The purpose of this study is to attempt to determine what influence configuration and/or letter sequence have on the recognition thresholds of words. It is felt that low configuration and/or rare letter sequences will result in lower recognition thresholds, while high configuration and/or common letter sequence will result in higher recognition thresholds when stimulus words are presented to subjects by means of a tachistoscope.

Three genomic clones encompassing human DNA segments (designated LhX-3, LhX-4, and LhX5) were isolated from an X chromosome-specific library and subjected to analysis by physical mapping and DNA sequencing. It was found that these three clones are very rich in repetitive DNA sequence elements and retropseudogenes.

Master of Science / Department of Physics / Jeremy Schmit / In this work, we present a model for the kinetics of amyloid fibril aggregation. In the model we mapped the process of Hydrogen bond (H-bond) formation and breakage to a random-walk. we captured the effect of side chains using position dependent H-bonds free energies which allows us to calculated the residence time for different binding alignments with the fibril. The residence time can be compared to the diffusion-limited attachment rate to give net aggregation stability. This stability increases exponentially with increasing number of bonds or binding energy in homopolymer chains, however for chains with patterned sequences, the residence time shows strong effects of the binding alignment. Using the residence time for uniform structures combined with estimate of the diffusion rate, we modeled and simulated the kinetics of amyloid aggregation. Results of the simulations gives the bond energies and concentrations required for the onset of growth of aggregates.

by Tsui Kwok-wing. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1995. / Includes bibliographical references (leaves 188-216). / ACKNOWLEDGEMENTS --- p.ii / ABSTRACT --- p.iii / TABLE OF CONTENTS --- p.v / ABBREVIATIONS --- p.ix / Chapter CHAPTER 1 --- INTRODUCTION / Chapter 1.1 --- General introduction --- p.1 / Chapter 1.2 --- Human genome project --- p.5 / Chapter 1.3 --- Organization of human genome --- p.7 / Chapter 1.4 --- Adult human heart cDNA library --- p.9 / Chapter 1.5 --- Gene expression in adult human heart --- p.10 / Chapter 1.6 --- Polymerase chain reaction --- p.12 / Chapter 1.7 --- Purification of PCR products --- p.15 / Chapter 1.8 --- Automated DNA sequencing --- p.17 / Chapter 1.9 --- Sequence analysis by electronic mail server --- p.21 / Chapter 1.10 --- Effects of agar and agarose on Vent´ёØ and Taq DNA polymerases --- p.23 / Chapter 1.11 --- Transcription factors and zinc finger proteins --- p.25 / Chapter 1.12 --- LIM domain --- p.28 / Chapter 1.13 --- Cysteine-rich intestinal protein --- p.30 / Chapter CHAPTER 2 --- MATERIALS AND METHODS / Chapter 2.1 --- Plating out the adult human heart cDNA library --- p.32 / Chapter 2.2 --- Amplification by polymerase chain reaction --- p.33 / Chapter 2.3 --- Purification of the PCR products by Millipore filters --- p.35 / Chapter 2.4 --- Elimination of the purification of the PCR products before sequencing --- p.36 / Chapter 2.5 --- Cycle sequencing --- p.37 / Chapter 2.6 --- Unicycle sequencing --- p.38 / Chapter 2.7 --- Sequencing by T7 polymerase --- p.39 / Chapter 2.8 --- Gel electrophoresis in the automated A.L.F. sequencer --- p.41 / Chapter 2.9 --- Sequence analysis by commercially available softwares --- p.42 / Chapter 2.10 --- Sequence analysis through electronic