Researches On Changes Of Physiological Characteristics And Molecular Mechanisms Of Arabidopsis In Response To Methanol And Ethanol Stimulation

Recent studies have demonstrated that application of exogenous methanol can stimulate the growth of plants, carbon source hypothesis that methanol promote plant growth because of methanol as carbon source and assimilation of carbohydrate. In this laboratory studies,the of doctoral Wang Shasha indicate that methanol is oxidized to formaldehyde in Arabidopsis thaliana,this have formaldehyde and subsequently oxidized to formic acid and then through the C1metabolism of serine and enters the Calvin cycle for carbohydrate assimilation, through the use of compounds that inhibit methanol metabolism of methanol metabolism and stimulate confirmed the growth of Arabidopsis thaliana have no correlation, methanol may serve as a signal molecule to promote the assimilation of CO2material by inducing expression of photosynthetic genes, thereby enhancing the growth and biomass accumulation in Arabidopsis thaliana. Ethanol is a kind of structure with methanol similar simple alcohols, ethanol application effect of stimulation of plant growth also confirmed a number of research. Through the study on containing sucrose and high concentration glucose on MS solid medium growth of Arabidopsis root applying low concentration methanol and ethanol, methanol and ethanol affect the expression and physiological characteristics of photosynthesis in Arabidopsis genes; through the comparison of the WT and FDH (formate dehydrogenase) expression and physiological characteristics of photosynthesis related genes mutants response of methanol and ethanol stimulation, further research on the mechanism of promoting plant growth of alcohols. Finally the application of Arabidopsis thaliana cDNA chip analysis and identification of Arabidopsis thaliana in methanol and ethanol response genes, and metabolic pathways in response to methanol and ethanol stimulation related genes involved in the regulation of physiological characteristics of Arabidopsis thaliana were analyzed at the transcriptional level, in order to clarify the mechanism of methanol and ethanol stimulation of Arabidopsis growth, provides the theory basis for the widespread use of alcohols.The main research results are as follows: Glucose is an important product of photosynthesis, and its content increases as a signal molecule inhibits the expression of genes related to photosynthesis and plant growth, to examine whether the methanol and ethanol has to ease the ability of high concentration glucose inhibit the growth and photosynthetic gene expression in Arabidopsis thaliana, Seedlings of WT Arabidopsis in solid MS containing sucrose and3%glucose medium, growth analysis after two weeks in the greenhouse were found in the culture medium containing glucose, Arabidopsis growth than in sucrose medium containing the difference. On the base of2mM methanol or ethanol were added to increase the fresh weight of relative growth rate of Arabidopsis thaliana root culture, that lower concentration of methanol or ethanol can alleviate the inhibitory effect of glucose on Arabidopsis growth and ethanol, the effect is good in methanol. Further analysis showed the presence of methanol or ethanol decreased the content and oxidation of a in Arabidopsis thaliana chlorophyll stress level, the results of RT-PCR analysis confirmed the presence of methanol or ethanol release high concentration glucose on most photosynthesis in Arabidopsis related genes such as chlorophyll a/b binding protein (CAB1/2, Lhb4), photosystem II and I related protein (Psbp/Psbo/Psao), CO2assimilation genes (RCA, Rubisco) inhibited transcription.Medium with gas and solid H13CHO WT and FDH in sucrose containing solid MS (FDH) mutant24h, H13CHO metabolic profiling analysis using13CNMR, the results found to accumulate a large amount of H13COOH mutant FDH, H13COOH mutant of metabolism was significantly inhibited, the ability to absorb formaldehyde reduced, and production [U-13C]Gluc and N-amino acid transport reduction, confirmed that the FDH mutant H13CHO assimilation capacity decline. Analysis of WT and FDH metabolism of13CH3OH spectrum, the accumulation of free13CH3OH in the FDH mutant was significantly higher than that of WT, but the generation of [U-13C]Gluc and N-amino acid transport is less than WT, confirmed the ability of FDH mutant13CH3OH worse than WT assimilation. Analysis of WT and FDH to add culture response of methanol or ethanol medium in MS sucrose solid, results show that methanol or ethanol stimulation of WT and FDH growth has the same effect, methanol or ethanol effects on anthocyanin and chlorophyll a/b content of WT and FDH mutants showed similar effects.13CNMR analysis confirmed that NaH13CO3metabolism of WT and FDH mutant spectra are also similar, indicating that FDH mutations do not affect the ability of the Arabidopsis NaH13CO3assimilation, methanol or ethanol to promote WT and FDH mutant NaH13CO3assimilation effect is the same. RT analysis showed that transcription of WT and FDH mutants photosynthesis related genes of similar response patterns in methanol or ethanol stimulation. These results demonstrate that reduce Arabidopsis methanol metabolism ability does not affect its to methanol or ethanol stimulation response, metabolism of alcohols and stimulating the growth ability of plant.In order to better understand the molecular mechanism of growth using methanol and ethanol stimulation of Arabidopsis thaliana, methanol or ethanol treatment after24h WT Arabidopsis transcriptome changes using the cDNA chip technology, results showed that there were2880genes significantly influenced by2mM methanol, methanol up-regulated genes and1953down-regulated genes were927, methanol a. The classification of the1011functions of known genes up-regulated expression results indicated that methanol, the largest proportion of signal transduction and transcription regulation related genes (28%), followed by energy and metabolism related genes (21%). Expression of2mM ethanol significantly affect4223genes, including genes were up-regulated and down-regulated genes were3019and1204. Clustering of1151functions of known ethanol upregulates the expression of gene, which is the greatest proportion of signal transduction and transcription regulation related genes (37%), followed by cell growth/structure and plant development related gene (14%). Path analysis results showed that the expression of methanol and ethanol were raised chlorophyll synthesis and chlorophyll a/b binding protein, photosynthetic light response and CO2assimilation pathway, light respiration pathway, the pentose phosphate pathway, three of citric acid cycle and oxidative phosphorylation pathways, nitrogen metabolism, ascorbic acid and glutathione synthesis pathway, gene, only by way of methanol regulation including metabolism of fructose and mannose pathway, Cys and Met metabolic pathway, alkaloid biosynthesis pathways, is only way to ethanol regulation of Ala/Asp/Glu metabolic pathway, Val/Leu/Ile pathway, RNA splicing pathway and the proteasome pathway.