca2+signaling pathway,and combining NCBI BLAST sequence alignment to obtain the full length of the gene,which was named TaCDPK6.In this experiment,the function of TaCDPK6 gene during the interaction between wheat and leaf rust fungus was studied,and the experimental results were shown as follows:1.TaCDPK6 was screened by combining RNA-seq data and NCBI genomic data.Bioinformatics analysis found that CDS purpose gene sequence length is 1530 bp,encoding 509 amino acids,coding for 57 kDa protein relative molecular mass,with an isoelectric point of 7.61,and-0.419 total average hydrophilic for hydrophilic protein.From the N terminal to C terminal,there are variable domains,especially protein kinase domains,linkage domains and regulatory domains.The RNA-seq data analysis showed that the expression level of the gene in the incompatible combination was significantly higher than that in the compatible combination,and the expression pattern of the incompatible combination was roughly the same as that in the compatible combination.It was speculated that the gene may actively participate in the disease resistance activities of wheat.2.RT-qPCR and Western Blotting were used to detect the expression pattern of TaCDPK6 gene and its expression protein in the incompatible combination and the compatible combination during the interaction between wheat and Puccinia triticina.The results showed that in incompatible combination,with the extension of inoculation time,the purpose of gene and protein expression first rose;thereafter,they showed a downward trend,and were significantly higher than the compatible combination and reached the maximum in 4 to 8 h.The expression quantity and RNA-seq expression database changes were basically identical.3.The fusion expression of TaCDPK6 and GFP fluorescent protein in tobacco leaves showed that TaCDPK6 was localized in the cytoplasm,which was consistent with the analysis results of Cello V.2.5 subcellular localization prediction website.4.We used VIGS and RNAi technology on wheat varieties,TcLr26 silence TaCDPK6.We then inoculated Puccinia triticina physiological races of 260,and found out that compared with the control plants,TaCDPK6 gene silencing of the plant HR area is significantly reduced,and the number of HMC has significantly increased in TaCDPK6 silence.In the process of wheat resistance to Puccinia triticina infection,the ability of HR generation and resistance to Puccinia triticina was decreased,suggesting that TaCDPK6 functions to regulate base defense in wheat.