mail server --- p.44 / Chapter 2.11 --- Database for storing the result of each clone --- p.46 / Chapter 2.12 --- Effects of agar and agarose on Vent´ёØ and Taq DNA polymerase --- p.47 / Chapter 2.13 --- Mini-preparation of plasmid DNA --- p.50 / Chapter 2.14 --- Large scale preparation of plasmid DNA --- p.51 / Chapter 2.15 --- Cloning the human cysteine rich heart protein (hCRHP) into the pAED4 vector --- p.53 / Chapter 2.16 --- Expression of hCRHP in E coli --- p.56 / Chapter 2.17 --- Northern hybridization --- p.58 / Chapter 2.18 --- Partial protein sequencing of hCRHP --- p.59 / Chapter CHAPTER 3 --- RESULTS / Chapter 3.1 --- The sequencing results of adult human heart cDNA clones --- p.60 / Chapter 3.2 --- Accuracy of sequencing results --- p.63 / Chapter 3.3 --- Catalogues of genes expressed in the adult human heart --- p.65 / Chapter 3.4 --- Effects of agar and agarose on Vent´ёØ and Taq DNA polymerases --- p.94 / Chapter 3.5 --- Elimination of the purification of the PCR products before sequencing --- p.102 / Chapter 3.6 --- Sequence analysis of hCRHP --- p.104 / Chapter 3.7 --- Northern hybridization of hCRHP --- p.109 / Chapter 3.8 --- Expression of hCRHP in E. coli --- p.112 / Chapter CHAPTER 4 --- DISCUSSION / Chapter 4.1 --- Random sequencing of adult human heart cDNA clones --- p.118 / Chapter 4.2 --- Catalogues of genes expressed in the adult human heart --- p.130 / Chapter 4.3 --- Gene expression in the adult human heart --- p.137 / Chapter 4.4 --- Importance of nonhuman matches --- p.170 / Chapter 4.5 --- Effects of agar and agarose on Vent´ёØ and Taq DNA polymerases --- p.177 / Chapter 4.6 --- Elimination of the purification of the PCR products before sequencing --- p.180 / Chapter 4.7 --- The possible role of CRIP and hCRHP --- p.184 / Chapter 4.8 --- Future prospect --- p.186 / REFERENCE --- p.188

by Li Leung Wah. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves 76-81). / Acknowledgements / Abstract / Chapter 1. --- Protein modeling --- p.1 / Chapter 1.1 --- Genetic Engineering --- p.1 / Chapter 1.2 --- Protein Engineering --- p.2 / Chapter 1.2.1 --- The basic concept --- p.2 / Chapter 1.2.2 --- The importance of protein modeling --- p.3 / Chapter 1.2.3 --- Applications --- p.4 / Chapter 1.2.3.1 --- Industry --- p.4 / Chapter 1.2.3.2 --- Medicine --- p.4 / Chapter 1.3 --- The structure of protein molecule --- p.5 / Chapter 2. --- About this thesis --- p.8 / Chapter 2.1 --- Methods on protein tertiary structure prediction --- p.8 / Chapter 2.1.1 --- Energy minimization method --- p.9 / Chapter 2.1.2 --- Sequence homology method --- p.9 / Chapter 2.1.3 --- Hierarchical assembly method --- p.11 / Chapter 2.2 --- Artificial Intelligence and molecular modeling --- p.11 / Chapter 2.3 --- Computer graphics and molecule display --- p.13 / Chapter 2.3.1 --- Molecular model in computer graphics --- p.13 / Chapter 2.3.2 --- Interactive graphic operations --- p.16 / Chapter 2.4 --- The objective of this thesis --- p.17 / Chapter 3. --- Algorithms for protein secondary structure prediction --- p.20 / Chapter 3.1 --- Hydrophobicity --- p.20 / Chapter 3.2 --- Algorithms for protein secondary structure prediction --- p.22 / Chapter 3.2.1 --- The Chou and Fasman method --- p.23 / Chapter 3.2.1.1 --- Method --- p.24 / Chapter 3.2.1.2 --- Results --- p.25 / Chapter 3.2.2 --- The GOR method --- p.26 / Chapter 3.2.2.1 --- Theory --- p.26 / Chapter 3.2.2.2 --- Method and results --- p.26 / Chapter 3.3 --- A proposed algorithm --- p.28 / Chapter 3.3.1 --- Procedure of our algorithm --- p.30 / Chapter 4. --- A protein tertiary structure prediction method --- p.31 / Chapter 4.1 --- The linkage between two amino acids --- p.32 / Chapter 4.2 --- Rotation angle between two peptide planes --- p.34 / Chapter 4.2.1 --- Helical structure --- p.35 / Chapter 4.2.1.1 --- Concept --- p.35 / Chapter 4.2.1.2 --- Procedure --- p.36 / Chapter 4.2.2 --- Sheet structure --- p.37 / Chapter 4.2.3 --- Turn structure --- p.38 / Chapter 4.2.4 --- Anti-parallel sheet and turn structure --- p.40 / Chapter 4.3 --- Random factor in rotation angle of peptide planes --- p.41 / Chapter 4.4 --- Atomic size --- p.41 / Chapter 4.5 --- Tertiary structure prediction algorithm --- p.42 / Chapter 5. --- Implementation --- p.45 / Chapter 5.1 --- Hardware --- p.45 / Chapter 5.2 --- User-defined data types and data structures --- p.46 / Chapter 5.3 --- Technique in molecule displaying --- p.48 / Chapter 5.4 --- Image processing --- p.50 / Chapter 5.5 --- Options in our program --- p.52 / Chapter 5.6 --- Steps in protein tertiary structure prediction --- p.54 / Chapter 6. --- Results --- p.59 / Chapter 6.1 --- The results of protein secondary structure prediction --- p.59 / Chapter 6.2 --- The results of protein tertiary structure prediction --- p.66 / Chapter 7. --- Conclusion --- p.70 / Chapter 7.1 --- Comments on protein secondary structure prediction algorithm --- p.70 / Chapter 7.1.1 --- Advantages and disadvantages --- p.70 / Chapter 7.1.2 --- Further development --- p.71 / Chapter 7.2 --- Discussion on X-ray crystallographic data --- p.72 / Chapter 7.3 --- Comments on the protein tertiary structure prediction algorithm --- p.73 / Chapter 7.3.1 --- Advantages and disadvantages --- p.73 / Chapter 7.3.2 --- Further development --- p.74 / Chapter 7.3.2.1 --- Rotation angle between two peptide planes --- p.74 / Reference --- p.76 / Glossary --- p.82 / Appendix A An algorithm to determine hydrophobic value --- p.83 / Appendix B Chou and Fasman algorithm --- p.84 / Appendix C GOR algorithm --- p.87 / Appendix D Shading algorithm --- p.88

by Agnes, Lo Shuk Yee. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1995. / Includes bibliographical references (leaves [27-33] (2nd gp.)). / Chapter PART 1: --- Human Heart cDNA Library Sequencing / Chapter A) --- Introduction of human heart cDNA library sequencing / Chapter A.1 --- Human genome project / Chapter A.2 --- The aim of human genome project / Chapter A.3 --- Automatic sequencing / Chapter A.4 --- Cycle sequencing reaction / Chapter A.5 --- Human heart cDNA library sequencing project / Chapter B) --- Methods and materials / Chapter (I) --- Preparation of plating bacterial-Y1090 / Chapter (II) --- Plating the bacteriophage with blue-white visual selection / Chapter (III) --- Amplification of bacteriophage cDNA clones by PCR / Chapter (IV) --- Purification and quantitation of PCR products / Chapter (V) --- Cycle DNA sequencing of PCR products / Chapter (VI) --- Casting the sequencing gel / Chapter (VII) --- Sequencing by Pharmacia LKB A.L.F. DNA Sequencer / Chapter (VIII) --- Editing and saving the DNA sequence / Chapter (IX) --- Sending the DNA sequence to Genbank by E-mail / Chapter (X) --- Usage of the Genbank database / Chapter C) --- Results / Chapter D) --- Discussions / Chapter D.1 --- Application of human genomic project / Chapter D.2 --- Interpretation of the sequencing results / Chapter D.3 --- Quality of cDNA libraries and representation of mRNA population / Chapter D.4 --- "Gene expression profile in three different organs-heart, brain and liver" / Chapter