Differential Display Of Wheat Powdery Mildew On Four Wheat Cultivars By Means Of Reverse Transcription-polymerase Chain Reaction

Wheat Powdery mildew caused by Blumeria graminis f.sp.tritic(Bgt)i is considered a major disease in the main wheat regions all over the world. The main strategy for management of Bgt is resistant cultivars. But, due to pathogen variance, the resistance of new cultivars was lost soon. It is a serious threat to the yield and quality of wheat. Therefore, the study of pathogen variability of Bgt is of great significance. In this study, Differential Display reverse transcription PCR technology was utilized to discriminate Bgt inoculated on different wheat cultivars as well as looking for different expressed gene.Bgt was purified and inoculated on four different wheat cultivars (Chuannong26, Chuanyu20, Xike2, Neimail2), which contain similar resistance gene, we carried out the cytological observations using wheat cultivars Chuannong26, Chuanyu20, Xike2, Neimail2. After artificial inoculation with the prevalent Bgt race, we observed the length changes of germ tube of Bgt and papillae formed. The length of germ tube was longer in Neimai12, and Papillae formed at a similar time on four cultivars.After 10 days in incubator, four samples of Bgt conidiums were collected. RNA was extracted and reverse transcription was finished by Thermo scientific RevertAid First Strand cDNA Synthesis Kit.78 pairs primers were employed in Differential Display reverse transcription PCR. Results showed that 44 differential DNA fragments were obtained, four of them were sequenced and their functions were analysed using biological information method.B-1 DNA fragment, is from sample CY20,129bp in length. The molecular weight, isoelectric point, instability index of the putative protein were 5.131KD,11.543,58.54, respectively, indicating that the protein is unstable. Besides, the aliphatic index and average hydrophily were 54.42 and -1.342 respectively. While the on line PredictProtein predicted that the secondary structure composition of the protein is α-helix 16.28%, β-sheet 23.26% and β-turn 60.47%. Moreover, the grand average of hydropathicity of the putative protein was between -3.356 and 1.344.B-2 DNA fragment, is from sample XK2,674bp in length. The molecular weight, isoelectric point, instability index of the putative protein were 9.496KD,5.421,49.04, respectively, indicating that the protein is unstable. Besides, the aliphatic index and average hydrophily were 98.74 and -0.014 respectively. While the on line PredictProtein predicted that the secondary structure composition of the protein is α-helix 6.90%, β-sheet 22.99% and β-turn 70.11%. Moreover, the grand average of hydropathicity of the putative protein was between -1.765 and 2.322.B-3 DNA fragment, is from sample CN26,323bp in length. Its DNA sequence had similarity with diguanylate cyclase related genes. The molecular weight, isoelectric point, instability index of the putative protein were 10.661KD,9.032,29.01, respectively, indicating that the protein is stable. Besides, the aliphatic index and average hydrophily were 123.26 and 0.764 respectively. While the on line PredictProtein predicted that the secondary structure composition of the protein is α-helix 72.63%, β-sheet 4.21% and β-turn 23.16%. Moreover, the grand average of hydropathicity of the putative protein was between -3.044 and 3.333.B-4 DNA fragment, is from sample NM12,204bp in length. The molecular weight, isoelectric point, instability index of the putative protein were 6.387KD,9.18,75.63, respectively, indicating that the protein is unstable. Besides, the aliphatic index and average hydrophily were 62.68 and -1.125 respectively. While the on line PredictProtein predicted that the secondary structure composition of the protein is α-helix 0%, β-sheet 23.21% and β-turn 76.79%. Moreover, the grand average of hydropathicity of the putative protein was between 1.022 and -3.189.