Cheung Ming-yan. / Thesis submitted in 2000. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 169-185). / Abstracts in English and Chinese. / Thesis committee --- p.i / Statement --- p.ii / Abstract --- p.iii / Acknowledgement --- p.vi / General abbreviations --- p.viii / Abbreviations of chemicals --- p.x / List of figures --- p.xii / List of tables --- p.xv / Table of Contents --- p.xvii / Chapter Chapter 1 --- General Introduction - Using transgenic plants as bioreactor --- p.1 / Chapter 1.1 --- Plant as Bioreactor --- p.1 / Chapter 1.1.1 --- Plant transformation historical milestones --- p.1 / Chapter 1.1.2 --- Applications of transgenic plants --- p.5 / Chapter 1.1.2.1 --- Examples of in situ Application --- p.5 / Chapter 1.1.2.2 --- Examples of ex situ application of transgenic plant --- p.9 / Chapter 1.2 --- Plant Hosts for Transformation: Arabidopsis thaliana and Glycine max --- p.18 / Chapter 1.2.1 --- Essential components for plant transformation --- p.18 / Chapter 1.2.1.1 --- Marker genes --- p.18 / Chapter 1.2.1.2 --- Promoters --- p.18 / Chapter 1.2.2 --- Arabidopsis thaliana --- p.20 / Chapter 1.2.2.1 --- Agrobacterium-mediated transformation --- p.20 / Chapter 1.2.2.2 --- Transformation methods for A. thaliana --- p.21 / Chapter 1.2.3 --- Glycine max (soybean) --- p.22 / Chapter 1.2.3.1 --- Soybean cultivars for transformation --- p.23 / Chapter 1.2.3.2 --- Soybean regeneration systems --- p.24 / Chapter 1.2.3.3 --- Soybean transformation systems --- p.26 / Chapter 1.3 --- Target Pharmaceutical and Agricultural Proteins: Lymphocytic choriomeningitis virus and Goldfish Growth hormones I and II --- p.29 / Chapter 1.3.1 --- Production of pharmaceutical proteins --- p.29 / Chapter 1.3.1.1 --- Lymphocytic choriomeningitis virus --- p.30 / Chapter 1.3.1.2 --- Nucleoprotein of LCMV --- p.33 / Chapter 1.3.2 --- Agricultural protein category --- p.34 / Chapter 1.3.2.1 --- Carassius auratus --- p.34 / Chapter 1.3.2.2 --- Growth hormones I and II --- p.35 / Chapter 1.4 --- Hypothesis and Objectives --- p.37 / Chapter Chapter 2 --- Materials and Methods --- p.38 / Chapter 2.1 --- Materials --- p.38 / Chapter 2.1.1 --- "Plants, bacterial strains and vectors" --- p.38 / Chapter 2.1.2 --- Chemicals and Regents --- p.43 / Chapter 2.1.3 --- Commercial kits --- p.44 / Chapter 2.1.4 --- Primers and Adaptors --- p.45 / Chapter 2.1.5 --- Equipments and Facilities used --- p.47 / Chapter 2.1.6 --- "Buffer, solution and medium" --- p.47 / Chapter 2.2 --- Methods --- p.48 / Chapter 2.2.1 --- Molecular Techniques --- p.48 / Chapter 2.2.1.1 --- Bacterial cultures for recombinant DNA and plant transformation --- p.48 / Chapter 2.2.1.2 --- Recombinant DNA techniques --- p.48 / Chapter 2.2.1.3 --- "Preparation and transformation of DH5a, DE3 and Agrobacterium competent cells" --- p.49 / Chapter 2.2.1.4 --- Gel electrophoresis --- p.52 / Chapter 2.2.1.5 --- "DNA, RNA and protein extractions" --- p.53 / Chapter 2.2.1.6 --- Generation of cRNA probes for Southern and Northern blot analyses --- p.56 / Chapter 2.2.1.7 --- Southern blot analysis --- p.56 / Chapter 2.2.1.8 --- Northern blot analysis --- p.57 / Chapter 2.2.1.9 --- Expression of Lymphocytic choriomeningitis virus nucleoprotein (LCMV NP) in bacterial system --- p.58 / Chapter 2.2.1.10 --- Western blot analysis for LCMV NP --- p.59 / Chapter 2.2.1.11 --- Protein dot blot for detecting the presence of recombinant LCMV-NP generated from transgenic plants --- p.62 / Chapter 2.2.1.12 --- PCR techniques --- p.62 / Chapter 2.2.1.13 --- Sequencing --- p.63 / Chapter 2.2.2 --- Plant tissue culture and transformation --- p.64 / Chapter 2.2.2.1 --- Arabidopsis thaliana --- p.64 / Chapter 2.2.2.2 --- Soybean --- p.65 / Chapter 2.2.3 --- In vitro transcription and translation of target genes in rabbit reticulocyte and wheat germ systems --- p.68 / Chapter 2.2.3.1 --- In vitro transcription of target genes with with Ribomix large scale RNA production systems-T7 