| Wheat is an important food crops in China. Its yield is related to national food security. Therefore, High and stable yields of wheat play a key role in agricultural development. Wheat powdery mildew, caused by the fungal pathogen Blumeria graminis f. sp. tritici(Bgt), is an serious wheat disease. Wheat powdery mildew which could lead to serious yield losses and even no harvest has become the key factors restraining the yield and quality of wheat in China. The wheat powdery mildew fungus could mutate and become a new physiological race in a relatively short time and the cultivar resistance to powdery mildew, containing a single resistance gene, could decrease or even lose in the short term, therefore, finding and cloning the multiple genes which have resistance to Bgt are important for wheat breeding.In this study, genes were isolated from a high resistance to powdery mildew hybrid wheat alien disomic addition line germplasm SN6306, which originated from the intergeneric hybridization between wheat YN15 and Thinopyrum intermedium(Host) Nevski. The results are list as follows:(1)Functional Analysis of TiASP GeneIn this study, we established the mature embryo callus of YN15 as the receptor material, and the plasmid containing the TiASP gene and the pAHC20 plasmid containing the Bar gene were transformed into the mature embryo transformation system. According to the research on the method of callus induction, it was found that the percentage of the whole embryo induction was high, the quality of the callus was good, and it could make full use of the nutrients in the seed. In this study, we also studied the ratio of the differentiated medium to determine the hormone combination of 2.0 mg/L 6-BA+1.0 mg/L KT. In this study, we used the gene gun transformation to transform TiASP into the YN15 of the physiological races of E09, and the transgenic plants were obtained. The transgenic plants were obtained by PCR. The wheat powdery mildew infection of the transgenic plants and the control, observe two groups wheat leaves, after a week the results showed that the spots areas of transgenic plants is smaller than non transgenic control YN15. Transgenic plants with enhanced resistance, that TiASP may be a powdery mildew resistance gene in Thinopyrum intermedium.(2) Cloning and Functional Analysis of TaCPS1 Gene in WheatObtained in the early SN6306 after induced by Powdery Mildew in up-regulated expression results, after analysis obtained in the number 47969 unigene. Specific primers TaCPS1F/R were used to amplify target gene and get size 2500 bp band, obtained after sequencing length sequence of 2531 bp TaCPS1. In this study,the c DNA of TaCPS1 which was related to resistant to powdery mildew was identified from the wheat-Thinopyrum alien addition line SN6306 by m RNA differential display. The c DNA had the length of 2394 bp, which encoded 797 amino acid residues. Bioinformatics analysis indicated that the encoded protein which was consisted of the hydrophilic amino acids did not contain signal peptide. It was located in cytoplasm belonging to the Isoprenoid-Biosyn-C1 superfamily.(3) A Preliminary Analysis of the Function of TaCPS1 Gene in WheatRTFQ PCR analysis showed that the expression of TaCPS1 was up-regulated about 45 times than YN15 after they were induced by E09. Thus we hypothesized that TaCPS1 gene might be related to SN6306 resistance to wheat powdery mildew. The expression of TaCPS1 were increased nearly 15 times and 2.4 times under the induction of SA and Me JA respectively, which suggested that TaCPS1 may be involved in the regulation of SA and Me JA in the powdery mildew resistance mechanism. In this study, we constructed a VIGS system, which will contain a part of fragment of TaCPS1 that was connected to the p Ca-γb LIC vector, and successfully transformed into EHA105 agrobacterium, and pave the way for the preliminary analysis of the function of TaCPS1. |
