| ACKNOWLEDGEMENTS | 第6-8页 |
| ABSTRACT | 第8-10页 |
| TABLE OF CONTENTS | 第11-16页 |
| LIST OF FIGURES | 第16-19页 |
| LIST OF TABLES | 第19-20页 |
| ABBREVIATIONS | 第20-23页 |
| CHAPTER Ⅰ Introduction | 第23-34页 |
| 1.1 Desiccation damage | 第23页 |
| 1.2 Definition of desiccation tolerance | 第23-24页 |
| 1.3 Desiccation tolerance in seeds | 第24-25页 |
| 1.4 Mechanisms of desiccation tolerance | 第25-26页 |
| 1.5 Antioxidative mechanism | 第26-30页 |
| 1.5.1 Reactive oxygen species (ROS) sources | 第26-27页 |
| 1.5.2 Damage risks by ROS accumulation | 第27页 |
| 1.5.3 Signal role of ROS in metabolism | 第27-28页 |
| 1.5.4 Antioxidant enzymes | 第28-30页 |
| 1.5.4.1 Superoxide dismutases (SOD) | 第28页 |
| 1.5.4.2 Catalase (CAT) | 第28-29页 |
| 1.5.4.3 Enzymes of the ascorbate-glutathione cycle | 第29-30页 |
| 1.6 Production of protective proteins during seed development | 第30-32页 |
| 1.6.1 Late embryogenesis abundant (LEA) proteins | 第30-31页 |
| 1.6.2 Small heat shock proteins (HSPs) | 第31-32页 |
| 1.7 Jatropha curcas as an experimental material | 第32页 |
| 1.8 Aim of the present work | 第32-34页 |
| CHAPTER Ⅱ Materials and methods | 第34-65页 |
| 2.1 Plant materials | 第34-35页 |
| 2.2 Measurement of seed weight, volume and moisture content | 第35页 |
| 2.3 Germination test | 第35页 |
| 2.4. Measurement of relative growth rate | 第35-36页 |
| 2.5 Desiccation treatments | 第36页 |
| 2.6 Conductivity test | 第36页 |
| 2.7 Evaluation of hydrogen peroxide (H_2O_2) content | 第36-37页 |
| 2.8 Evaluation of lipid peroxidation product | 第37页 |
| 2.9 Antioxidant enzyme assays | 第37-39页 |
| 2.9.1 Superoxide dismutases (SOD) assay | 第38页 |
| 2.9.2 Catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) activity assays | 第38-39页 |
| 2.9.3 Dehydro ascorbate reductase (DHAR) assay | 第39页 |
| 2.10 Native polyacrylamide gel electrophoresis (Native-PAGE) analysis of antioxidant enzymes | 第39-43页 |
| 2.10.1 Native-PAGE solutions | 第39-40页 |
| 2.10.2 Preparing the gels | 第40-41页 |
| 2.10.3 SOD activity | 第41-42页 |
| 2.10.4 CAT activity | 第42页 |
| 2.10.5 APX activity | 第42页 |
| 2.10.6 GR activity | 第42-43页 |
| 2.11 Protein isolation and analysis | 第43-49页 |
| 2.11.1 Crude protein | 第43页 |
| 2.11.2 Heat stable protein | 第43页 |
| 2.11.3 Protein quantification | 第43-44页 |
| 2.11.4 Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) | 第44-47页 |
| 2.11.4.1 SDS-PAGE solutions | 第44-45页 |
| 2.11.4.2 Preparing the gels | 第45-47页 |
| 2.11.5 Western blot analysis | 第47-49页 |
| 2.11.5.1 Western blot buffers | 第47-48页 |
| 2.11.5.2 Western blot procedures | 第48-49页 |
| 2.12 First-strand cDNA Synthesis | 第49-52页 |
| 2.12.1 Total RNA extraction | 第49-50页 |
| 2.12.2 Determination of RNA concentration | 第50-51页 |
| 2.12.3 DNase digestion | 第51页 |
| 2.12.4 First-strand cDNA synthesis reaction | 第51-52页 |
| 2.13 Polymerase Chain reaction (PCR) | 第52-62页 |
| 2.13.1 Common procedure for PCR | 第52页 |
| 2.13.2 Separation of DNA by agarose gel electrophoresis | 第52-53页 |
| 2.13.3 Semi-quantitative reverse transcriptase polymerase chain reaction | 第53页 |
| 2.13.4 Rapid Amplification of cDNA 3' Ends (3'-RACE) | 第53-56页 |
| 2.13.4.1 First-strand cDNA synthesis for 3'-RACE | 第54-55页 |
| 2.13.4.2 3'-RACE reaction | 第55-56页 |
| 2.13.5 Quantitative real-time polymerase chain reaction analysis (qRT-PCR) | 第56-62页 |
| 2.13.5.1 qRT-PCR overview | 第56-59页 |
| 2.13.5.2 Primer design and PCR efficiency | 第59-60页 |
| 2.13.5.3 qRT-PCR reaction | 第60-61页 |
| 2.13.5.4 Thermal cycling conditions | 第61-62页 |
| 2.14 E. coli transformation | 第62-64页 |
| 2.14.1 Purification of DNA fragment from agarose gel | 第62页 |
| 2.14.2 Solutions and media | 第62-63页 |
| 2.14.3 Preparing competent cells | 第63-64页 |
| 2.14.4 Transformation procedure | 第64页 |
| 2.15 Sequence analysis | 第64-65页 |
| CHAPTER Ⅲ Changes in production and scavenging of hydrogen peroxide during natural and artificial dehydration of developing Jatropha curcas seeds | 第65-100页 |
| 3.1 Introduction | 第65-67页 |
| 3.2 Results | 第67-91页 |
| 3.2.1 Changes during the maturation and natural desiccation phase | 第67-79页 |
