Metabolic Profiling Analysis Of High Temperature Mediatedgene N For Immune Deficiency Of TMV

Tobacco infected tobacco mosaic virus is a serious disease which occurs in China and all over the world. The disease seriously affected the quantity and quality of tobaccos. The N gene-mediated HR is temperature dependent. At temperatures below 28°C, HR is elicited in the tobacco plant bearing the N gene upon infection with TMV. However, at temperatures higher than 28°C, the plants lose their ability to mount HR, and TMV spreads systemically. We use the methods of metabonomics to reveal the changes of metabolites before and after the inactivation of gene N at different temperatures and different infection time, which supplemented by molecular means to verify. And we carried out a preliminary study to eliciting resistance of two substances(malic acid and GABA).The results were as follows:1. This research studied the metabolic differences between TMV-free and TMV-infected Nicotiana tabacum var. Samsun NN 12 h、24 h and 48 h after infection at 25°C and 31°C respectively. We used gas chromatography-mass spectrometry and determined different metabolites with PCA and OPLS-DA. By using the GC-MS, 49 kinds of metabolites were detectable, containing amino acids, sugars, organic acids, organic acids, and polyamine. And we concluded, TMV infected N. tabacum var. Samsun NN would cause the raise of malic acid, salicylic acid, γ-aminobutyric acid, proline and ethanolamine at 25°C while N. tabacum var. Samsun NN would lose the function of gene-N with the derease of sucrose and inositol and the raise of glucose, fructose and amino acids at 31°C.2. We use quantitative PCR to analyse the relative levels of expression of important metabolic pathways related enzymes genes including malate dehydrogenase, glutamate decarboxylase and γ-aminobutyric acid aminotransferase at different temperatures in different infection period.The results show : malate dehydrogenas was the most significant increase at 25°C in 12 h infection while occured in 24 h at 31°C. And also the increase of glutamate dehydrogenase is higher at 25°C than 31°C. For γ-aminobutyric acid transferase, significant increase occured in 48 h infection at 25°C, while no significant change at 31°C.3. From the differenet metabolites, we chose γ- aminobutyric acid and malic acid to do the preliminary study of eliciting resistance. The results show : with the treatment of γ- aminobutyric acid and malic acid, the tobaccos infected with TMV revealed eliciting resistance. The most effective dose of γ- aminobutyric acid in eliciting resistance was 5 mmol/L. The average disease severities were more than 55.3% lowered. The most effective dose of malic acid in eliciting resistance was 100 mmol/L. The average disease severities were more than 56.3% lowered.