Several isoforms of LPP have now been identified and cloned but remain to be purified. In the present study, a bacterial expression system was established and hexa- and deca-histidine epitope tagged-LPP1 and LPPla expressed in E. coli. In addition, a maltose binding protein (MBP) epitope tagged-LPP Ia was expressed in E. coli. Hexa- and deca-histidine LPP1 and LPPla, were partially purified using immobilised affinity chromatography. MBP-LPPla was expressed to higher levels than hexa- and deca-histidine LPP1 and LPPla in E. coli, most probably within insoluble inclusion bodies. In all cases, recovery of LPP activity was low. Membranes derived from HEK293 cells that stably over-express LPP I, LPP I a, LPP2 or LPP3 were used to demonstrate the differential hydrolysis of several molecular species of PA, LPA(18: 1), C8-CIP and SlP. Kinetic analysis using a multisubstrate assay system revealed that the LPP isoforms do not follow typical Michaelis-Menten kinetics towards most substrates under the assay conditions employed. The LPPs appear to show differential kinetics depending on the complement of substrates accessible to the enzymes. Stable over-expression of LPPI, LPPla, LPP2, but not LPP3, in HEK293 cells has previously been shown to attenuate the activation of ERK-1/2 by G-protein coupled receptors agonists such as SIP, LPA, PA and thrombin. The present study extended these observations by showing that basal growth rates were unaffected and that levels of mRNA transcript for the SIP, /EDGI receptor were reduced in the LPP stable cell lines but that this did not correlate with attenuation of the SIP-stimulated ERK-1/2 response. In addition, transient overexpression of LPPI, LPPIa and LPP2, but not LPP3 in HEK293 cells and GPASM cells also resulted in the attenuation of SIPinduced ERK-1/2 activation. Furthermore, transient transfection of a plasmid construct encoding the antisense sequence for LPP1 was also found to attenuate SIPinduced ERK-1/2 activation whereas the PMA-stimulated response was unaffected. Many questions remain to be fully answered in order to determine the physiological and pathophysiological. roles of the LPPs and the reason for the molecular diversity of the enzyme family.

This thesis reports eight experiments which investigate voluntary emotional expressions in head-injured and normal children and four experiments which investigate spontaneous expressiveness in head-injured and normal children. A novel technique using dynamic video film was used to elicit more truly spontaneous expressions and then surreptitiously video the child's expressive responses to the film clips. In the voluntary condition children posed expressions on verbal request and imitated expressions depicted in photographs. Independent adult raters then viewed still images of the children's' expressions and categorised them for emotion shown and rated intensity of expression. Results indicated that for the spontaneous expressions not all film clips elicited expressions equally as well. The best clips were those that elicited happiness, sadness and disgust and differences in expressiveness were shown to exist between the head-injured and normal children. The acquired head-injury children as a group were significantly less expressive than the controls while differential effects on positive and negative spontaneous expression were found in left and right congenital hemiplegics. In right hemiplegics (left brain damage) the spontaneous expressions of disgust, sadness and happiness were poorer than controls, while in left hemiplegics (right brain damage) only negative spontaneous expressions of disgust and sadness were poorer than controls. These findings add new information to the existing body of knowledge in that they suggest that the left hemisphere is more important than one thought for the expression of negative emotion. The results from the voluntary production conditions indicated that the head-injured children could accurately imitate expressions suggesting that their ability to manipulate facial muscles was intact. However, the acquired head-injury children were found to be poorer than controls at producing expressions on verbal request particularly so in the left brain damage group.

Exposures to ionising radiation resulting from natural sources or medical diagnostics are generally very low. In contrast, exposures to therapeutic radiation, although, they are often partial exposures, represent much higher doses. Similar levels of exposure may also occur as a consequence of a radiological accident, where it would be necessary to quickly separate individuals requiring urgent medical attention from the “worried-well”. The well-established biodosimetry techniques based on cytogenetics, particularly scoring dicentric chromosomes, are accurate and sensitive, yet, they are unsuitable for mass screening due to limited capacity and the time required for providing dose estimates. Measuring gene expression changes following radiation exposure was suggested over a decade ago to be an alternative method for dose estimation, as it is a quick, sensitive and suitable technique for high throughput application. The qPCR protocol was extensively optimised for increased reproducibility and sensitivity in order to be suitable for biodosimetry purposes. Radiation-responsive transcripts were identified and characterised in terms of temporal- and dose-responses. Finally, candidate transcripts were validated in human blood irradiated ex vivo and in vivo in blood samples obtained from cancer patients undergoing radiotherapy treatment. The data generated here serve as a proof of principle that qPCR-based gene expression assays can be used for radiation biodosimetry purposes to aid classical cytogenetics tools in an event of mass causality.