| 3.2.1.1 Changes in J. curcas fruits and seed size during development | 第67-69页 |
| 3.2.1.2 Changes in fresh and dry weight and water content of jatropha curcas seeds during development | 第69-70页 |
| 3.2.1.3 Germinability and leakage rate of electrolyte of whole seeds and isolated embryos during development | 第70-71页 |
| 3.2.1.4 Hydrogen peroxide contents and lipid peroxidation | 第71-73页 |
| 3.2.1.5 Antioxidant enzyme activities | 第73-76页 |
| 3.2.1.6 Expression profiling of antioxidant enzyme genes | 第76-79页 |
| 3.2.2 Effect of artificial dehydration on developing seeds | 第79-84页 |
| 3.2.2.1 Effect of dehydration on seed water content | 第79-80页 |
| 3.2.2.2 Effect of dehydration on seed germinability | 第80-82页 |
| 3.2.2.3 Effect of dehydration on membrane leakage | 第82-83页 |
| 3.2.2.4 H_2O_2 and MDA contents | 第83-84页 |
| 3.2.3 Comparison between natural and artificial dehydration effects on H_2O_2 levels and antioxidant enzyme activities of developing seeds | 第84-91页 |
| 3.2.3.1 Seed development and acquisition of desiccation tolerance | 第85-86页 |
| 3.2.3.2 H_2O_2 and MDA contents | 第86-87页 |
| 3.2.3.3 Antioxidant enzyme activities | 第87-91页 |
| 3.3 Discussion | 第91-100页 |
| 3.3.1 Changes in seed germinability during development and after artificial dehydration | 第91-92页 |
| 3.3.2 Changes in H_2O_2 and MDA | 第92-94页 |
| 3.3.3 Changes in antioxidant enzymes | 第94-100页 |
| CHAPTER Ⅳ Changes in protein expression profiles during the maturation and after artificial dehydration of developing Jatropha curcas seeds | 第100-109页 |
| 4.1 Introduction | 第100-101页 |
| 4.2 Results | 第101-108页 |
| 4.2.1 Total protein profile during seed development | 第101-104页 |
| 4.2.2 Changes in total protein profile after artificial dehydration of developing seeds | 第104页 |
| 4.2.3 Changes in heat shock protein profile during dehydration at different developmental stages | 第104-105页 |
| 4.2.4 Changes in membrane leakage during dehydration at different developmental stages | 第105-108页 |
| 4.3 Discussion | 第108-109页 |
| CHAPTER Ⅴ Identification and characterization of two dehydrin cDNAs during maturation of Jatropha curcas seeds | 第109-128页 |
| 5.1 Introduction | 第109-111页 |
| 5.2 Results | 第111-125页 |
| 5.2.1 Dehydrins were induced by maturation and natural dehydration process | 第111-113页 |
| 5.2.2 Identification and characterization of dehydrin cDNAs from J. curcas seeds | 第113-121页 |
| 5.2.2.1 Jatropha curcas dehydrin-1 (JcDHN-1) | 第114-116页 |
| 5.2.2.1.1 Characterization of the full length cDNA of JcDHN-1 | 第114-115页 |
| 5.2.2.1.2 Homology analysis of JcDHN-1 protein with dehydrins from other plants | 第115-116页 |
| 5.2.2.2 Jatropha curcas dehydrin-2 (JcDHN-2) | 第116-121页 |
| 5.2.2.2.1 Characterization of the full length cDNA of JcDHN-2 | 第116-119页 |
| 5.2.2.2.2 Homology analysis of JcDHN-2 protein with dehydrins from other plants | 第119-121页 |
| 5.2.3 Alignment of JcDHN-1 and JcDHN-2 proteins | 第121-123页 |
| 5.2.4 Analysis of Jatropha curcas DHNs gene expression by qRT-PCR | 第123-125页 |
| 5.3 Discussion | 第125-128页 |
| CHAPTER Ⅵ Identification and characterization of two small heat shock proteins cDNAs during maturation of Jatropha curcas seeds | 第128-143页 |
| 6.1 Introduction | 第128-129页 |
| 6.2 Results | 第129-141页 |
| 6.2.1 Detection of sHSPs during maturation and natural dehydration process | 第129页 |
| 6.2.2 Identification and characterization of detected sHSPs | 第129-137页 |
| 6.2.2.1 Jatropha curcas heat shock protein-1 (JcHSP-1) | 第131-134页 |
| 6.2.2.1.1 Characterization of the full length cDNA of JcHSP-1 | 第131页 |
| 6.2.2.1.2 Homology analysis of JcHSP-1 protein | 第131-134页 |
| 6.2.2.2 Jatropha curcas heat shock protein-2 (JcHSP-2) | 第134-137页 |
| 6.2.2.2.1 Characterization of the full length cDNA of JcHSP-2 | 第134页 |
| 6.2.2.2.2 Homology analysis of JcHSP-2 protein | 第134-137页 |
| 6.2.3 Alignment of JcHSP-1 and JcHSP-2 | 第137-138页 |
| 6.2.4 Analysis of Jatropha curcas sHSPs gene expression by qRT-PCR | 第138-141页 |
| 6.3 Discussion | 第141-143页 |
| General discussion | 第143-145页 |
| References | 第145-164页 |
| 摘要 | 第164-165页 |
