The Role Of TaRanGAP2 In Wheat Resistance To Puccinia Triticina Infection

Puccinia triticina is a highly specialized living nutrient fungus.In the incompatible interaction between wheat and Puccinia triticina,the host produces a hypersensitive reaction(HR)to resist the infection.In the preliminary work of the laboratory,we proved that a NB-LRR protein TaNRH plays a very important role in the wheat resistance to Puccinia triticina,Then we confirmed that TaNRH can physically interact with TaRanGAP2 by yeast two-hybrid and BiFC technology.On the basis of,this study uses Co-IP technology to further detect whether TaNRH and TaRanGAP2 interact in the plants.The wheat near-isogenic line TcLr26 and physiological race 260 of Puccinia triticina were used to form incompatible combinations,and the recurrent parent Tatcher(TC)and physiological race 260 of Puccinia triticina were used to form compatible combinations.We aim to investigate the role of TaRanGAP2 gene in HR induction.These results will provide new insights into the function of TaNRH and will be of great significance for elucidating the mechanism of leaf rust resistance and breeding for disease resistance in wheat.The results of the experiment are as follows:1.The pCM1307-TaRanGAP2-HA vector was constructed and the pSuper1300-TaNRH-GFP vector constructed in the previous work was used,through Co-IP technology,it was found that TaNRH and TaRanGAP2 can interact in plants.2.The pSuper1300-TaRanGAP2-GFP vector was constructed.Compared with TaNRH alone,when TaRanGAP2 and TaNRH were co-expressed,in tobacco by transiently transforming,the induction of HR by TaNRH was enhanced3.By bioinformatics analysis,it was found that the total-length CDS of TaRanGAP2,is 1665bp,whcih encod 554 amino acids,TaRanGAP2protein contains a WPP domain and 4 LRR repeat sequences.4.Using RT-qPCR technology,the expression level of TaRanGAP2 gene at different times after inoculation were detected..The results showed that the expression of TaRanGAP2 was upregulated at 8 h and was the highest at 16 h after inoculation in the incompatible combination,after inoculation,while the expression of TaRanGAP2 in the compatible combination did not change significantly.5.The Agrobacterium containing pSuper1300-TaRanGAP2-GFP vector was transformed into tobacco transiently,it was observed with Confocal laser scanning microscope and showed that TaRanGAP2 protein was localized in the cytoplasm.6.Virus induced gene silencing(VIGS)technology were used to silence TaRanGAP2 gene in TcLr26.After inoculation physiological race 260 of Puccinia triticina,compared with the control group,the HR area of single infection point was significantly increased in TaRanGAP2 gene-silencing plants leaves,the number of HMC increased significantly,It is proved that TaRanGAP2 gene plays an important role in HR induction of wheat resistance to Puccinia triticina infection.