| ABSTRACT | 第1-9页 |
| 摘要 | 第9-13页 |
| Chapter 1: REVIEW OF LITERATURE WITH AMBITION OF EXPERIMENT | 第13-20页 |
| ·Historical basis of the Pepper | 第13-14页 |
| ·Importance of the Pepper | 第14-15页 |
| ·Importance of polygalacturonase | 第15-17页 |
| ·Electron Microscopy | 第17页 |
| ·Agrobacterium Mediated Transformation | 第17-18页 |
| ·Function of Polygalacturonase in some other plants | 第18-19页 |
| ·Purpose of study | 第19-20页 |
| Chapter 2: Materials and Methods | 第20-45页 |
| ·Materials | 第20-24页 |
| ·Plant Material | 第20页 |
| ·Tissue culture propagation | 第20-24页 |
| ·Agrobacterium Strains and plasmid | 第24页 |
| ·Agrobacterium culture media with growing conditions | 第24页 |
| ·Ambient environment for In-Vitro propagation | 第24页 |
| ·Salient Chemicals and other Materials | 第24页 |
| ·Methods | 第24-36页 |
| ·Gene Cloning | 第24-25页 |
| ·Sequence Identification | 第25页 |
| ·Designing of Primers | 第25页 |
| ·Extraction of Plant DNA | 第25-26页 |
| ·Extraction of Total RNA | 第26-27页 |
| ·Construction of cDNA Library | 第27页 |
| ·The PCR Operations | 第27-29页 |
| ·Purification of PCR amplified DNA | 第29页 |
| ·Link amplified DNA with Vectors | 第29-31页 |
| ·Preparation of Competent E. coli cells using CaCl2or MgCl2 | 第31-32页 |
| ·Transformation of CaPG into E. coli | 第32页 |
| ·Plasmid Isolation from E. coli | 第32-36页 |
| ·Transformation | 第36-40页 |
| ·Transformation into Agrobacterium | 第36页 |
| ·Plasmid Isolation from Agrobacterium | 第36-37页 |
| ·The PCR Analysis | 第37-39页 |
| ·Validation of Gene Transformation by Enzymes Digestion Analysis | 第39-40页 |
| ·Ultra-structure Studies | 第40页 |
| ·Semi quantitative RT-PCR | 第40-41页 |
| ·Polygalacturonase Enzyme Assay | 第41-42页 |
| ·Collection of Crude Enzyme | 第41-42页 |
| ·Determination of Enzyme Activity | 第42页 |
| ·Morphological Studies | 第42-43页 |
| ·Statistical Analysis | 第43页 |
| ·Maintenance of Plant Materials during In-Vitro propagation | 第43-44页 |
| ·Surface sterilization of Pepper, Micro-Tom and Tobacco seeds | 第43页 |
| ·Micro-propagation of Pepper, Tomato and Tobacco Explants | 第43页 |
| ·Transformation of cotyledon explants | 第43-44页 |
| ·Post-Transformation | 第44-45页 |
| Chapter 3: Results | 第45-60页 |
| ·Verification of CaPG gene and E8 promoter cloning | 第45-48页 |
| ·Verification of linkage of imported CaMV35S promoter | 第46页 |
| ·Verification of linkage of imported NOS terminal | 第46页 |
| ·PCR verification of CaPG gene and E8 promoter within Plasmids | 第46-48页 |
| ·Enzymes Digestion verification within Plasmids | 第48页 |
| ·RT-PCR for determining CaPG gene Expression | 第48-49页 |
| ·Polygalacturonase Assay | 第49-50页 |
| ·Ultra structure Observations | 第50-56页 |
| ·Electron Microscopy at Green Stage | 第50-51页 |
| ·Electron Microscopy at Breaker Stage | 第51页 |
| ·Electron Microscopy at Fully Ripened Stage | 第51-56页 |
| ·Morphological Aspects | 第56页 |
| ·Transgenic Pepper, Micro-Tom and Tobacco lines | 第56-60页 |
| ·Induction of Callus | 第56-57页 |
| ·Development of Shoots and Roots | 第57-58页 |
| ·Development and hardening in Soil | 第58-59页 |
| ·Validation of CaPG gene in B-72 and Tobacco cultivars | 第59-60页 |
| Chapter 4: Discussion | 第60-64页 |
| ·Role of polygalacturonase in mechanism of pepper fruit ripening | 第60-61页 |
| ·Quantification of CaPG gene expression by RT-PCR | 第61页 |
| ·Quantification of CaPG gene expression by Transmission Electron Microscopy | 第61-62页 |
| ·Prospects of Expression vector pVBG2307 | 第62-63页 |
| ·Validation of CaPG Gene in the transgenic plants | 第63-64页 |
| Conclusion | 第64-66页 |
| ACKNOWLEDGMENT | 第66-67页 |
| References | 第67-72页 |
| Appendix A: Agrobacterium Mediated Transformation | 第72-73页 |
| APPENDIX B: Plant Tissue Culture media having different compositions | 第73-74页 |
| APPENDIX C: Bacterial Culture Medium | 第74页 |
| APPENDIX D: Liquid Broth for Bacterial Cultivation | 第74页 |
| APPENDIX E: Plasmid Isolation Solutions and TE Buffer | 第74-75页 |
| APPENDIX F: TRI reagent | 第75页 |
| APPENDIX G: CTAB Buffer | 第75-76页 |
| APPENDIX H: 50×TAE (Tris-acetate) Buffer (1 L) | 第76页 |
| APPENDIX I: Preparation Solutions for Developing Competent Cells | 第76-77页 |
| APPENDIX J: Restriction enzymes in multiple cloning sites of vector pVBG2307 to provide greater manipulation of DNA fragments | 第77页 |
| APPENDIX K: List of cloned genes, cleaved elements and binary vectors sequenced in pVBG2307, including their cutting sites and sizes | 第77-78页 |
| APPENXIX L: Sequences of CaPG gene and E8 promoters BLAST from NCBI | 第78页 |
| APPENDIX M BLAST result of cloned CaPG gene | 第78-81页 |
| Curriculum vitae | 第81-78页 |
| APPENDIX M BLAST result of cloned CaPG gene | 第78-81页 |
| Curriculum vitae | 第81-85页 |