D.5 --- Population study of the cDNA library / Chapter D.6 --- Isolation of a large number of novel genes by substraction cDNA library / Chapter D.7 --- Screening method to find out the complete coding sequence of interesting genes / Chapter D.8 --- Technical problems encountered and managed / Chapter PART 2: --- Identification of human cytosolic malate dehydrogenase by large scale human heart cDNA library sequencing / Chapter CHAPTER 1: --- Introduction of malate dehydrogenase / Chapter 1.1 --- Malate dehydrogenase--Kreb's cycle enzyme / Chapter 1.2 --- Two stereospecific forms of dehydrogenase / Chapter 1.3 --- NAD-binding domain / Chapter 1.4 --- The active site / Chapter 1.5 --- Comparison of surface properties between cMDH and mMDH / Chapter 1.6 --- N-terminal region and mitochondrial import / Chapter 1.7 --- Subunit-subunit interactions / Chapter 1.8 --- Physiological importance of malate dehydrogenase / Chapter 1.9 --- Secondary structure-total 11 β-strands and 9 α-helixes / Chapter 1.10 --- Objectives of the thesis / Chapter CHAPTER 2: --- Cloning and sequence analysis of human cytosolic malate dehydrogenase (hcMDH) / Chapter 2.1 --- Cloning of human cytosolic malate dehydrogenase (hcMDH) / Chapter 2.1.1 --- Methods and materials / Chapter 2.1.1.1 --- Cloning full length of hcMDH into expression vector pAED4 / Chapter 2.1.1.2 --- Preparation of competent cell-JM109 for transformation / Chapter 2.1.1.3 --- Minipreparation of plasmid DNA / Chapter 2.1.1.4 --- Midi-preparation of bacteriophage λDNA by QIAGEN´ёØ / Chapter 2.1.1.5 --- Titration of bacteriophage λ of human adult heart cDNA library / Chapter 2.1.1.6 --- Preparation of soft-agarose lysates / Chapter 2.1.1.7 --- Elution of DNA from agarose gel by GENECLEAN´ёØ / Chapter 2.1.2 --- Results / Chapter 2.1.3 --- Discussions / Chapter 2.2 --- Sequence analysis of human cytosolic malate dehydrogenase (hcMDH) / Chapter 2.2.1 --- Methods and materials: Autoread sequencing / Chapter (I) --- Annealing of primer to double-stranded template / Chapter (II) --- Sequencing / Chapter 2.2.2 --- Results and discussions / Chapter 2.3 --- Amino acids and protein structure analysis of cMDH / Chapter CHAPTER 3 : --- "Protein expression, partial purification and folding experiments of human cytosolic malate dehydrogenase (hcMDH)" / Chapter 3.1 --- Protein expression of hcMDH in E. coli / Chapter 3.1.1 --- Methods and materials / Chapter 3.1.1.1 --- Protein expression induced by IPTG / Chapter 3.1.1.2 --- Isoelectric focusing (IEF)-two dimensional gel electrophoresis / Chapter (I) --- First dimensional electrofocusing / Chapter (II) --- The second dimension SDS-PAGE electrophoresis / Chapter (III) --- Sample preparation / Chapter 3.1.2 --- Results / Chapter 3.1.3 --- Discussions / Chapter 3.1.3.1 --- The properties of expressed protein of hcMDH / Chapter 3.1.3.2 --- T7 expression system / Chapter 3.1.3.3 --- Strong φ 10 promoter / Chapter 3.1.3.4 --- E.coli BL21 host cell / Chapter 3.2 --- Partial purification and folding experiments of hcMDH / Chapter 3.2.1 --- Methods and materials / Chapter 3.2.1.1 --- Partial purification of hcMDH expressed protein / Chapter (I) --- Preparation of supernatant from E.coli crude extract / Chapter (II) --- Ion-exchange column chromatography / Chapter (III) --- Affinity chromatography / Chapter (IV) --- Gel filtration on a Sepharose CL-6B column / Chapter 3.2.1.2 --- Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE) / Chapter 3.2.1.3 --- Staining the protein gel by the Coomassie Blue R-250 method / Chapter 