by Sham Lok To. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 143-171). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgement --- p.iv / Abbreviations --- p.v / Table of contents --- p.vi / List of figures --- p.ix / List of tables --- p.xi / Chapter Chapter One --- Literature Review --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.1.1 --- About L. edodes --- p.3 / Chapter 1.1.2 --- Nutritional and medicinal values of L. edodes --- p.4 / Chapter 1.1.3 --- Life cycle of L. edodes --- p.6 / Chapter 1.1.4 --- Environmental factors affecting fruiting body formation in L. edodes --- p.6 / Chapter 1.2 --- Molecular mechanisms of fruiting body development in L. edodes --- p.8 / Chapter 1.2.1 --- Expression profiling and identification of differentially expressed genes during fruiting --- p.8 / Chapter 1.2.2 --- Changing in membrane structure --- p.11 / Chapter 1.2.3 --- The signal transduction cascade --- p.12 / Chapter 1.3 --- Transformation in L. edodes and in other fungi --- p.14 / Chapter 1.3.1 --- Transformation of L. edodes --- p.14 / Chapter 1.3.2 --- Transformation in other fungi --- p.17 / Chapter 1.4 --- Bioinformatics tools for comparative promoter analysis --- p.22 / Chapter 1.5 --- Objectives and significance --- p.26 / Chapter Chapter Two --- Promoter analysis of differentially expressed genes (DEGs) in the fruiting body development in L. edodes --- p.27 / Chapter 2.1 --- Introduction --- p.27 / Chapter 2.2 --- Materials and methods --- p.29 / Chapter 2.2.1 --- Strains and cultivation conditions --- p.29 / Chapter 2.2.2 --- Genome walking of the 5' flanking region of the DEGs --- p.29 / Chapter 2.2.3 --- Annealing Control Primed (ACP) PCR --- p.31 / Chapter 2.2.4 --- Construction of genomic DNA library --- p.36 / Chapter 2.2.5 --- Nested PCR to amplify the target sequences --- p.37 / Chapter 2.2.6 --- Cloning and sequencing of the 5' flanking region --- p.38 / Chapter 2.2.7 --- Determination of transcription start site by the Neural Network algorithm --- p.39 / Chapter 2.2.8 --- Identification of putative transcription factor binding sites --- p.40 / Chapter 2.3 --- Results --- p.41 / Chapter 2.3.1 --- Construction of adaptor linked template for genome walking --- p.41 / Chapter 2.3.2 --- Sequence analysis and quality control --- p.41 / Chapter 2.3.3 --- Comparison of various methods in genome walking --- p.42 / Chapter 2.3.4 --- Promoter analysis --- p.42 / Chapter 2.4 --- Discussion --- p.58 / Chapter Chapter Three --- In-silico analysis of transcription factor binding sites and identification transcription factors expressed in L. edodes --- p.64 / Chapter 3.1 --- Introduction --- p.64 / Chapter 3.2 --- Material and methods --- p.67 / Chapter 3.2.1 --- Sequence manipulation and extraction of homologous ESTs from C. cinereus --- p.67 / Chapter 3.2.2 --- Extraction of 5' flanking region of the corresponding ESTs and promoter prediction --- p.67 / Chapter 3.2.3 --- Positional cloning of mating type factor A --- p.68 / Chapter 3.3 --- Results --- p.70 / Chapter 3.3.1 --- Sequence extraction and manipulation --- p.70 / Chapter 3.3.2 --- In-silico analysis of transcription factor binding sites in