| 摘要 | 第7-9页 |
| ABSTRACT | 第9-11页 |
| Chapter 1 Review of literature | 第16-31页 |
| 1.1 Introduction | 第16-17页 |
| 1.2 Role of iron in plants | 第17-19页 |
| 1.2.1 Iron deficiency, toxicity and Its impacts on rice | 第17-18页 |
| 1.2.2 Mechanism of iron (Fe) uptake and transport in rice | 第18-19页 |
| 1.3 Subcellular regulation of iron | 第19-23页 |
| 1.3.1 Vacuoles; as largest pool of iron trafficking | 第20页 |
| 1.3.2 Plasma membrane ferric reductase activity | 第20页 |
| 1.3.3 Mitochondrial ferric reductase activity | 第20-21页 |
| 1.3.4 Role of mitochondrial iron transporter (MIT) & nucleus in iron (Fe) transport | 第21-23页 |
| 1.4 Loading of iron (Fe) into grain | 第23-24页 |
| 1.5 Gene families related to iron (Fe) homeostasis | 第24-26页 |
| 1.5.1 Nicotinamide Synthesis (NAS) Gene Family | 第24-25页 |
| 1.5.2 Natural resistance-associated macrophage (NRAMP) gene family | 第25页 |
| 1.5.3 Yellow stripe (YSL) gene family | 第25页 |
| 1.5.4 Iron-regulated transporter (IRT) gene family | 第25-26页 |
| 1.5.5 Ferric reduction oxidase (FRO) gene family | 第26页 |
| 1.6 Rice FRO genes and their major functional domains | 第26-28页 |
| 1.6.1 Subcellular localization and role of FROs in plant development | 第27页 |
| 1.6.2 Transcriptomic changes of FRO genes during imbalance supply of iron | 第27-28页 |
| 1.7 Approaches towards Iron (Fe) biofortification | 第28-30页 |
| 1.7.1 Transgenic approach | 第29页 |
| 1.7.2 Discovery of novel genes and their functional characterization | 第29-30页 |
| 1.8 Aims and objectives of research work | 第30-31页 |
| Chapter 2 Expansion and Evolutionary Mechanism of Plant FRO Gene Family | 第31-53页 |
| 2.1 Introduction | 第31页 |
| 2.2 Methodology | 第31-34页 |
| 2.2.1 Information of plant genomic databases | 第31页 |
| 2.2.2 Identification of FRO family members | 第31-32页 |
| 2.2.3 Phylogenetic analysis, chromosomal location and duplication | 第32-33页 |
| 2.2.4 Gene structure, conserved domain and motif analysis | 第33-34页 |
| 2.2.5 Analysis of promoter region | 第34页 |
| 2.3 Results | 第34-49页 |
| 2.3.1 Identification, classification, and annotation of FRO family members | 第34-40页 |
| 2.3.2 Systematic evolutionary relationship of FRO genes | 第40-42页 |
| 2.3.3 Structural diversity of FRO genes | 第42页 |
| 2.3.4 Analysis of promoter region, conserved motifs and domains | 第42-47页 |
| 2.3.5 Chromosomal location and gene duplication of FRO genes | 第47-49页 |
| 2.4 Discussion | 第49-52页 |
| 2.4.1. FRO genes expansion, duplication, and structural diversity | 第49-51页 |
| 2.4.2. Evolution and functional diversity of conserved domains in plant FRO gene | 第51-52页 |
| 2.5 Conclusion | 第52-53页 |
| Chapter 3 Expression Analysis of Rice FRO Genes in Response to Abiotic Stresses, Metalsand Hormone Applications | 第53-70页 |
| 3.1 Introduction | 第53-54页 |
| 3.1.1 Instruments | 第53-54页 |
| 3.1.2 Reagents | 第54页 |
| 3.2 Plant materials and growth conditions | 第54-58页 |
| 3.2.1 Stress treatments and sample collection | 第54-55页 |
| 3.2.2 Expression analysis | 第55页 |
| 3.2.3 RNA extraction and c DNA synthesis | 第55-56页 |
| 3.2.4 RNA extraction | 第56页 |
| 3.2.5 cDNA Synthesis | 第56-57页 |
| 3.2.6 qRT-PCR analysis | 第57-58页 |
| 3.3 Antioxidant enzyme activities under metal stresses | 第58-59页 |
| 3.4 Statistical analysis | 第59页 |
| 3.5 Results | 第59-67页 |
| 3.5.1 Tissue-specific expression profile of OsFRO1 and OsFRO7 genes | 第59页 |
| 3.5.2 Inducible expression analysis of rice OsFRO1 and OsFRO7 gene under abioticand hormone applications | 第59-64页 |
| 3.5.3 Inducible expression analysis of rice OsFRO1 and OsFRO7 genes againstdifferent heavy metals stresses | 第64-66页 |