3.2.1.4 --- Staining the protein gel by the Silver staining Method / Chapter 3.2.1.5 --- Quantitation of protein by the Bradford Method / Chapter 3.2.1.6 --- Native gel electrophoresis / Chapter 3.2.1.7 --- Malate dehydrogenase MDH enzyme staining method / Chapter 3.2.1.8 --- Malate dehydrogenase MDH enzyme assay / Chapter 3.2.1.9 --- Fast protein liquid chromatography (FPLC) / Chapter 3.2.1.10 --- Protein folding experiment / Chapter 3.2.1.11 --- Eukaryotic expression of hcMDH / Chapter 3.2.2 --- Results / Chapter 3.2.2.1 --- Partial purification by chromatography / Chapter 3.2.2.2 --- Native gel / Chapter 3.2.2.3 --- FPLC / Chapter 3.2.2.4 --- To aid folding of protein by adding NADH / Chapter 3.2.2.5 --- Eukaryotic expression / Chapter 3.2.3 --- Discussions / Chapter 3.2.3.1 --- Purification of malate dehydrogenase MDH / Chapter 3.2.3.2 --- "Methods for visualizing dehydrogenase enzymes, e.g. malate dehydrogenase" / Chapter 3.2.3.3 --- The presence of unfold hcMDH protein in bacteria / Chapter 3.2.3.4 --- Folding of protein by heat shock protein GroE / Chapter 3.2.3.5 --- Eukaryotic expression / Chapter CHAPTER 4: --- Master screening of single base change by PCR-SSCP (Single Strand Conformational Polymorphism) / Chapter 4.1 --- Theory of SSCP / Chapter 4.2 --- Methods and materials / Chapter 4.3 --- Results / Chapter 4.4 --- Discussions / Chapter 4.4.1 --- The procedure of SSCP / Chapter 4.4.2 --- An alternative quick detection method for polymorphism of hcMDH at position 565--by automatic sequencing / Chapter 4.4.3 --- Other detection methods-- RNA-PCR and ddF / Chapter 4.4.4 --- Parameters affecting sensitivity of SSCP / Chapter 4.4.5 --- Application of SSCP / Chapter CHAPTER 5: --- Southern hybridization and In situ hybridization / Chapter 5.1 --- Southern blot analysis of human cytosolic malate dehydrogenase (hcMDH) / Chapter 5.1.1 --- Methods and materials / Chapter (I) --- Transfer genomic DNA to Nylon membrane / Chapter (II) --- Synthesis of radiolabelling cDNA probe / Chapter (III) --- Pre-hybridization and hybridization reaction / Chapter 5.1.2 --- Results / Chapter 5.1.3 --- Discussions / Chapter 5.2 --- In situ hybridization / Chapter 5.2.1 --- Methods and materials / Chapter (I) --- Preparation of Dig labelling probe by random primed labelling / Chapter (II) --- Estimating the yield of Dig-labelled nucleic acids / Chapter (III) --- Denaturation and hybridization of the hcMDH probe with animal tissues / Chapter (IV) --- Color development of the tissue / Chapter 5.2.2 --- Results / Chapter 5.2.3 --- Discussions / Chapter 5.2.3.1 --- Cellular distribution of hcMDH / Chapter 5.2.3.2 --- The principle of in situ hybridization / Chapter 5.2.3.3 --- Specimen preparation / Chapter 5.2.3.4 --- Hybridization conditions / Chapter 5.2.3.5 --- "Ontogeny of MDH in rabbit fetal brain, heart and lung" / Appendixes: / "Appendix I: 531 random cDNA clones from clone no. J950 to K951 in human heart cDNA library sequencing project. The name of clones, accession number, the length of the partial sequence and percentage of match are listed" / Appendix II: The new accession no. of Novel clones in Genbank / "Appendix III: The enzymatic reaction, molecular weigth, specific activity and Michaelis constants of different sources of malate dehydrogenase" / Appendix IV: The full sequence of nucleic acids and amino acids of human cytosolic malate dehydrogenase hcMDH. Accession no. of hcMDH is U20352 in Genbank / Appendix V: Nucleotide sequences of the mouse cMDH gene / Appendix VI: Nucleotide sequences of the mouse mMDH gene / Appendix VII: Structural organization of the mouse cytosolic malate dehydrogenase and its comparison with that of the mouse mitochondrial malate dehydrogenase gene

