| Wheat (Triticum aestivum L.) is one of the main crop in the world, and its grain yield and quality are prone to be damaged by bacteria, fugus and virus etc. Wheat powdery mildew is a severe foliar disease caused by Blumeria graminis f. sp. Tritici (Bgt) in regions with temperate and maritime climates. Pm21, a resistant gene discoverd and utilized firstly by Chinese scientist, has broad-spectrum ability to resist powdery mildew. The gene located originally in shord arm of 6V in Haynaldia villosa (2n=2x=14, VV), and it was translocated into wheat. The gene is immune to all available isolates of Bgt in China.In this research, wheat cultivar'Bainong', which is susceptible to Bgt without any resistant genes against powdery mildew, and the near-isogenic line'W2132'with a resistance gene Pm21 were used. Differencial protein profiles were detected to determine the important proteins in resistance to powdery mildew by two dimensional electrophyresis (2DE) and MALDI-TOF-MS. To understand the resistant mechanism of Pm21 and obtain the important functional proteins or genes will be beneficial to the study in gene clone and transformation, and lay foundation for inhancing abilities in resistance to powdery mildew. And it has important significance in high grain yield and molecular breeding in wheat.Main results were listed as follow:1. Optimized the 2D technology of wheat leaves in our laboratory, and established the experiment systems for wheat proteomics: add 10%PVPP with milling wheat leaves; add one step to remove the ions, enhancing the voltages to 500V and enlonging the time to 4h; add 4% DTT in hydration; adjust the amount of APS (0.08%) and TEMED (0.04%), and add one step to vacuum the sample.2. Obtained the 2DE maps with well separation and many proteins. After 2DE and staining with CBB, detection with ImageMasterTM 2D Platinum software, about 400 spots were obtained in gels with pH3-10, and about 1100 spots were obtained in pH4-7.3. Differential proteins were determined by software. A total of 56 spots differentially expressed were studied. Sixteen proteins were identified by MALDI-TOF-MS in the two lines exhibited more than 1.5 fold differentially. Among the 13 determined proteins, spot 3 was detected as Rubisco activase and Spot 12 as Rubisco SSU, enhancing energy metabolism; Spot 1 was detected as oxidoreductase family protein and Spots 4 and 13 were belong to proline-rich family proteins, being related with defence; Spot 11 which was class II transposable element, may relate with gene; Spots 7 and 15 were belong to dehydrin proteins,making stabilization of proteins; Spot 14 was ribosomal protein L34,being related to stress response regulation.4. Determined one important proteins by MS, belong to Proline-rich Proteins (PRP). Besides the most proteins involoving in many physiological and biochemistry process, one PRP protein, is one main composition in cell wall and has important role in anti-stress in plants.5. Obvious difference in cell structure existed between the two materials. Mypha of the Bgt could not invade the powdery mildew resistant materials, and when the mypha invading the non-resistance materials, the chloroplasts were damaged; in addition, the cell wall of resistance materials were thicker than the non-resistance. The PRP, determined in MS, was highly expressed in resistant material. Therefore, the changing in cell wall structure might be one mechanism in powdery milder. |
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