| 摘要 | 第1-7页 |
| Abstract | 第7-13页 |
| Chapter 1 Literature review | 第13-23页 |
| ·Wuzhishan minipigs | 第13-14页 |
| ·Fluorescent proteins | 第14-16页 |
| ·The development history of red fluorescent protein as a marker | 第14-15页 |
| ·Monomeric red fluorescent protein (mRFPl) | 第15页 |
| ·Animal modal expressing fluorescent proteins | 第15-16页 |
| ·Transgeaic animals | 第16-22页 |
| ·Techniques for genetic modifications pig | 第16-18页 |
| ·Pronuclear microinjection | 第16-17页 |
| ·Retroviral vectors | 第17页 |
| ·Sperm-mediated gene transfer (SMGT) | 第17-18页 |
| ·Embryonic stem cells | 第18页 |
| ·Somatic cell nuclear transfer | 第18页 |
| ·Generation of transgenic animal by somatic cell nuclear transfer | 第18-20页 |
| ·The preparation of donor cells | 第19-20页 |
| ·Calcium Phosphate Precipitation | 第19-20页 |
| ·Electroporation | 第20页 |
| ·Lipid transfection | 第20页 |
| ·Analysis of transgenic animals | 第20-21页 |
| ·The development prospects of genetically modified animals | 第21-22页 |
| ·Aim of this study | 第22-23页 |
| Chapter 2 Methods and materials | 第23-31页 |
| ·Animals | 第23页 |
| ·Chemicals and media | 第23页 |
| ·Main equipment | 第23-24页 |
| ·Methods | 第24-30页 |
| ·Construction of transgenic pCX-mRFPl-pgk-neoR vectors | 第24-25页 |
| ·Generation of Wuzhishan minipigs ear fibroblast cell line (WMP) | 第25页 |
| ·Selection of susceptive concentration of antibiotics G418 in Wuzhishan minipigs ear fibroblast cell line | 第25页 |
| ·Transfection of pCX-mRFPl-pgk-neoR into ear fibroblasts | 第25-27页 |
| ·FuGENE(?) HD Transfection Reagent were used to transfect ear fibroblasts | 第25-26页 |
| ·Lipofectamine TM 2000 Transfection Reagent were used to transfect ear fibroblasts | 第26页 |
| ·Electroporation were used to transfect ear fibroblasts | 第26-27页 |
| ·WMP/mRFPl transfected fibroblasts by PCR assay | 第27页 |
| ·Production of SCNT embryos | 第27-28页 |
| ·Embryo transfer | 第28页 |
| ·Identification of positive clones | 第28-29页 |
| ·Karyotyping | 第29页 |
| ·Fluorescence in situ hybridization (FISH) | 第29页 |
| ·Detection of red fluorescence | 第29-30页 |
| ·Statistical analysis | 第30-31页 |
| Chapter 3 Results and analysis | 第31-42页 |
| ·Selection of Susceptive Concentration of Antibiotics G418 in Wuzhishan minipigs ear fibroblast cell line | 第31页 |
| ·Comparison of transfection efficiency of pCX-mRFPl-pgk-neoR into ear fibroblasts | 第31-33页 |
| ·Generation of cloned embryos by nuclear transfer of mRFPl-expressing cells | 第33-34页 |
| ·In vitro development and mRFPl expression of SCNT embryos | 第34-35页 |
| ·Generation of transgenic Wuzhishan minipigs | 第35-36页 |
| ·Analysis of mRFPl expression in transgenic pigs | 第36-42页 |
| Chapter 4 Discussion | 第42-47页 |
| ·Comparison of transfection efficiency of pCX-mRFPl-pgk-neoR into ear fibroblasts | 第42-44页 |
| ·Expression of red fluorescent protein transgenic Wuzhishan minipigs | 第44页 |
| ·Generation of transgenic Wuzhishan minipigs by somatic cell nuclear transfer | 第44-45页 |
| ·Analysis of transgenic Wuzhishan minipigs | 第45-47页 |
| Chapter 5 Conclusion | 第47-48页 |
| Reference | 第48-53页 |
| Appendix Components of basic medie | 第53-55页 |
| Appendix B | 第55-56页 |
| Acknowledgement | 第56-57页 |