by Agnes Pui-Yee Chan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1992. / Includes bibliographical references (leaves 162-177). / ACKNOWLEDGEMENTS --- p.i / ABSTRACT --- p.ii / TABLE OF CONTENTS --- p.iv / ABBREVIATIONS --- p.vii / Chapter CHAPTER 1 --- INTRODUCTION / Chapter 1.1 --- Physiology of growth --- p.3 / Chapter 1.2 --- The anterior pituitary --- p.4 / Chapter 1.3 --- Chemistry of GH and the GH gene family --- p.7 / Chapter 1.4 --- Biochemical effects and mode of action of GH --- p.8 / Chapter 1.5 --- Control of GH at cellular level --- p.10 / Chapter 1.6 --- Control of GH gene expression at molecular level / Chapter 1.6.1 --- Introduction --- p.11 / Chapter 1.6.2 --- Tissue-specific expression of GH gene / Chapter 1.6.2.1 --- Tissue-specific transcription factors of pituitary cells --- p.20 / Chapter 1.6.2.2 --- Non-tissue specific transcription factors of pituitary cells --- p.27 / Chapter 1.6.2.3 --- Negatively-acting transcription factors of non-pituitary cells --- p.34 / Chapter 1.6.2.4 --- Theory for tissue-specific GH gene activation --- p.39 / Chapter 1.7 --- Characteristic of growth in fish --- p.40 / Chapter 1.8 --- Objectives of the present study --- p.42 / Chapter CHAPTER 2 --- MATERIALS AND METHODS / Chapter 2.1 --- General techniques / Chapter 2.1.1 --- Preparation of DNA / Chapter 2.1.1.1 --- Minipreparation of DNA --- p.46 / Chapter 2.1.1.2 --- Preparation of DNA using Qiagen column --- p.47 / Chapter 2.1.1.3 --- Preparation of phage DNA --- p.48 / Chapter 2.1.2 --- Elution of DNA from agarose gel --- p.51 / Chapter 2.1.3 --- Preparation of competence cells and transformation --- p.52 / Chapter 2.1.4 --- Ligation of DNA fragments --- p.53 / Chapter 2.1.5 --- Cell feeding and subculturing --- p.54 / Chapter 2.2 --- Special techniques / Chapter 2.2.1 --- DNA sequencing --- p.56 / Chapter 2.2.2 --- Polymerase chain reaction (PCR) --- p.67 / Chapter 2.2.3 --- Direct sequencing of PCR products --- p.72 / Chapter 2.2.4 --- Nested-deletion --- p.75 / Chapter 2.2.5 --- DNA transfection --- p.81 / Chapter 2.2.6 --- CAT assay --- p.86 / Chapter CHAPTER 3 --- RESULTS / Chapter 3.1 --- Sequencing of the grass carp GH gene / Chapter 3.1.1 --- Introduction --- p.93 / Chapter 3.1.2 --- Sequencing strategy --- p.94 / Chapter 3.2 --- Sequence analysis of the grass carp GH gene --- p.108 / Chapter 3.3 --- Functional analysis of the grass carp GH gene --- p.115 / Chapter CHAPTER 4 --- DISCUSSIONS / Chapter 4.1 --- DNA sequence comparison between grass carp GH gene and other organisms --- p.137 / Chapter 4.2 --- Amino acid comparisons between grass carp GH and other organisms --- p.143 / Chapter 4.3 --- Tissue-specific expression of GH gene / Activation of transcription --- p.154 / Repression of transcription --- p.155 / Chapter 4.4 --- Electroporation of zebrafish eggs --- p.157 / Chapter 4.5 --- Further studies --- p.160 / REFERENCES --- p.162 / APPENDIX --- p.178

