Tobacco Leaf Proteome Research Of Flue-cured Tobacco Producing Areas Of Sichuan

Tobacco (Nicotiana tabacum) is a widely adaptable economic crop. The Contribution of the taxation of China’s tobacco industry has been the top in all industries for the past decade. Although tobacco is wildly distributed between60degrees north and45degrees south latitudes, It’s a sensitive plant to environmental conditions. The yield and quality of flue-cured tobacco are under a greater influence of growing areas than their genotypes. The suitable ecological conditions played an important role in the production of high-quality tobacco leaves. Particular ecological conditions give tobaco unique characteristics. Hence, it is of practical significance to look for the causal factors of unique quality formation of flue-cured tobacco leaves from different producing ecotypes in Sichuan Province.In this study, the major cultivar "Yunyan87" were chosen as experimental materials. We established technique system of proteome of "Yunyan87" and identified the differential proteins of the flue-cured tobacco leaves among four typical ecotypes of Sichuan, including Yibin, Guangyuan, Xichang and Miyi, using the method of comparative proteomics. We focused on analyzing the proteins involved in the aroma substances related metabolic processes, such as starch synthesis, sugar metabolism, nitrogenous metabolism, terpene metabolism and carbohydrate synthesis. We aimed at finding out the key proteins which play important roles in the regulation and biosynthesis of the quality related substances of tobacco leaves. The main results were presented as follows:1. High quality proteins of tobacco leaves were extracted using BPP method, and satisfied to proteomic studies in the next step. High resolution and repeatability gel maps of1-DE and2-DE were obtained using GAP staining method. The identification of protein spots had a high success rate using mass spectrum.2. Comparing the2-DE maps of tobacco leaves of the four different ecotypes of Sichuan, we obtained119of differentially expressed protein spots with good separability.115out of the119spots were successfully identified using MALDI TOF and MALDI TOF/TOF mass spectrometry, and they represented91different kinds of proteins. These proteins were divided into12categories by COGs function classification, and photosynthesis, respiration metabolism, molecular chaperone proteins were the main parts. Subcellular location study using Targt P indicated that,44proteins were located to the chloroplast, one protein was located in the mitochondria, two proteins in secretion pathway,28proteins in other cell compartments, and the other40proteins localization remained unknown.3. By Blast2Go analysis, the retrieved50proteins of the identified differential proteins belonged to29kinds of enzymes, involved in30different metabolic pathways, including photosynthetic carbon fixation, glyoxylic acid and carboxylic acid metabolism, glycolysis/gluconeogenesis, methane metabolism, nitrogen metabolism, pentose phosphorylation, glutathione metabolism, oxidative phosphorylation and so on. Among them, many proteins and enzymes were involved in photosynthesis, respiration metabolism, glycolysis, nitrogen metabolism, and pentose phosphorylation cascades, indicating these metabolic pathways were more activity than others in tobaco leaves. In addition, We also found differential proteins involved in one terpenoids and four amino acid metabolic pathways.4. In this study, we identified some signal transduction related proteins, like14-3-3protein; photosynthese carbon assimilation associated RuBP proteins subunits; secondary metabolism associated Glyceric acid phosphate kinase; molecular chaperone family protein tcp-1cpn60, and so on. This work provided the reference for the further study of tobacco quality formation and the response mechanism to external environment changes.