C. cinereus . --- p.70 / Chapter 3.3.3 --- Comparison of putative TFBS between L. edodes and C. cinereus --- p.71 / Chapter 3.3.4 --- Identification of transcription factors in L. edodes by positional cloning --- p.71 / Chapter 3.4 --- Discussion --- p.85 / Chapter Chapter Four --- Identification，expression profiling and promoter analysis of hydrophobin genes --- p.91 / Chapter 4.1 --- Introduction --- p.91 / Chapter 4.2 --- Material and methods --- p.92 / Chapter 4.2.1 --- Clustering and grouping of the hydrophobin ESTs --- p.92 / Chapter 4.2.2 --- Identification of the consensus sequences of the hydrophobin groups --- p.93 / Chapter 4.2.3 --- RNA Sources and Preparation --- p.93 / Chapter 4.2.4 --- Expression profiling of hydrophobin genes by RT-PCR --- p.95 / Chapter 4.2.5 --- Promoter cloning and analysis of hydrophobin genes --- p.95 / Chapter 4.3 --- Results --- p.97 / Chapter 4.3.1 --- Isolation and characterization of four newly found hydrophobin genes --- p.97 / Chapter 4.3.2 --- Expression levels of hydrophobins --- p.100 / Chapter 4.3.3 --- Promoter sequencing of the hydrophobins --- p.103 / Chapter 4.4 --- Discussion --- p.103 / Chapter Chapter Five --- Transformation of L. edodes --- p.110 / Chapter 5.1 --- Introduction --- p.110 / Chapter 5.2 --- Materials and methods --- p.112 / Chapter 5.2.1 --- Vectors and primers design --- p.112 / Chapter 5.2.2 --- Maxi-preparation of plasmids --- p.112 / Chapter 5.2.3 --- Cultural condition and optimization of protoplasts release --- p.114 / Chapter 5.2.4 --- PEG mediated transformation --- p.115 / Chapter 5.2.5 --- Electroporation mediated transformation --- p.116 / Chapter 5.2.6 --- PCR screening of regenerated transformant --- p.116 / Chapter 5.2.7 --- Particle bombardment --- p.117 / Chapter 5.3 --- Results --- p.121 / Chapter 5.4 --- Discussion --- p.128 / Chapter Chapter Six --- General discussions --- p.132 / References --- p.143

Lee Ming Tsung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 160-177). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.v / Abbreviations --- p.vi / Table of contents --- p.vii / List of Figures --- p.xii / List of Tables --- p.xiv / Chapter Chapter 1 --- Literature Review --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Nutritional values --- p.2 / Chapter 1.3 --- Medicinal values --- p.3 / Chapter 1.3.1 --- Anti-tumor effect --- p.3 / Chapter 1.3.2 --- Anti-viral and anti-caries effect --- p.4 / Chapter 1.3.3 --- Immunopotentiating effect --- p.4 / Chapter 1.3.4 --- Hypocholesterolaemic effect --- p.5 / Chapter 1.4 --- Life cycle and morphology --- p.6 / Chapter 1.5 --- Growth requirements --- p.9 / Chapter 1.5.1 --- Nutritional factors --- p.9 / Chapter 1.5.2 --- Physical and chemical factors --- p.10 / Chapter 1.6 --- Application of L. edodes --- p.12 / Chapter 1.7 --- Endocytosis --- p.13 / Chapter 1.7.1 --- Different types of endocytosis --- p.13 / Chapter 1.7.1.1 --- Phagocytosis --- p.14 / Chapter 1.7.1.2 --- Pinocytosis --- p.15 / Chapter 1.7.1.3 --- Receptor-mediated endocytosis --- p.15 / Chapter 1.7.2 --- The Endocytic Pathway --- p.17 / Chapter 1.7.3 --- Endocytosis in fungi --- p.20 / Chapter 1.7.4 --- Rab GTPases --- p.21 / Chapter 1.7.4.1 --- Control of the active and inactive state of Rab proteins --- p.22 / Chapter 1.7.4.2 --- Regulation of docking and fusion of membrane in endosomal trafficking --- p.23 / Chapter 1.7.4.3 --- Rab7 GTPase --- p.26 / Chapter 1.8 --- Aims of the project --- p.28 / Chapter Chapter 2 --- Protein-protein Interaction Study of Le.Rab7 by in vivo and in vitro Interaction Assay --- p.29 / Chapter 2.1 --- Introduction --- p.29 / Chapter 2.2 --- Materials and Methods --- p.36 / Chapter 2.2.1 --- Yeast two-hybrid screening --- p.36 / Chapter 2.2.1.1 --- Confirmation of the clones Le.Rab7-pGBK.T7 --- p.36 / Chapter 2.2.1.1.1 --- Bacterial transformation --- p.36 / Chapter 2.2.1.1.2 --- PCR screening for positive transformants --- p.38 / Chapter 2.2.1.1.3 --- Plasmid preparation and confirmation of transformants --- p.38 / Chapter 2.2.1.1.4 --- Sequencing --- p.39 / Chapter 2.2.1.2 --- Confirmation of Le.Rab7 protein expression in yeast --- p.40 / Chapter 2.2.1.2.1 --- Yeast transformation --- p.40 / Chapter 2.2.1.2.2 --- Yeast protein extraction --- p.40 / Chapter 2.2.1.2.3 --- Western Blotting --- p.41 / Chapter 2.2.1.3 --- Yeast Two-hybrid screening by Yeast-mating --- p.42 / Chapter 2.2.1.4 --- Identification of Preys --- p.44 / Chapter 2.2.1.4.1 --- PCR screening for clones grown on plates --- p.44 / Chapter 2.2.1.4.2 --- Colony lift filter assay --- p.45 / Chapter 2.2.1.4.3 --- Sequencing --- p.47 / Chapter 2.2.1.5 --- Confirmation of interaction by Co-transformation assay --- p.47 / Chapter 2.2.1.5.1 --- Plasmid preparation of positive clones --- p.47 / Chapter 2.2.1.5.2 --- Transformation and bacterial plasmid preparation --- p.48 / Chapter 2.2.1.5.3 --- Yeast two-hybrid screening by co-transformation --- p.48 / Chapter 2.2.1.5.4 --- Colony lift filter assay --- p.50 / Chapter 2.2.2 --- Rapid Amplification of cDNA 5'ends --- p.51 / Chapter 2.2.2.1 --- RNA preparation --- p.51 / Chapter 2.2.2.1.1 --- Strains and culture conditions --- p.51 / Chapter 2.2.2.1.2 --- RNA extraction --- p.51 / Chapter 2.2.2.2 --- 5' RACE --- p.52 / Chapter 2.2.2.2.1 --- RNA processing --- p.52 / Chapter 2.2.2.2.2 --- Reverse transcription --- p.53 / Chapter 2.2.2.2.3 --- Nested PCR for 5'RLM-RACE --- p.53 / Chapter 2.2.2.3 --- "Gel analysis of products, TA cloning of RACE product and sequencing" --- p.54 / Chapter 2.2.2.4 --- Cloning of full-length Le.Rab5 --- p.54 / Chapter 2.2.3 --- In vitro protein-protein interaction assay --- p.55 / Chapter 2.2.3.1 --- Plasmid extraction from E.coli --- p.55 / Chapter 2.2.3.2 --- In vitro translation --- p.56 / Chapter 2.2.3.3 --- In vitro co-immunoprecipitation --- p.56 / Chapter 2.3 --- Results --- p.57 / Chapter 2.3.1 --- Yeast two-hybrid analysis by yeast mating assay --- p.57 / Chapter 2.2.1.1 --- Confirmation of the clones Le.Ra67-pGBKT7 --- p.57 / Chapter 2.3.1.1.1 --- PCR screening for positive transformants --- p.57 / Chapter 2.3.1.1.2 --- Plasmid preparation and confirmation of transformants --- p.58 / Chapter 2.3.1.1.3 --- Sequencing --- p.59 / Chapter 2.2.1.2 --- Confirmation of protein expression in yeast --- p.60 / Chapter 2.3.1.2.1 --- Yeast transformation --- p.60 / Chapter 