Siu Shu Shun. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 100-106). / Abstracts in English and Chinese. / Acknowledgements --- p.i / Contents --- p.ii / Abstract / 摘要 --- p.iv / Abbreviations --- p.vi / List of figures --- p.vii / List of tables --- p.x / Chapter Chapter 1: --- Introduction / Chapter 1.1 --- "Human EST sequencing project, the role and goal" --- p.1 / Chapter 1.2 --- Human liver cDNA sequencing --- p.2 / Chapter 1.3 --- The role of membrane-associated proteins in hepatocellular functions --- p.3 / Chapter 1.3.1 --- Outline of the liver function --- p.3 / Chapter 1.3.2 --- Basic structure of hepatocyte --- p.4 / Chapter 1.3.3 --- Category of membrane associated proteins --- p.5 / Chapter 1.4 --- Identification of human OAT2 gene --- p.7 / Chapter 1.5 --- The multispecific transporter family --- p.8 / Chapter 1.5.1 --- Classification --- p.8 / Chapter 1.5.2 --- The human OAT family --- p.9 / Chapter 1.6 --- The characteristics of rat multispecific OAT2 --- p.11 / Chapter 1.7 --- Clinical significance of organic anion transport proteins --- p.14 / Chapter Chapter 2: --- Materials and Methods / Chapter 2.1 --- Human liver EST sequencing project --- p.16 / Chapter 2.1.1 --- Plating out the adult human liver phage library --- p.16 / Chapter 2.1.2 --- PCR detection and amplification of the cDNA clone --- p.17 / Chapter 2.1.3 --- Automatic cDNA sequencing --- p.18 / Chapter 2.2 --- Cloning of hOAT2 gene into TA cloning vector pT-Adv --- p.19 / Chapter 2.2.1 --- Amplification of hOAT2 by PCR --- p.19 / Chapter 2.2.2 --- Ligation reaction --- p.19 / Chapter 2.2.3 --- Transformation of recombinant plasmid into competent cells --- p.20 / Chapter 2.3 --- Sequence analysis and structural prediction --- p.20 / Chapter 2.4 --- Cloning of the hOAT2 gene into the pQE30 expression vector --- p.21 / Chapter 2.4.1 --- PCR amplification and restriction endonuclease cutting --- p.21 / Chapter 2.4.2 --- Gene clean --- p.22 / Chapter 2.4.3 --- Preparation of bacterial competent cells --- p.23 / Chapter 2.5 --- Small scale synthesis of plasmid DNA --- p.24 / Chapter 2.6 --- Large scale synthesis of plasmid DNA --- p.25 / Chapter 2.7 --- Cloning of the hOAT2 gene into the pSecTag2B mammalian expression vector --- p.26 / Chapter 2.8 --- Cloning of the hOAT2 gene into the pEGFP-C2 fluorescent vector --- p.27 / Chapter 2.8.1 --- Tissue culture and transfection --- p.27 / Chapter 2.8.2 --- Fluorescence microscopy examination --- p.28 / Chapter 2.9 --- Chromosomal mapping of the hOAT2 gene --- p.29 / Chapter 2.9.1 --- Somatic cell hybrids mapping --- p.29 / Chapter 2.9.2 --- Radiation hybrids mapping --- p.29 / Chapter 2.10 --- Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) --- p.30 / Chapter 2.11 --- Western hybridization --- p.32 / Chapter 2.11.1 --- Preparation of anti-hOAT2 antibodies --- p.32 / Chapter 2.11.1.1 --- Synthetic peptide conjugation --- p.32 / Chapter 2.11.1.2 --- Immunizing rabbit polyclonal antibodies for human OAT2 --- p.32 / Chapter 2.11.1.3 --- Purification of the rabbit polyclonal IgG antibodies --- p.33 / Chapter 2.11.2 --- Western blot analysis --- p.33 / Chapter 2.11.2.1 --- Protein isolation from rat liver --- p.33 / Chapter 2.11.2.2 --- Prote in preparation from cell lysate --- p.34 / Chapter 2.11.2.3 --- Quantitation of total proteins by Bradford protein assay --- p.35 / Chapter 2.11.2.4 --- Blotting and hybridization --- p.35 / Chapter Chapter 3: --- Results / Chapter 3.1 --- Catalogue of the 500 liver ESTs --- p.37 / Chapter 3.2 --- Nomenclature of human NLT gene --- p.47 / Chapter 3.3 --- Cloning and characterization of the hOAT2 sequence --- p.48 / Chapter 3.3.1 --- Isolation of hOAT2 cDNA from human liver cDNA library --- p.48 / Chapter 3.3.2 --- The primary and secondary structural analysis of hOAT2 --- p.53 / Chapter 3.3.3 --- Motif search and prediction --- p.61 / Chapter 3.3.4 --- Homology alignment --- p.64 / Chapter 3.4 --- Chromosomal mapping of hOAT2 gene --- p.67 / Chapter 3.4.1 --- Somatic