Der transkriptionelle Repressor Blimp-1 wurde ursprünglich als essentiell für die terminale Differenzierung von B-Zellen zu Antikörper-produzierenden Plasmazellen beschrieben. Im Rahmen dieser Arbeit wurde die Expression von Blimp-1 in humanen T-Zellen untersucht. Die Versuchsergebnisse zeigen, dass Blimp-1 auch in humanen T-Zellen sowohl auf mRNA- als auch auf Proteinebene exprimiert wird. Es ist deshalb anzunehmen, dass Blimp-1 auch für die terminale Differenzierung von T-Zellen eine wichtige Rolle spie / none

Thesis (M. Phil.)--Chinese University of Hong Kong. / Bibliography: leaves 158-185.

La réponse cellulaire aux dommages de l'ADN provoqués par l'irradiation (IR) est relativement bien étudiée mais de nombreuses observations montrent l'implication de l'expression de nombreux gènes. Nous souhaitons identifier les différentes formes de la réponse transcriptionnelle à l'IR et reconstruire un réseau de régulation génique impliqué dans son contrôle. La problématique réside dans l'exploitation de dynamiques d'expression de gènes dans des conditions de perturbations génétiques et dans l'intégration d'informations biologiques systémiques. Nous définissons une approche constituée d'une étape automatisée de déduction de régulations à partir de perturbations et de deux étapes d'induction qui permettent d'analyser la dynamique d'expression des gènes et d'extraire des régulations des données additionnelles. Cela nous a permis d'identifier, chez la levure, une réponse complexe à l'IR et de proposer un modèle de régulation dont certaines relations ont été validées expérimentalement. / The cellular response to the DNA damage provoked by irradiation (IR) is relatively well studied, however, many observations show the involvement of the expression of many genes. We propose to identify the different potential patterns of the transcriptional response to IR and to reconstruct a gene regulatory network involved in its control. The first point of this work lies in the exploitation of the gene expression dynamics in conditions of genetic perturbations. The second point lies in the integration of systemic biological informations. We define an approach composed of one step of automated logical deduction of regulations from a strategy of perturbations and two induction steps that allow the analysis of the gene expression dynamics and the extraction of potential regulation from additional data. This approach allowed to identify, for the yeast, a complex response to IR and allowed to propose a regulation model which some relations have been experimentally validated.

No description available.

Chum Wing Yan, Winnie. / Thesis submitted in 2002. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 157-178). / Abstracts in English and Chinese. / English Abstract --- p.i / Chinese Abstract --- p.iii / Acknowledgements --- p.v / Abbreviations --- p.vi / Table of Contents --- p.vii / List of Figures --- p.x / List of Tables --- p.xiii / Chapter Chapter One --- Literature Review / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Life cycle --- p.2 / Chapter 1.3 --- Nutritional value --- p.4 / Chapter 1.4 --- Medicinal value --- p.4 / Chapter 1.4.1 --- Antitumor ability --- p.5 / Chapter 1.4.2 --- Antimicrobial ability --- p.5 / Chapter 1.4.3 --- Hypocholesterolaemic effect --- p.6 / Chapter 1.4.4 --- Anti-viral effect --- p.6 / Chapter 1.4.5 --- Anticaries effects --- p.7 / Chapter 1.6 --- Commercial value --- p.7 / Chapter 1.6.1 --- Biodecolorization --- p.7 / Chapter 1.6.2 --- Bioconversion --- p.7 / Chapter 1.6.3 --- Biodegradation --- p.8 / Chapter 1.6.4 --- Indicator --- p.9 / Chapter 1.7 --- Cultivation --- p.9 / Chapter 1.8 --- Content --- p.10 / Chapter 1.9 --- Transformation --- p.11 / Chapter 1.10 --- Gene regulation for growth and fruiting body development --- p.13 / Chapter 1.11 --- Serial Analysis of Gene Expression --- p.18 / Chapter 1.11.1 --- Introduction --- p.18 / Chapter 1.11.2 --- Principles --- p.20 / Chapter 1.11.3 --- SAGE Application in Cancer and Immunology Studies --- p.22 / Chapter 1.11.4 --- Improvement of SAGE --- p.23 / Chapter 1.11.5 --- Bioinformatics --- p.24 / Chapter 1.12 --- DNA Microarray --- p.27 / Chapter 1.12.1 --- Introduction --- p.27 / Chapter 1.12.2 --- Application --- p.28 / Chapter 1.12.3 --- Method of cDNA Microarray --- p.28 / Chapter Chapter Two --- Serial Analysis of Gene Expression / Chapter 2.1 --- Introduction --- p.30 / Chapter 2.2 --- Material and Methods --- p.32 / Chapter 2.2.1 --- Mushroom cultivation and RNA extraction --- p.32 / Chapter 2.2.2 --- RNA Quality Estimation --- p.33 / Chapter 2.2.3 --- mRNA Isolation --- p.34 / Chapter 2.2.4 --- Serial Analysis of Gene Analysis (SAGE) --- p.34 / Chapter 2.2.4.1 --- Binding mRNA to magnetic beads for cDNA synthesis --- p.34 / Chapter 2.2.4.2 --- DNA synthesis verification --- p.35 / Chapter 2.2.4.3 --- NlaIII digestion --- p.36 / Chapter 2.2.4.4 --- NlaIII digestion verification --- p.36 / Chapter 2.2.4.5 --- Adapters ligation --- p.37 / Chapter 2.2.4.6 --- Cleaving with tagging enzyme --- p.37 / Chapter 2.2.4.7 --- Ditags creation --- p.38 / Chapter 2.2.4.8 --- PCR optimization and scale-up --- p.38 / Chapter 2.2.4.9 --- Polyacrylamide gel electrophoresis --- p.39 / Chapter 2.2.4.10 --- Eluting DNA from the gel --- p.40 / Chapter 2.2.4.11 --- NlaIII Cleavage and polyacrylamide gel electrophoresis --- p.40 / Chapter 2.2.4.12 --- Concatemers Ligation --- p.41 / Chapter 2.2.4.13 --- Cloning Concatemers into pZErO®-1 --- p.42 / Chapter 2.2.5 --- One Shot® TOP 10 Electrocomp´ёØ E. Coli transformation --- p.42 / Chapter 2.2.6 --- PCR Screening and Sequencing --- p.44 / Chapter 2.2.7 --- Sequence Analysis --- p.44 / Chapter 2.3 --- Results --- p.45 / Chapter 2.3.1 --- RNA Extraction --- p.45 / Chapter 2.3.2 --- cDNA Synthesis --- p.45 / Chapter 2.3.3 --- NlaIII digestion --- p.45 / Chapter 2.3.4 --- PCR amplification --- p.46 / Chapter 2.3.5 --- Gel-purification of the 100bp Ditags --- p.46 / Chapter 2.3.6 --- Isolation of the 26bp Ditags --- p.46 / Chapter 2.3.7 --- Concatemers Generation --- p.47 / Chapter 2.3.8 --- PCR Screening --- p.47 / Chapter 2.3.9 --- The Abundance and Identity of SAGE Tags --- p.48 / Chapter 2.4 --- Discussion --- p.86 / Chapter 2.4.1 --- RNA Extraction --- p.86 / Chapter 2.4.2 --- cDNA Synthesis and NlaIII digestion --- p.86 / Chapter 2.4.3 --- PCP amplification --- p.87 / Chapter 2.4.4 --- SAGE Tags Analysis --- p.88 / Chapter Chapter Three --- Microarray / Chapter 3.1 --- Introduction --- p.96 / Chapter 3.2 --- Materials and Methods --- p.97 / Chapter 3.2.1 --- Microarray chip preparation --- p.97 / Chapter 3.3.2 --- Sample preparation --- p.97 / Chapter 3.2.3 --- cDNA Synthesis and Sample labeling --- p.98 / Chapter 3.2.4 --- cDNA Purification --- p.99 / Chapter 3.2.5 --- cDNA analysis --- p.99 / Chapter 3.2.6 --- Array Hybridization --- p.102 / Chapter 3.2.6.1 --- Sample Preparation --- p.102 / Chapter 3.2.6.2 --- Hybridization Procedure --- p.102 / Chapter 3.2.7 --- Stringency Washes --- p.103 / Chapter 3.2.8 --- Detection with TSA --- p.103 / Chapter 3.2.9 --- Scanning and Analysis --- p.105 / Chapter 3.3 --- Results --- p.109 / Chapter 3.4 --- Discussion --- p.120 / Chapter Chapter Four --- Full Length Sequencing / Chapter 4.1 --- Introduction --- p.124 / Chapter 4.2 --- Material and Methods --- p.124 / Chapter 4.3 --- Results and Discussion --- p.125 / Chapter Chapter Five --- General Discussion --- p.149 / Appendix --- p.155 / References --- p.157