2.3.1.2.2 --- SDS-PAGE and Western blotting of Le.Rab7 in yeast --- p.61 / Chapter 2.2.1.3 --- Yeast two-hybrid screening by yeast mating assay --- p.62 / Chapter 2.2.1.4 --- Identification of Preys --- p.63 / Chapter 2.3.1.4.1 --- PCR screening for clones grown on plates --- p.63 / Chapter 2.3.1.4.2 --- Colony lift assay --- p.65 / Chapter 2.3.1.4.3 --- Sequencing --- p.67 / Chapter 2.3.2 --- Confirmation of interactions by co-transformation assay --- p.70 / Chapter 2.2.2.1 --- Yeast two-hybrid analysis by co-transformation assay --- p.70 / Chapter 2.2.2.2 --- Colony lift filter assay --- p.70 / Chapter 2.2.2.3 --- Selection of prey plasmids for in vitro binding assay --- p.72 / Chapter 2.3.3 --- Rapid amplification of cDNA ends (RACE) --- p.76 / Chapter 2.2.3.1 --- TA cloning of RACE product and sequencing --- p.76 / Chapter 2.2.3.2 --- Cloning of full-length Le.Rab5 --- p.79 / Chapter 2.3.4 --- In vitro protein-protein interaction assay --- p.80 / Chapter 2.4 --- Discussion --- p.82 / Chapter Chapter 3 --- Temporal and Spatial expression of Le.Rab7，Le.Rab5 and Le.RACKl --- p.87 / Chapter 3.1 --- Introduction --- p.87 / Chapter 3.2 --- Materials and Methods --- p.93 / Chapter 3.2.1 --- Northern blot analysis --- p.93 / Chapter 3.2.1.1 --- RNA fractionation by formaldehyde gel electrophoresis --- p.93 / Chapter 3.2.1.2 --- Northern blotting --- p.94 / Chapter 3.2.1.2.1 --- Transfer of RNAs --- p.94 / Chapter 3.2.1.2.2 --- Probe preparation --- p.95 / Chapter 3.2.1.2.3 --- "Hybridization, Stringency washes and Signal detection" --- p.96 / Chapter 3.2.2 --- Quantitative RT-PCR --- p.97 / Chapter 3.2.2.1 --- cDNA synthesis from different developmental stages --- p.97 / Chapter 3.2.2.1.1 --- RNA preparation extraction --- p.97 / Chapter 3.2.2.1.2 --- DNase I treatment --- p.97 / Chapter 3.2.2.1.3 --- Reverse transcription --- p.98 / Chapter 3.2.2.2 --- Real time PCR --- p.98 / Chapter 3.2.2.2.1 --- Primer design and verification --- p.98 / Chapter 3.2.2.2.2 --- Real time PCR reaction and data analysis --- p.100 / Chapter 3.2.3 --- In situ RNA-RNA hybridization --- p.101 / Chapter 3.2.3.1 --- Preparation of samples and probes --- p.101 / Chapter 3.2.3.1.1 --- Tissue preparation --- p.101 / Chapter 3.2.3.1.2 --- RNA probe synthesis --- p.101 / Chapter 3.2.3.2 --- Hybridization and Signal development --- p.102 / Chapter 3.2.3.3 --- Image viewing --- p.103 / Chapter 3.3 --- Results --- p.105 / Chapter 3.3.1 --- Northern blot analysis --- p.105 / Chapter 3.3.2 --- Quantitative RT-PCR assays --- p.109 / Chapter 3.3.3 --- In situ RNA-RNA hybridization --- p.113 / Chapter 3.4 --- Discussion --- p.119 / Chapter Chapter 4 --- Existence of endocytosis and Protein localization of Le.Rab7 in L. edodes --- p.123 / Chapter 4.1 --- Introduction --- p.123 / Chapter 4.2 --- Materials and Methods --- p.127 / Chapter 4.2.1 --- Tracing the endocytie pathway using FM4-64 dye --- p.127 / Chapter 4.2.1.1 --- Strains and culture conditions --- p.127 / Chapter 4.2.1.2 --- FM4-64 internalization in mycelium and gill tissue of L. edodes --- p.127 / Chapter 4.2.2 --- Drug treatment effect on the internalization of FM4-64 dye --- p.128 / Chapter 4.2.3 --- Double labeling with AM4-64 and anti-Le.Rab7 antibody --- p.129 / Chapter 4.2.3.1 --- Synthesis of Le.Rab7 antibody --- p.129 / Chapter 4.2.3.1.1 --- Customization of Le.Rab7 antiserum --- p.129 / Chapter 4.2.3.1.2 --- Validation of anti-Le.Rab7 polyclonal antiserum --- p.129 / Chapter 4.2.3.2 --- Double immunofluorescence labeling --- p.130 / Chapter 4.2.4 --- Immunohistochemistry of young and mature fruiting body --- p.131 / Chapter 4.2.4.1 --- Tissue preparation --- p.131 / Chapter 4.2.4.2 --- Immunohistochemical staining --- p.132 / Chapter 4.2.4.3 --- Image viewing --- p.133 / Chapter 4.3 --- Results --- p.134 / Chapter 4.3.1 --- Presence of endocytosis in L .edodes --- p.134 / Chapter 4.3.2 --- Validation of active transport of FM4-64 --- p.137 / Chapter 4.3.3 --- Dye internalization at specific structures in L. edodes --- p.138 / Chapter 4.3.4 --- Presence of Le.Rab7 protein in the endosomal structures along the endocytic pathway --- p.142 / Chapter 4.3.5 --- Presence of Le.Rab7 protein in the pre- and hymenophore of fruiting body --- p.145 / Chapter 4.4 --- Discussion --- p.148 / Chapter Chapter 5 --- General discussion --- p.152 / References --- p.160

Chum Wing Yan Winnie. / "August 2006." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 190-223). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese.

by Shih Sheung Mei. / Thesis submitted in: July 2003. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 206-215). / Abstracts in English and Chinese. / Abstract --- p.ii / Achnoledgements --- p.vi / Abbreviations --- p.viii / List of contents --- p.viv / List of tables --- p.xiii / List of figures --- p.xv / Chapter Chapter One --- Literature Review / Chapter 1.1 --- Introducation of Lentinula edodes --- p.1 / Chapter 1.1.1 --- Life cycle of Basidiomycete --- p.1 / Chapter 1.1.2 --- Differentially Expressed Genes in stages of Lentinula edodes --- p.3 / Chapter 1.2 --- Relationship of Monokaryons and Dikaryons in Basidiomycetes --- p.4 / Chapter 1.2.1 --- Mating Type Gene in Filamentous Fungi --- p.4 / Chapter 1.2.3 --- Dikaryon Formation and Homeodomain Proteins --- p.6 / Chapter 1.2.4 --- Clamp Connection formation in Dikaryon --- p.9 / Chapter 1.3 --- Stuctural Protein of Mushroom --- p.11 / Chapter 1.3.1 --- Hydrophobin --- p.11 / Chapter 1.3.1.1 --- General Introduction --- p.11 / Chapter 1.3.1.2 --- Structure of hydrophobin --- p.11 / Chapter 1.3.1.3 --- Formation of Disulphide bonds and Glycosylation --- p.12 / Chapter 1.3.1.4 --- Functions of Hydrophobins --- p.13 / Chapter 1.4 --- Genomics of filamentous fungi --- p.15 / Chapter 1.5 --- Genetic analysis of filamentous fungi --- p.18 / Chapter 1.6 --- Objectives of the Project --- p.20 / Chapter Chapter Two --- Identification of Differentially Expressed Genes in Dikaryons of Lentinula edodes by Microarray of Primordium Expressed Sequence Tags / Chapter 2.1 --- Introduction --- p.23 / Chapter 2.2 --- Materials and Methods --- p.27 / Chapter 2.2.1 --- Construction of EST database --- p.27 / Chapter 2.2.2 --- Construction of EST Microarray cDNA gene-chip --- p.27 / Chapter 2.2.2.1 --- Amplification of the primordium EST clones --- p.27 / Chapter 