| Rice is one of the most important staple foods in the world. But rice production was confined by the lack of freshwater resources. As we know that, in our country agricultural production used too much freshwater resources, in which about 70% was consumed by rice. So studying the water-saving and drought resistance rice (WDR) was currently necessarily. At the present time, traditional breeding method is very significant effect in improving drought resistance of rice, but knows little about its molecular theory. As the complex truth of drought resistance theory, we must find and study more genes and gene network involved in drought response.In our study, we treated the rice that were planted in the soil with drought stress, then carried out a genome-wide profiling and analysis of miRNAs and mRNAs by microarray between drought-challenged rice and control. Two stress-responsive genes were studied for their expression levels under water stress. OsI17 gene was over-expressed in rice to test the ability on improving drought resistance. We found OsI17 gene could regulate the mRNA profiling and improve the drought resistance of rice, obviously. The main resalts are as follows:1. To gaina deep understanding of the mechanism of drought tolerance in rice, genome-wide profiling and analysis of miRNAs was carried out in drought-challenged rice across a wide range of developmental stages, from tillering to inflorescence formation, using a microarray platform. Among the miRNAs familes identified as significantly down- or up-regulated under the drought stress, miR170, miR172, miR397, miR408, miR529, miR896, miR1030, miR1035, miR1050, miR1088, and miR1126 were down-regulated miRNAs. miR395, miR474, miR845, miR851, miR854, miR901, miR903, and miR1125 were up-regulated miRNAs. miR156, miR168, miR170, miR171, miR172, miR319, miR396, miR397, and miR408 showed opposite expression to that observed in drought-stressed Arabidopsis. All of these revealed for the first time to be induced by drought stress in plants. The most conserved down-regulated miRNAs were ath-miR170, the miR171 family, and ath-miR396, and the most conserved up-regulated miRNAs were ptc-miR474 and ath-miR854a.2. We carried out a genome-wide profiling and analysis of mRNAs in drought-challenged rice by a microarray platform. A lot of metabolism pathways and genes were identified, which involved in drought-resistance of rice, in which the most important pathway was carbon fix. Maintaining a reasonable rate of synthesis of carbon-hydrogen compounds helps to enhance stress tolerance under drought-prone conditions3. Analyzed the expression profiling of OsDr1and OsI17 in rice. Up-regulated expression of OsDr1and OsI17 genes induced by various abiotic stresses (for e.g. Drought, salty, cold) and stress-related phytohormone (for e.g. ABA, JA, SA, and H2O2) treatments.4. We had cloned the full length cDNA of OsI17 in rice by RT-PCR method. Then we constructed the over-expression, antisense-RNA expression and OsI17-EGFP fuse vectors, and induced into Zhonghua11 (oryza sativa L.ssp. Japonica).6. OsI17 protein contains an AT-hook motif followed by a DUF296 domain, which was located in nucleus. AT-hook motif and DUF296 domain has nucleus location ability independently.7. The yeast two-hybrid testing showed that, the OsI17 protein (even if the DUF296 domain) had a homolog combining ability, that was necessary to playing its role.8. Under drought conditions, the transgenic rice has more content of total chlorophyll and lower chl a/b ratio than the WT. The drought stress testing at reproduce stage shows that, transgenic rice of OsI17 has higher POD activity, more small Vascular Bundles and bulliform cell groups in the flag leaf, higher RWC and Water-Retaining Capability of leaves. All of this shows that OsI17 enhanced the drought tolerance and drought avoidance of rice.9. Analysis of the transgenic rice profiling with microarray platform showed that, compared to WT,73 genes were up-regulated, and the PSII system was changed in evidence.10. The drought, salty and cold stress testing at seedling stage shows that, compared to the wild type, transgenic rice of OsI17 exhibited increased of stress tolerance. Compared to the wild type, the seed-setting rate, production per plant, biomass per plant and 100-seed weight of transgenic rice, exhibited 17-23%, more than 27%,25%～50% and more than 29% increase, respectively. The drought stress testing at reproduce stage shows that transgenic rice of OsI17 showed increased of drought tolerance.11. The OsI17 protein and the promoter all have sequences polymorphism among the cultivated rice varieties and wild rice lines (O.rufipogon). Phylogenetic tree of the OsI17 promoter showed that, the variety between japonica and indica had been exist among wild rice lines (O.rufipogon).12. The OsDr1 full length cDNA sequence was cloned by 5’-RACE and 3’-RACE method. OsDr1 expression was up-regulated by water stress, stress-related phytohormone; OsDr1 was expressed in circadian rhythms and regulated by light. |
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