Duan Meijuan. / Thesis submitted in: August 2003. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 170-194). / Abstracts in English and Chinese. / Acknowledgements --- p.i / Abstract --- p.iii / Chinese abstract --- p.vi / List of Tables --- p.viii / List of Figures --- p.ix / List of Abbreviations --- p.xiv / Chapter Chapter 1. --- General Introduction --- p.1 / Chapter Chapter 2. --- Literature Review --- p.3 / Chapter 2.1 --- Hybrid rice: Genetics and breeding --- p.3 / Chapter 2.1.1 --- Classification in rice --- p.5 / Chapter 2.1.2 --- Heterosis in rice --- p.6 / Chapter 2.1.2.1 --- Performance of heterosis in rice --- p.6 / Chapter 2.1.2.2. --- Genetic mechanism of heterosis in rice --- p.7 / Chapter 2.1.3 --- Utilization of heterosis in rice --- p.9 / Chapter 2.2 --- Grain quality in rice --- p.11 / Chapter 2.2.1 --- Composition of rice grain quality --- p.11 / Chapter 2.2.1.1 --- Appearance quality --- p.11 / Chapter 2.2.1.2 --- Milling quality --- p.11 / Chapter 2.2.1.3 --- Nutritional quality --- p.12 / Chapter 2.2.1.4 --- Cooking and eating quality --- p.17 / Chapter 2.2.2 --- Genetic and breeding for high nutritional quality rice --- p.17 / Chapter 2.2.3 --- "Structural, physiological and biochemical changes during rice seed development" --- p.18 / Chapter 2.3 --- Molecular biological characteristics of rice seed storage protein --- p.20 / Chapter 2.3.1 --- "Property, classification and structure" --- p.20 / Chapter 2.3.1.1 --- Property and classification --- p.20 / Chapter 2.3.1.2 --- Composition structure --- p.20 / Chapter 2.3.1.2.1 --- Glutelin --- p.21 / Chapter 2.3.1.2.2 --- Prolamin --- p.22 / Chapter 2.3.1.2.3 --- Globulin and Albumin --- p.23 / Chapter 2.3.2 --- "Structure, expression and regulation of genes encoding rice seed storage protein genes" --- p.24 / Chapter 2.3.2.1 --- Structure --- p.24 / Chapter 2.3.2.1.1 --- Glutelin gene family --- p.24 / Chapter 2.3.2.1.2 --- Prolamin gene family --- p.26 / Chapter 2.3.2.1.3 --- Albumin and globulin gene family --- p.27 / Chapter 2.3.2.2 --- Expression of storage proteins in rice seed development --- p.28 / Chapter 2.3.2.3 --- Regulation of expression of seed storage protein genes --- p.29 / Chapter 2.3.2.3.1 --- Regulation at transcriptional level --- p.29 / Chapter 2.3.2.3.2 --- Regulation at post-transcriptional level --- p.31 / Chapter 2.3.2.3.3 --- Regulation at translational level --- p.31 / Chapter 2.3.3 --- "Synthesis, processing and deposition of rice seed storage proteins" --- p.32 / Chapter 2.4 --- Molecular characteristics of starch in rice grain --- p.34 / Chapter 2.4.1 --- Property of rice starch --- p.34 / Chapter 2.4.2 --- Starch biosynthesis in rice --- p.37 / Chapter 2.4.3 --- Enzymes involved in starch biosynthesis in rice --- p.39 / Chapter 2.4.3.1 --- ADP-glucose pyrophosphorylase (AGPase) --- p.39 / Chapter 2.4.3.2 --- Soluble starch synthase (SSS) --- p.41 / Chapter 2.4.3.3 --- Granular-bound starch synthase (GBSS) --- p.42 / Chapter 2.4.3.4 --- Starch branching enzyme (SBE) --- p.44 / Chapter 2.4.3.5 --- Starch debranching enzyme (SDE) --- p.46 / Chapter 2.5 --- Aspartate family amino acid biosynthetic pathway in rice --- p.48 / Chapter 2.5.1 --- Introduction --- p.48 / Chapter 2.5.2 --- Two key regulatory enzymes involved in lysine synthesis pathway --- p.50 / Chapter 2.5.2.1 --- Aspartate kinase (AK) --- p.50 / Chapter 2.5.2.2 --- Dihydrodipicolinate synthase (DHPS) --- p.51 / Chapter 2.5.2.3. --- Regulation of lysine and the other end products of AK pathway --- p.52 / Chapter 2.6 --- Proteomics in plants --- p.53 / Chapter 2.7 --- Approaches for grain quality improvement in rice --- p.56 / Chapter 2.7.1 --- Improvement of nutrition quality --- p.56 / Chapter 2.7.2 --- Improvement of eating and cooking quality --- p.57 / Chapter 2.8 --- Objectives of my project --- p.58 / Chapter Chapter 3. --- Materials and Methods --- p.60 / Chapter 3.1 --- Materials --- p.60 / Chapter 3.1.1 --- Chemicals --- p.60 / Chapter 3.1.2 --- Apparatus and commercial kits --- p.60 / Chapter 3.1.3 --- Plant materials --- p.61 / Chapter 3.1.4 --- DNA sequencing --- p.61 / Chapter 3.1.5 --- Software --- p.61 / Chapter 3.2 --- Methods --- p.62 / Chapter 3.2.1 --- Search for protein and DNA sequences of all genes --- p.62 / Chapter 3.2.1.1 --- Genes encoding rice glutelin family --- p.62 / Chapter 3.2.1.2 --- Genes encoding rice prolamin family --- p.63 / Chapter 3.2.1.3 --- Genes encoding rice albumin family --- p.63 / Chapter 3.2.1.4 --- Genes encoding rice globulin family --- p.64 / Chapter 3.2.1.5 --- Genes encoding rice starch synthesis enzymes --- p.64 / Chapter 3.2.2 --- Alignment of homologous DNA sequence between family member genes --- p.65 / Chapter 3.2.2.1 --- Seeds storage protein gene families of rice seeds --- p.65 / Chapter 3.2.2.2 --- Rice starch synthase gene families --- p.67 / Chapter 3.2.3 --- Primer design --- p.69 / Chapter 3.2.4 --- Collection of developing hybrid rice seeds --- p.71 / Chapter 3.2.5 --- Total RNA extraction --- p.75 / Chapter 3.2.6 --- Quantification of total RNA and determination of internal control --- p.75 / Chapter 3.2.7 --- RT-PCR (Reverse-transcription polymerase chain reaction) --- p.77 / Chapter 3.2.8 --- Northern blot analysis --- p.78 / Chapter 3.2.9 --- DNA sequencing --- p.79 / Chapter 3.2.10 --- Protein extraction --- p.80 / Chapter 3.2.10.1 --- Extraction of four kinds of storage proteins --- p.80 / Chapter 3.2.10.2 --- Extraction of the Wx protein --- p.81 / Chapter 3.2.11 --- Tricine SDS-PAGE --- p.82 / Chapter 3.2.12 --- "Determination of crude protein and amylose content in P64S,9311 and F1 hybrid" --- p.83 / Chapter 3.2.12.1 --- Determination of crude protein --- p.83 / Chapter 3.2.12.2 --- Determination of amylose content --- p.84 / Chapter 3.2.13 --- Two-dimension gel electrophoresis --- p.85 / Chapter 3.2.13.1 --- Clean up of protein sample for 2-D gel --- p.85 / Chapter 3.2.13.2 --- Quantification of protein samples --- p.86 / Chapter 3.2.13.3 --- First-dimension IEF (isoelectric focusing) --- p.86 / Chapter 3.2.13.4 --- IPG strips equilibration --- p.87 / Chapter 3.2.13.5 --- Second-dimension SDS PAGE --- p.87 / Chapter 3.2.13.6 --- Silver staining of 2-D gel --- p.88 / Chapter 3.2.14 --- MALDI-ToF mass spectrometry (Matrix Assisted Laser Desorption Ionization-Time of Flight) --- p.88 / Chapter 3.2.14.1 --- Sample destaining --- p.88 / Chapter 3.2.14.2 --- In-gel digestion with trypsin enzyme --- p.89 / Chapter 3.2.14.3 --- Desalination of the digested sample with Zip Tip --- p.90 / Chapter 3.2.14.4 --- Mass spectrometry --- p.90 / Chapter Chapter 4. --- Results --- p.91 / Chapter 4.1 --- Quantification of the total RNA from developing seeds at different developingstages --- p.91 / Chapter 4.2 --- Determination of internal control --- p.92 / Chapter 4.3 --- DNA sequence analysis --- p.95 / Chapter 4.4 --- Profiling the expression of genes encoding rice seed storage proteins --- p.97 / Chapter 4.4.1 --- The glutelin genes --- p.97 / Chapter 4.4.1.1 --- The Gtl (GluA-2) gene --- p.100 / Chapter 4.4.1.2 --- The Gt2 (GluA-1) gene --- p.100 / Chapter 4.4.1.3 --- The Gt3 (GluA-3) gene --- p.101 / Chapter 4.4.1.4 --- Comparison of the expression profiles of GluA subfamily genes --- p.101 / Chapter 4.4.1.5 --- The GluB-1 gene --- p.101 / Chapter 4.4.1.6 --- The GluB-2 gene --- p.102 / Chapter 4.4.1.7 --- The GluB-4 gene --- p.102 / Chapter 4.4.1.8 --- Comparing of the expression profiles of GluB subfamily genes --- p.102 / Chapter 4.4.1.9 --- Profiling the expression of glutilin family genes in developing hybrid rice seeds --- p.103 / Chapter 4.4.1.10 --- Profiling glutelin gene expression in developing seeds at protein level --- p.103 / Chapter 4.4.2 --- Profiling the expression of genes encoding prolamin familyin developing hybrid rice seeds --- p.105 / Chapter 4.4.2.1 --- The 10-kDa prolamin gene --- p.105 / Chapter 4.4.2.2 --- The RP5 gene --- p.108 / Chapter 4.4.2.3 --- The RP6 gene --- p.108 / Chapter 4.4.2.4 --- The Prol 7 gene --- p.109 / Chapter 4.4.2.5 --- The Prol 14 gene --- p.109 / Chapter 4.4.2.6 --- The Prol 17 gene --- p.109 / Chapter 4.4.2.7 --- Expression profiles of prolamin family genes --- p.110 / Chapter 4.4.2.8 --- Expression profiles of prolamin genes in developing hybrid rice seeds at protein level --- p.111 / Chapter 4.4.3 --- Profiling the expression of genes encoding globulin familyin developing hybrid rice seed --- p.113 / Chapter 4.4.3.1 --- The 26-kDa globulin (alpha-globulin) gene --- p.113 / Chapter 4.4.3.2 --- The globulin 1 gene --- p.113 / Chapter 4.4.3.3 --- The globulin 2 gene --- p.115 / Chapter 4.4.3.4 --- The Low molecular weight (LMW) globulin gene --- p.115 / Chapter 4.4.3.5 --- Profiling the expression of the globulin family genes --- p.115 / Chapter 4.4.3.6 --- Expression profiles of globulin proteins in developing hybrid rice seeds at protein level --- p.117 / Chapter 4.4.4 --- Profiling the expression of genes encoding rice albumin familyin developing hybrid rice seeds --- p.118 / Chapter 4.4.4.1 --- The RA5 gene --- p.118 / Chapter 4.4.4.2 --- The RA 14 gene --- p.119 / Chapter 4.4.4.3 --- The RA 17 gene --- p.119 / Chapter 4.4.4.4 --- Profiling the expression of the albumin family genes --- p.121 / Chapter 4.4.4.5 --- Albumin gene expression in developing hybrid rice seeds at protein level --- p.121 / Chapter 4.4.5 --- Comparison of expression pattern of all genes encoding rice seed storage proteins in developing hybrid rice seeds --- p.123 / Chapter 4.4.6 --- Profiling the total proteins in developing hybrid rice seeds --- p.126 / Chapter 4.5 --- Profiling the expression of genes encoding rice starch synthasesin developing hybrid rice seeds --- p.127 / Chapter 4.5.1 --- Rice ADP-glucose pyrophosphorylase (AGPase) genes --- p.127 / Chapter 4.5.1.1 --- The AGPase large subunit gene --- p.127 / Chapter 4.5.1.2 --- The AGPase small subunit gene --- p.127 / Chapter 4.5.2 --- "The Wx (Granule bound starch synthase, GBSS) gene" --- p.129 / Chapter 4.5.3 --- Genes encoding rice SSS (Soluble starch synthase) family --- p.132 / Chapter 4.5.3.1 --- The SSS1 gene --- p.132 / Chapter 4.5.3.2 --- The SSS II-1 gene --- p.132 / Chapter 4.5.3.3 --- The SSS II-2 gene --- p.132 / Chapter 4.5.3.4 --- The SSS II-3 gene --- p.135 / Chapter 4.5.3.5 --- The SSS III-2 gene --- p.135 / Chapter 4.5.3.6 --- The SSS IV-1 gene --- p.135 / Chapter 4.5.3.7 --- The SSS IV-2 gene --- p.135 / Chapter 4.5.3.8 --- Profiling the expression of SSS family genes --- p.136 / Chapter 4.5.4 --- Genes encoding rice starch branching enzyme (SBE) family --- p.138 / Chapter 4.5.4.1 --- The SBE-1 gene --- p.138 / Chapter 4.5.4.2 --- The SBE-3 gene --- p.138 / Chapter 4.5.4.3 --- The SBE-4 gene --- p.138 / Chapter 4.5.4.4 --- Profiling the expression of SBE family genes --- p.140 / Chapter 4.5.5 --- Genes encoding rice starch debranching enzyme (SDE) family --- p.141 / Chapter 4.5.5.1 --- The isoamylase gene --- p.141 / Chapter 