2.2.2.2 --- Purification of the amplified EST clones --- p.28 / Chapter 2.2.2.3 --- Spotting of the amplified EST clones onto chips --- p.29 / Chapter 2.2.3 --- Screening of the Differentially Expressed Genes in Dikaryons by Primordium Microarray --- p.31 / Chapter 2.2.3.1 --- Mycelium Cultivation and Preparation of Total RNA --- p.31 / Chapter 2.2.3.2 --- cDNA synthesis and labeling --- p.32 / Chapter 2.2.3.3 --- cDNA purification --- p.33 / Chapter 2.2.3.4 --- Probe Storage Conditions --- p.34 / Chapter 2.2.3.5 --- cDNA analysis --- p.35 / Chapter 2.2.3.6 --- Microarray hybridization --- p.37 / Chapter 2.2.3.7 --- Stringency washes --- p.39 / Chapter 2.2.3.8 --- Detection with TSA --- p.39 / Chapter 2.2.3.9 --- Microarray scanning and data anlysis --- p.41 / Chapter 2.3 --- Results --- p.45 / Chapter 2.3.1 --- Amplification of primordium ESTs --- p.45 / Chapter 2.3.2 --- Purification of PCR products --- p.45 / Chapter 2.3.3 --- Data Analysis of Microarray Data --- p.47 / Chapter 2.3.3.1 --- Generation of Primordium EST Microarray Image for analysis --- p.47 / Chapter 2.3.3.2 --- Normalization of the Data --- p.49 / Chapter 2.3.3.3. --- Transciption Profile of Dikaryon compared with Monokaryon --- p.79 / Chapter 2.3.3.4. --- Differentially Expression of Dikaryon L54 --- p.80 / Chapter 2.4 --- Discussion --- p.85 / Chapter Chapter Three --- Enrichment of Genes with Differentially Expression in Dikaryons by Construction of Full-length Subtractive Library / Chapter 3.1 --- Introduction of Subtraction Cloning --- p.93 / Chapter 3.2 --- Materials and Methods --- p.97 / Chapter 3.2.1 --- Construction of Full-length Dikaryotic Subtractive library --- p.97 / Chapter 3.2.1.1 --- Isolation of PolyA+ mRNA of Dikaryon for Subtraction --- p.97 / Chapter 3.2.1.2 --- Enrichment of Differentially Expressed Genes in Dikaryon L54 by Subtraction with Monokaryons A and B --- p.99 / Chapter 3.2.1.3 --- First-Strand cDNA Synthesis --- p.102 / Chapter 3.2.1.4 --- cDNA Amplification by Long-Distance PCR --- p.102 / Chapter 3.2.1.5 --- Proteinase K Digestion --- p.103 / Chapter 3.2.1.6 --- Sfi Digestion --- p.104 / Chapter 3.2.1.7 --- cDNA size fractionation by CHROMA SPIN-400 --- p.104 / Chapter 3.2.1.8 --- Determination of the Ligation Efficiency --- p.106 / Chapter 3.2.1.9 --- Ligation of cDNA to lamda TriplEx2 Vector --- p.107 / Chapter 3.2.1.10 --- Lamda-phage Packaging Reaction --- p.107 / Chapter 3.2.1.11 --- Titering the Unamplifled Library and Determining the Percentage of Recombinant Clones --- p.108 / Chapter 3.2.1.12 --- Library Amplification --- p.109 / Chapter 3.2.1.13 --- Conversion of λTriplEx2 Recombinant Clones to pTriplEx2 Recombinant Plasmids --- p.111 / Chapter 3.2.2 --- Screening of the Subtractive library --- p.114 / Chapter 3.2.2.1 --- Verification of the enrichment by Plaque Lifting hybridization --- p.114 / Chapter 3.2.2.1.1 --- Lifting the Plaques --- p.114 / Chapter 3.2.2.1.2 --- Synthesis of the Probes for Plaque Lift Hybridization --- p.115 / Chapter 3.2.2.1.3 --- Hybridization to the Membranes --- p.116 / Chapter 3.2.2.2 --- Screening the Subtractive library by Macroarray Hybridization --- p.117 / Chapter 3.2.2.2.1 --- Colony Picking by QPik System --- p.117 / Chapter 3.2.2.2.2 --- Gridding of Macroarray --- p.118 / Chapter 3.2.2.2.3 --- Filter Processing of Gridded Membrane --- p.119 / Chapter 3.2.2.2.4 --- Hybridization to the Macroarray Membrane --- p.120 / Chapter 3.3 --- Results and Discussion --- p.121 / Chapter 3.3.1 --- Enrichment of Differentially Expressed Genes in Dikaryon L54 by Subtraction with Monokaryons A and B --- p.121 / Chapter 3.3.2 --- Construction of the full-length subtractive library --- p.123 / Chapter 3.3.3 --- Conversion of A TriplEx2 Recombinant Clones to pTriplEx2 Recombinant Plamid --- p.124 / Chapter 3.3.4 --- Verification the Enrichment of Subtractive library by Plaque lifting Hybridization --- p.125 / Chapter 3.3.5 --- Screening of the Subtractive library by Macroarray --- p.125 / Chapter 3.4 --- Discussion --- p.126 / Chapter Chapter Four --- Identification of Genes with Differentially Expression in Dikaryons by Subtactive cDNA Library Microarray / Chapter 4.1 --- Introduction --- p.135 / Chapter 4.2 --- Materials and Methods / Chapter 4.2.1 --- Selection and Amplification of clonesin SubtractionLlibrary for Microarray screening --- p.140 / Chapter 4.2.2 --- PCR product Purification --- p.141 / Chapter 4.2.3 --- Generation of Subtractive Dikaryotic Library Microarray Chip --- p.142 / Chapter 4.2.4 --- Screening the Differentially Expressed Genesin Dikaryon L54 by the Subtraction Dikaryotic Library cDNA Microarray Analysis --- p.143 / Chapter 4.2.4.1 --- Preparation of Total RNA --- p.143 / Chapter 4.2.4.2 --- Synthesis and fluorescent labeling of total cDNA --- p.145 / Chapter 4.2.4.3 --- Purification of labeled cDNA --- p.146 / Chapter 4.2.4.4 --- Storage Condition of Probe --- p.147 / Chapter 4.2.4.5 --- Analysis of labeled total cDNA --- p.148 / Chapter 4.2.4.6 --- Microarray hybridization --- p.150 / Chapter 4.2.4.7 --- Stringency washes --- p.152 / Chapter 4.2.4.8 --- Detection with TSA --- p.153 / Chapter 4.2.4.9 --- Image generation and data analysis --- p.155 / Chapter 4.2.5 --- Sequence analysis of clones showing differentially expressed in dikaryons in microarray screening --- p.157 / Chapter 4.2.5.1 --- Single-pass partial sequencing of 3´ة-end of subtractive cDNA clones --- p.157 / Chapter 4.2.5.2 --- Compiling dikaryotic EST database --- p.158 / Chapter 4.2.6 --- Comparison microarray analysis with SAGE analysis of the differentially expressed genes --- p.159 / Chapter 4.3 --- Results --- p.161 / Chapter 4.3.1 --- Preparation of clones for microarray hybridization --- p.161 / Chapter 4.3.2 --- Screening the differentially expressed genesin dikaryon L54 by the subtractive dikaryotic library cDNA microarray analysis --- p.162 / Chapter 4.3.2.1 --- Image capture and microarray data analysis --- p.162 / Chapter 4.3.2.2 --- Comparision of dikaryon L54 with monokaryons A and B --- p.163 / Chapter 4.3.2.3 --- Sequenced and comparison of the differentially expressed genes in dikaryon --- p.166 / Chapter 4.3.3 --- Comparison microarray analysis with SAGE analysis of the differentially expressed genes --- p.169 / Chapter Chapter Five --- Conclusion and Future Perpectives --- p.198 / References --- p.206