| 3.5.4 Determination of antioxidant enzyme activities | 第66-67页 |
| 3.6 Discussion | 第67-69页 |
| 3.6.1 Tissue-specific gene expression of rice FROs and response to metal stress | 第67-68页 |
| 3.6.2 Inducible expression pattern of OsFRO1 and OsFRO7 genes against abioticstresses and hormones treatments | 第68-69页 |
| 3.6.3 Changes in antioxidant activity under iron (Fe) and chromium (Cr) stress | 第69页 |
| 3.7 Conclusion | 第69-70页 |
| Chapter 4 Functional Characterization of Os RO1 Gene and Stress Tolerance Mechanism | 第70-93页 |
| 4.1 Introduction | 第70页 |
| 4.2 Materials, Reagents and Instruments | 第70-74页 |
| 4.2.1 Antisense RNA Vector Construction | 第71-72页 |
| 4.2.2 Subcellular localization of OsFRO1 | 第72-73页 |
| 4.2.3 Histochemical beta-glucuronidase (GUS) analysis | 第73页 |
| 4.2.4 Transfection of cloning vector into agrobacterium strain EHA-105 | 第73-74页 |
| 4.3 Genetic transformation of Os FRO1-RNAi into rice | 第74-75页 |
| 4.3.1 Bacterial suspension and agrobacterium infection and co-cultivation | 第74页 |
| 4.3.2 Regeneration and transgene conformation | 第74-75页 |
| 4.4 Tissue Culture | 第75-76页 |
| 4.4.1 Surface sterilization of seeds and Callus Induction | 第75页 |
| 4.4.2 Selection, Regeneration and recovery of Transgenic Plants | 第75-76页 |
| 4.5 Expression analysis of Os FRO1-RNAi | 第76-77页 |
| 4.5.1 Tissue specific and inducible expression profile | 第76页 |
| 4.5.2 Determination of antioxidant activity and Fe concentration in rice shoots | 第76页 |
| 4.5.3 Determination of leaf pigments and root activity | 第76-77页 |
| 4.6 Experimental data and statistical analysis | 第77页 |
| 4.7 Results | 第77-80页 |
| 4.7.1 Cloning of OsFRO1 gene and construction of expression vector | 第77页 |
| 4.7.2 Agrobacterium mediated transformation | 第77-78页 |
| 4.7.3 Agrobacterium growth and infection | 第78页 |
| 4.7.4 Selection and transgene analysis | 第78页 |
| 4.7.5 Subcellular localization of OsFRO1 in rice | 第78页 |
| 4.7.6 Histochemical beta-glucuronidase (GUS) analysis | 第78-80页 |
| 4.8 Analysis of expression profile of osfro1 in different tissues and young panicle | 第80-89页 |
| 4.8.1 Phenotypic characterization of osfro1 | 第80-83页 |
| 4.8.2 Reduced tolerance of osfro1 silenced in rice roots | 第83-84页 |
| 4.8.3 Knockdown effect of osfro1 in ROS network by modulation of enzymaticactivity | 第84页 |
| 4.8.4 Knockdown osfro1 leads to Fe-sensitivity during germination and seedlinggrowth | 第84-86页 |
| 4.8.5 Knockdown effect of osfro1 on abiotic stresses and hormones | 第86-87页 |
| 4.8.6 Expression of osfro1 and chlorophyll content under Fe stress | 第87-89页 |
| 4.9 Discussion | 第89-93页 |
| Chapter 5 General Summary and Future Perspectives | 第93-96页 |
| 5.1 General summary | 第93-95页 |
| 5.2 Future perspectives | 第95-96页 |
| References | 第96-113页 |
| Appendices | 第113-127页 |
| Appendix-1. List of symbols and abbreviations | 第113-114页 |
| Appendix-2. Media used for japonica rice tissue culture & transformation | 第114-117页 |
| Appendix-3. Media composition for bacterial growth | 第117-118页 |
| Appendix-4. Preparation of different reagents used in tissue culture media | 第118-119页 |
| Appendix-5. Plasmid extraction method | 第119-120页 |
| Appendix-6. Plant Tissue SOD, POD, CAT and MDA determination | 第120-122页 |
| Appendix-7. Primers used for vector construction and q RT-PCR analysis | 第122-123页 |
| Appendix-8. The domain structure of the FRO proteins | 第123-125页 |
| Appendix-9. Site of amino acids and motif location of FROs protein | 第125-126页 |
| Appendix-10. Schematic diagram of amino acid motifs of FRO, s proteins | 第126-127页 |
| Acknowledgements | 第127-128页 |
| CURRICULUM VITAE | 第128-129页 |
