| Tomato (Solarium lycopersicum) is one of the most important economical vegetable crops in the Solanaceae family and also has been used as a model system. Drought stress straightly affects the growth and development of tomato seedlings, and causes of decreased quality and yield reduction. It has become one of urgent requirements to breed excellently drought-tolerant tomato variety to reduce the water deficit damage.The elucidation of the mechanisms of drought tolerance has been studied on the base of morphological and anatomical, physiological and biochemical, and molecular level. The plant adaptation to water deficit are caused by changed morphological and anatomical structure, such as thick waxy cuticle, altered stomata aperture, well developed root system, and strong parenchyma. The physiological and biochemical characteristics including the accumulation of osmotic adjustment materials and protective enzymes, the stability of membrane system, the presence of endogenous hormones and secondary metabolites also contribute to enhanced drought tolerance. During the process of molecular research on plant responses to drought stress, a large number of genes were cloned, classifying into two major groups according to their putative functional modes. The first group comprises the genes encoding structural proteins, which are stress tolerance downstream effectors. The second group genes encoded regulatory proteins that are early response transcriptional activators including the transcription factors and the protein kinases. The expression of drought-inducible genes can be governed by ABA-dependent or ABA-independent regulatory systems, forming the cross-talks to regulate the activity of transcription factors and signal transduction in response to drought stress.S. pennellii (LA0716), one of the wild crossable relatives of cultivated species, has a distinct ability to withstand desiccation in extreme arid condition. A collection of 50 introgression lines that covers the entire genome of their donor parent, S. pennellii, in the background of the cultivated species, S. lycopersicum M82, a drought-sensitive cultivar, were generated. In this study, we firstly use the drought-tolernat lines together with the currently expanding tomato research platforms such as genome sequencing and microarray applications to study the chacteraization of drought responsive genes and investigate the drought tolerance mechanism in tomato. The main results were as following:1. The structure characteristics analysis of S. pennellii. A comparative observation of morphological and anatomical structure between S. pennellii and M82 were performed to study the drought-tolerant structure characteristics. The aerial tissues including leaf, stem, calyx and fruit of S. pennellii plant are covered with dense trichomes and glandular secretion, which may be benefit in reducing of water loss and absorbing moisture in the air. The increased number of stomata on the upper epidermis of S. pennellii leaf might support the channel for adsorbed dew. The thickness ratio of stockade tissue to sponge tissue in S. pennellii leaf is higher than that in M82, which could reinfoce the mechanical strength under drought stress. The parenchyma cells are widely distributed throughout stem of S. pennellii, suggesting they are benefit for water store and stronger water retention capacity, while the root and leaf vein are weak, indicating these tissues play unimportant roles in drought tolerance.2. An analysis on index system of tomato drought tolerance evaluation. The correlation of index/index and index/drought tolerance under different drought stresses were studied. The relative water content can be used for selecting the drought-tolerant tomato genotype under mild, moderate and serve drought stress. The malondialdehyde content is also effective index for detecting the tomato drought tolerance under moderate drought stress.3. The expression characteristics of drought responsive genes in tomato. Gene expression profiles between two drought-tolerant lines identified from introgression line population of S. pennellii and M82 were investigated under drought stress using tomato Oligo chips TOM2. More than 1,200 drought-responsive genes were identified in each of tested genotypes. The "response to stress" and "response to abiotic stimulus" were among the largest groups in all three genotypes. Based on biological function of drought-responsive genes, the enhanced drought tolerance of tomato may be closely related to upstream regulation of transcription factors and signal regulators specifically identified in the drought tolerant lines. However, those genes expressed in both tolerant and sensitive genotypes may act as basal drought-responsive factors in tomato under drought stress. The specifically expressed gene involved in cell differentiation, growth and anatomical structure morphogenesis in the tolerant genotypes might also be beneficial to enhanced drought tolerance. The biochemical adaptation to water deficit in the tolerant genotypes may be performed by specifically expressed genes that were involved in biochemical pathways of gluconeogenesis, purine and pyrimidine nucleotides biosynthesis, tryptophan degradation, starch degradation, methionine biosynthesis, and removal of superoxide radicals. In addition,19 pathways belonging to the metabolism of secondary metabolites, hormones, nutrients, amino acids, sugars and polysaccharides, fatty acids and lipids, and C1 compounds were significantly affected in all three genotypes, which might be the primary biochemical processes in tomato under drought stress.4. The cloning, sequencing and protein structure analysis of candidate genes related to drought-tolerance. The differently expressed genes between tolerant and sensitive genotypes were selected to be subject for further analysis. Based on bioinformatics and biological technologies, the open reading frames of these genes were cloned and sequenced, whose amino acids and protein structure were deduced and predicted.5. The expression pattern of candidate genes in different tissues of tomato and in leaf treataed with different external factors. Semi-quantitative RT-PCR analysis of different tissues of tomato showed that SIWRKY, SpWRKY and SINAC had tissue-specific expression, while SITR, SpTR and SpNAC were constitutively expressed in all the tested tissues. All aimed genes were detected abundantly in leaves and stems, and weakly in fruits and/or roots of tomato. Most of them had higher expression under moderate and serve drought stress than mild treatment. Almost these genes were induced by salt, high and low temperature, longer exogenous ABA treatment and salicylic acid.6. The chromosome location of candidate genes. This study was conducted by PCR and digestion polymorphism based on tomato genome sequencing results. SpTR, SpWRKY and SpNAC were located on tomato chromosome 11,8 and 10, respectively.7. Function characterization of candidate genes. Overexpression vectors were transformed into one tomato cultivar by Agrobacterium-mediated transformation. The PCR analysis of putative transgenic plants showed that a total of 14,13 and 19 transgenic plants of SpTR, SpWRKY and SpNAC were obtained, respectively, indicating that T-DNA had integrated into the tomato genome. Semi-quantitative RT-PCR analysis of aimed genes expression showed that the higher expression abundance in the transgenic plants than wild type. Evaluation for abiotic stress tolerance of transgenic plants was measured. These results indicated that overexpression of SpTR and SpWRKY could improve the tolerance of drought, salt and low temperature.
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