and SP6 (Promega) --- p.68 / Chapter 2.2.3.2 --- In vitro translation with rabbit reticulocyte lysate and wheat germ extract --- p.69 / Chapter Chapter 3 --- Results --- p.71 / Chapter 3.1 --- Expression of Lymphocytic choriomeningitis virus nucleoprotein (LCMV NP) and goldfish growth hormones I and II (GHI and GHII) in transgenic Arabidopsis thaliana --- p.71 / Chapter 3.1.1 --- Expression of LCMV-NP in transgenic Arabidopsis thaliana --- p.71 / Chapter 3.1.1.1 --- Cloning of the gene encoding LCMV NP into the binary vector system W104 --- p.71 / Chapter 3.1.1.2 --- Transformation of W104-LCMV-NP into the Agrobacterium GV3101/pMP90 --- p.78 / Chapter 3.1.1.3 --- Transformation of LCMV-NP cDNA into Arabidopsis thaliana --- p.80 / Chapter 3.1.1.4 --- Southern blot and Northern blot analyses of transgenic plant containing the LCMV-NP cDNA --- p.83 / Chapter 3.1.1.5 --- Production of recombinant LCMV-NP protein in DE3 cells --- p.90 / Chapter 3.1.1.6 --- Detection of recombinant LCMV-NP protein in transgenic A.thaliana --- p.98 / Chapter 3.1.2 --- Expression of goldfish growth hormones I and II (GHI and GHII) in transgenic Arabidopsis thaliana --- p.105 / Chapter 3.1.2.1 --- "Screening of homozygous lines of goldfish, Carassius auratus, growth hormones transgenic Arabidopsis thaliana" --- p.105 / Chapter 3.1.2.2 --- Southern blot and Northern blot analyses of transgenic plant containing the LCMV-NP cDNA --- p.109 / Chapter 3.1.2.3 --- Detection of recombinant GHI and GHII from transgenic plant --- p.112 / Chapter 3.2 --- In vitro transcription and translation of target genes in rabbit reticulocyte and wheat germ systems --- p.117 / Chapter 3.2.1 --- Subcloning of target genes in pGEM-3Zf(+) vector --- p.117 / Chapter 3.2.1.1 --- Subcloning of LCMV-NP fragment into pGEM-3Zf(+) vector --- p.117 / Chapter 3.2.1.2 --- Subcloning of goldfish GHI and GHII fragments into pGEM-3Zf(+) vector --- p.120 / Chapter 3.2.2 --- In vitro transcription of target genes with Ribomix large scale RNA production systems-T7 and SP6 --- p.125 / Chapter 3.2.3 --- In vitro translation with rabbit reticulocyte lysate and wheat germ extract systems --- p.128 / Chapter 3.3 --- Establishment of Glycine max regeneration and transformation systems --- p.130 / Chapter 3.3.1 --- The Establishment of soybean regeneration system --- p.130 / Chapter 3.3.2 --- Establishment of soybean transformation system --- p.133 / Chapter 3.3.2.1 --- Definition of transformation efficiency --- p.133 / Chapter 3.3.2.2 --- Effects of plant hosts --- p.136 / Chapter 3.3.2.3 --- Effects of Agrobacterium strains --- p.138 / Chapter 3.3.2.4 --- The application of vacuum infiltration --- p.139 / Chapter 3.3.2.5 --- Effect of kanamycin --- p.140 / Chapter 3.3.2.6 --- Effect of cocultivation duration and light/ dark treatment during germination --- p.141 / Chapter 3.3.2.7 --- Application of the detergent Silwet-77 --- p.142 / Chapter 3.3.3 --- Verification of transformation results by PCR screening --- p.143 / Chapter Chapter 4 --- Discussion --- p.147 / Chapter 4.1 --- "Expression of LCMV-NP, GHI and GHII in A. thaliana" --- p.148 / Chapter 4.2 --- Establishing a soybean transformation system --- p.157 / Chapter 4.2.1 --- Plant hosts and explants --- p.158 / Chapter 4.2.2 --- Regeneration of explants --- p.159 / Chapter 4.2.3 --- Agrobacterium strains --- p.161 / Chapter 4.2.4 --- Bacteria-plant interaction --- p.161 / Chapter 4.2.5 --- Transient versus stable transformation --- p.165 / Chapter 4.3 --- Conclusion and perspective --- p.167 / References --- p.169 / Appendix --- p.186
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_323523 |
| Date | January 2001 |
| Contributors | Cheung, Ming-yan., Chinese University of Hong Kong Graduate School. Division of Biology. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xxi, 197 leaves : ill. ; 30 cm. |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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