4.5.5.2 --- The pullulanase gene --- p.141 / Chapter 4.5.5.3 --- Difference between isoamylose and pullulanase --- p.141 / Chapter 4.5.6 --- Comparison of the expression patterns of genes encoding the enzymes involved in starch synthesis in developing hybrid rice seeds --- p.143 / Chapter 4.6 --- Profiling the expression of genes encoding aspartate family amino acid biosynthetic pathway in rice in developing hybrid rice seeds --- p.145 / Chapter 4.6.1 --- Rice AK (aspartate kinase) gene --- p.145 / Chapter 4.6.2 --- The DHPS gene --- p.145 / Chapter 4.7 --- Two-dimension gel electrophoresis and MALDI-ToF seed proteins analysis of rice --- p.147 / Chapter Chapter 5. --- Discussion --- p.152 / Chapter 5.1 --- Super hybrid rice as experimental material and its significance --- p.152 / Chapter 5.2 --- RT-PCR and northern blotting as methods to profile gene expression --- p.153 / Chapter 5.3 --- Regulation of genes related to nutritional quality in rice --- p.155 / Chapter 5.3.1 --- Storage protein genes --- p.155 / Chapter 5.3.2 --- Lysine synthesis enzymes --- p.158 / Chapter 5.4 --- Regulation of genes related to cooking and eating quality in rice --- p.159 / Chapter 5.5 --- Heredity of genes expression in F1 hybrid --- p.161 / Chapter 5.6 --- Application of 2-D gel electrophoresis --- p.162 / Chapter 5.7 --- Future perspectives --- p.163 / Chapter Chapter 6. --- Conclusion --- p.164 / References --- p.170

Zhang Junjun. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 147-160). / Abstracts in English and Chinese. / Thesis Committee --- p.i / Statement from Author --- p.ii / Acknowledgements --- p.iii / Abstract --- p.v / 摘要 --- p.vii / Table of Contents --- p.ix / List of Tables --- p.xiv / List of Figures --- p.xvi / List of Abbreviations --- p.xviii / Chapter Chapter 1 --- General Introduction and Literature Review --- p.1 / Chapter 1.1 --- General Introduction --- p.1 / Chapter 1.2 --- Literature Review --- p.2 / Chapter 1.2.1 --- Rice as a Model Monocot Cereal --- p.2 / Chapter 1.2.1.1 --- Genome Size --- p.3 / Chapter 1.2.1.2 --- Synteny --- p.3 / Chapter 1.2.1.3 --- Well-mapped Genome --- p.4 / Chapter 1.2.1.4 --- Amenable to Transformation --- p.4 / Chapter 1.2.2 --- Rice Genome Sequencing --- p.4 / Chapter 1.2.3 --- Rice Functional Genomics --- p.6 / Chapter 1.2.3.1 --- International Collaboration on Rice Functional Genomics --- p.6 / Chapter 1.2.3.2 --- Recent Progress on Rice Functional Genomics in China --- p.7 / Chapter 1.2.4 --- Large-scale Expressed Sequenced Tag (EST) Analysis --- p.8 / Chapter 1.2.4.1 --- Introduction --- p.8 / Chapter 1.2.4.2 --- Large-scale EST Analysis vs. SAGE and MPSS: Advantages and Shortcomings --- p.12 / Chapter 1.2.4.3 --- Large-scale Rice EST analysis --- p.14 / Chapter 1.2.4.4 --- Seed Specific Large-scale EST Analysis --- p.16 / Chapter 1.2.4.5 --- Bioinformatics Tools Involved --- p.17 / Chapter 1.2.5 --- Profiling Gene Expression using cDNA Microarray --- p.20 / Chapter 1.2.5.1 --- Introduction --- p.20 / Chapter 1.2.5.2 --- Advantages of cDNA Microarray in Gene Expression Profiling --- p.22 / Chapter 1.2.5.3 --- Profiling Gene Expression by cDNA Microarrays --- p.24 / Chapter 1.2.5.4 --- Profiling Seed-specific Gene Expression --- p.26 / Chapter 1.2.6 --- Overview of Current Information on the Biosynthesis and Gene Regulation of Major Components during Rice Seed Formation --- p.30 / Chapter 1.2.6.1 --- Starch --- p.30 / Chapter 1.2.6.2 --- Storage Proteins --- p.38 / Chapter 1.2.6.3 --- Lipids --- p.42 / Chapter Chapter 2 --- Materials and Methods --- p.44 / Chapter 2.1 --- Materials --- p.44 / Chapter 2.1.1 --- Chemicals --- p.44 / Chapter 2.1.2 --- Facilities and Instruments --- p.45 / Chapter 2.1.3 --- Commercial Kits --- p.46 / Chapter 2.1.4 --- Oligonucleotides --- p.47 / Chapter 2.1.5 --- Plant Materials --- p.50 / Chapter 2.1.6 --- Bacterial Strains --- p.52 / Chapter 2.2 --- Methods --- p.52 / Chapter 2.2.1 --- mRNA Isolation --- p.52 / Chapter 2.2.2 --- Construction of cDNA Library --- p.53 / Chapter 2.2.2.1 --- Construction of cDNA Library in Phage --- p.53 / Chapter 2.2.2.2 --- Construction of cDNA Library in E.coli --- p.56 / Chapter 2.2.3 --- Screening of cDNA Library --- p.62 / Chapter 2.2.3.1 --- In situ Hybridization --- p.62 / Chapter 2.2.3.2 --- Probe Synthesis --- p.63 / Chapter 2.2.3.3 --- Hybridization and Detection --- p.64 / Chapter 2.2.4 --- Single-pass Sequencing of cDNAs --- p.65 / Chapter 2.2.5 --- Sequence Analysis --- p.67 / Chapter 2.2.6 --- BLAST Search --- p.71 / Chapter 2.2.7 --- Contig Analysis --- p.71 / Chapter 2.2.8 --- Database Management --- p.72 / Chapter 2.2.9 --- Selection Criteria for ESTs in Different Pathways --- p.72 / Chapter 2.2.10 --- Construction of Super Hybrid Rice Seed cDNA Microarray --- p.75 / Chapter 2.2.11 --- "Probe Synthesis, Microarray Hybridization and Detection" --- p.75 / Chapter 2.2.12 --- Quality Test of External Control --- p.78 / Chapter 2.2.13 --- Scanning of Microarray --- p.79 / Chapter 2.2.14 --- Analysis of Microarray Data --- p.79 / Chapter 2.2.15 --- Normalization --- p.79 / Chapter 2.2.16 --- Northern Blot Analysis --- p.80 / Chapter Chapter 3 --- Results and Discussion --- p.82 / Chapter 3.1 --- cDNA Libraries --- p.82 / Chapter 3.1.1 --- cDNA Library in Phage --- p.82 / Chapter 3.1.2 --- cDNA Library in E.coli --- p.84 / Chapter 3.2 --- Statistic Data of Screening and Single-pass Sequencing --- p.87 / Chapter 3.3 --- Genes Expressing in Developing Rice Seeds --- p.89 / Chapter 3.4 --- Novel ESTs in the Seed EST Set --- p.94 / Chapter 3.5 --- Unique Genes in the EST Set from the Phage cDNA Library --- p.94 / Chapter 3.6 --- Mapping ESTs onto Rice Genome --- p.95 / Chapter 3.7 --- The Most Abundant Transcripts Reflected by EST Redundancy --- p.98 / Chapter 3.8 --- ESTs and Representive Genes in Different Pathways --- p.102 / Chapter 3.9 --- ESTs Involved in Carbon Flows --- p.106 / Chapter 3.10 --- Microarray Configuration and Contents --- p.109 / Chapter 3.11 --- Quantification of Total RNA from Seeds at 5 Developing Stages --- p.110 / Chapter 3.12 --- Quality Test of External Control --- p.111 / Chapter 3.13 --- Scanning of Array Hybridization --- p.113 / Chapter 3.14 --- Reproducibility of Technical Duplicates --- p.116 / Chapter 3.15 --- Summary of Gene Expression Profiles in Developing Super Hybrid Rice Seeds --- p.117 / Chapter 3.16 --- Expression Profiles of Major Storage Compounds in Rice Seeds --- p.119 / Chapter 3.16.1 --- Expression profiles of storage protein genes --- p.119 / Chapter 3.16.2 --- Expression Profiles of Starch Synthesis Related Genes --- p.122 / Chapter 3.16.3 --- Expression Profiles of Lipid Synthesis Related Genes --- p.124 / Chapter 3.16.4 --- General Expression Patterns of Major Storage Compounds --- p.126 / Chapter 3.18 --- General Discussion --- p.131 / Chapter 3.18.1 --- Cautions when Interpreting the EST Data --- p.131 / Chapter 3.18.2 --- Comparison of EST Data Sets of Developing Seeds between Arabidopsis and Rice --- p.131 / Chapter 3.18.3 --- Comparison of cDNA Library Construction by Two Methods --- p.135 / Chapter 3.18.4 --- Comparison of Different Normalization Methods --- p.138 / Chapter 3.18.5 --- Comparison between Microarray and Northern Data --- p.140 / Chapter Chapter 4 --- Conclusion --- p.142 / Chapter Chapter 5 --- Future Perspectives --- p.145 / Reference Lists --- p.147