cell hybrid mapping of hOA T2 gene --- p.67 / Chapter 3.4.2 --- Radiation hybrid mapping of hOA T2 gene --- p.69 / Chapter 3.4.3 --- Identification of partial human genomic sequence --- p.73 / Chapter 3.5 --- Detection of the hOAT2 gene expression in human tissues by RT- PCR assay --- p.76 / Chapter 3.6 --- Detection of subcellular localization of hOAT2 protein by conjugating fluorescence protein --- p.81 / Chapter 3.7 --- Immunodetection of protein extracts from cultured cells --- p.83 / Chapter Chapter 4: --- Discussion / Chapter 4.1 --- Characterization of the hepatocellular ESTs --- p.85 / Chapter 4.1.1 --- Classification and frequency distribution of the 500 ESTs --- p.85 / Chapter 4.1.2 --- The expression pattern of membrane associated proteins --- p.87 / Chapter 4.2 --- Tissue distribution and expression profiles of hOAT2 --- p.88 / Chapter 4.3 --- HOAT2 in fetal development --- p.89 / Chapter 4.4 --- Predicting the topology of membrane proteins --- p.90 / Chapter 4.5 --- Chromosomal mapping of human OAT2 --- p.91 / Chapter 4.6 --- Possible functions of hOAT2 --- p.93 / Chapter 4.6.1 --- Hepato-renal relation --- p.93 / Chapter 4.6.2 --- Substrate diversity --- p.95 / Chapter 4.7 --- Fluorescence detection for subcellular localization --- p.96 / Chapter 4.8 --- Conclusion --- p.97 / Chapter 4.9 --- Further aspects --- p.99 / References --- p.100 / Appendix --- p.107

Ixodes ricinus is the most important human-biting tick in Europe and the principal vector of Lyme borreliosis. In addition, this hard tick species transmits a large number of microbial pathogens that are of importance to animal and human health. Little is known about the diversity and genetic population structure of I. ricinus across Europe. Genetic diversity of these tick populations may have implications on disease transmission. I. ricinus primers were designed for a number of mitochondrial genes and a Multilocus Sequence Typing-like Scheme (MLST) was devised. This was termed mitochondrial MLST (mtMLST). MLST has so far mainly been used for typing microbes, and the development of a MLST scheme for an arthropod vector is novel. Understanding the geographic structure of I. ricinus populations, in combination with studies regarding the migration of tick-borne microbial infections, e.g. Lyme borreliosis, is likely to illuminate important processes in the evolution and spread of tick-borne diseases.

Mathematics, physics, biology, and computer science are combined to create computational modeling, which studies the behaviors and reactions of complex biomedical problems. Modern biomedical research relies significantly on realistic computational human models or “virtual humans�. Relevant study areas utilizing computational human models include electromagnetics, solid mechanics, fluid dynamics, optics, ultrasound propagation, thermal propagation, and automotive safety research. These and other applications provide ample justification for the realization of the Visible Human Project® (VHP)-Female v. 4.0, a new platform-independent full body electromagnetic computational model. Along with the VHP-Female v. 4.0, a realistic and anatomically justified Dynamic Breathing Sequence is developed. The creation of such model is essential to the development of biomedical devices and procedures that are affected by the dynamics of human breathing, such as Magnetic Resonance Imaging and the calculation of Specific Absorption Rate. The model can be used in numerous application, including Breath-Detection Radar for human search and rescue.