Chan Ho Yu, Richard. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 115-122). / Abstracts in English and Chinese. / Declaration --- p.i / Acknowledgements --- p.ii / Abstract --- p.iii / 摘要 --- p.v / Table of Contents --- p.vii / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1 --- Potassium Channels --- p.1 / Chapter 1.1.1 --- Potassium Ions --- p.1 / Chapter 1.1.2 --- Potassium Channels --- p.1 / Chapter 1.1.3 --- Structure of K Channels --- p.2 / Chapter 1.1.4 --- Classification ofK Channels --- p.3 / Chapter 1.1.5 --- Mechanisms Contributed to K Channel Functions and Diversity --- p.5 / Chapter 1.1.5.1 --- RNA Editing --- p.5 / Chapter 1.1.5.2 --- Alternative Splicing --- p.6 / Chapter 1.1.5.3 --- Heteromultimeric Assembly of Principal Subunits --- p.6 / Chapter 1.1.5.4 --- Auxiliary Subunits --- p.7 / Chapter 1.1.5.5 --- Posttranslational Modifications --- p.7 / Chapter 1.2 --- Voltage-gated Potassium (Kv) Channels --- p.9 / Chapter 1.2.1 --- Diversity of Kv Channel Structure --- p.9 / Chapter 1.2.2 --- Early Origin of the Kv Family --- p.10 / Chapter 1.2.3 --- Structural Diversity of Kv Channels in Drosophila --- p.11 / Chapter 1.2.4 --- Structural Diversity of Kv Channels in Mammals --- p.11 / Chapter 1.2.5 --- Phylogenetic Tree of Kv Family --- p.13 / Chapter 1.2.6 --- Tissue Expression of Kv Channels --- p.13 / Chapter 1.2.7 --- "Three Main Functions of Kv Channels as Signaling Proteins: Ion Permeation, Gating and Sensing" --- p.16 / Chapter 1.2.7.1 --- Ion Permeation --- p.16 / Chapter 1.2.7.2 --- Gating --- p.18 / Chapter 1.2.7.2.1 --- Gating at the S6 Bundle Crossing --- p.18 / Chapter 1.2.7.2.2 --- Ball-and-Chain Gating --- p.19 / Chapter 1.2.7.2.3 --- Gating at the Selectivity Filter --- p.19 / Chapter 1.2.7.3 --- Sensing Mechanisms --- p.20 / Chapter 1.2.7.3.l --- Voltage Sensor --- p.20 / Chapter 1.2.7.3.2 --- Gating Sensors for Ligands --- p.21 / Chapter 1.3 --- KCNA10 --- p.22 / Chapter 1.3.1 --- "Rabbit Homologue of KCNA10, Kcnl" --- p.22 / Chapter 1.3.2 --- Genomic Localization of Human KCNA10 --- p.23 / Chapter 1.3.3 --- Human Gene for KCNA10 --- p.23 / Chapter 1.3.4 --- Basic Kinetic and Pharmacological Properties of KCNA10 --- p.25 / Chapter 1.3.5 --- "Regulation of KCNAlO by KCNA4B, a β -subunit" --- p.27 / Chapter 1.4 --- Aim of the Present Study --- p.30 / Chapter Chapter2: --- Materials and Methods --- p.31 / Chapter 2.1 --- Molecular Sub-Cloning ofKCNAlO --- p.31 / Chapter 2.1.1 --- Polymerase Chain Reaction (PCR) ofKCNA10 Fragment from KCNA Clone --- p.10 / Chapter 2.1.2 --- Separation and Purification of PCR Products --- p.32 / Chapter 2.1.2.1 --- Separation --- p.32 / Chapter 2.1.2.2 --- Purification --- p.33 / Chapter 2.1.3 --- Polishing the Purified PCR Products --- p.33 / Chapter 2.1.4 --- Ligation of PCR Products and pPCR-Script Amp SK(+) Cloning Vector --- p.34 / Chapter 2.1.5 --- Transformation --- p.34 / Chapter 2.1.6 --- Preparing Glycerol Stocks Containing the Bacterial Clones --- p.35 / Chapter 2.1.7 --- Plasmid DNA Preparation --- p.35 / Chapter 2.1.8 --- Clones Confirmation --- p.36 / Chapter 2.1.8.1 --- Restriction Enzyme Digestion --- p.36 / Chapter 2.1.8.2 --- Automatic Sequencing --- p.37 / Chapter 2.2 --- In situ Hybridization --- p.39 / Chapter 2.2.1 --- Probe Preparation --- p.39 / Chapter 2.2.1.1 --- Antisense KCNA10 RNA Probe --- p.39 / Chapter 2.2.1.2 --- Sense KCNA10 RNA Probe (Control Probe) --- p.40 / Chapter 2.2.2 --- Testing of DIG-Labeled RNA Probes --- p.43 / Chapter 2.2.3 --- Paraffin Sections Preparation --- p.43 / Chapter 2.2.4 --- In situ Hybridization: Pretreatment --- p.44 / Chapter 2.2.5 --- "Pre-hybridization, Hybridization and Post-hybridization" --- p.45 / Chapter 2.2.5.1 --- Pre-hybridization --- p.45 / Chapter 2.2.5.2 --- Hybridization --- p.45 / Chapter 2.2.5.3 --- Post-hybridization --- p.46 / Chapter 2.2.6 --- Colourimetnc Detection of Human KCNA10 --- p.46 / Chapter 2.3 --- Cell Culture --- p.47 / Chapter 2.3.1 --- Human Kidney Proximal Epithelial Cell Line (OK) --- p.47 / Chapter 2.3.2 --- Mouse Micro-vessel Endothelial Cell Line (H5V) --- p.48 / Chapter 2.3.3 --- Mouse Neuroblastoma Cell Line (NG108-15) --- p.48 / Chapter 2.3.4 --- Human Bladder Epithelial Cell Line (ECV304) --- p.48 / Chapter 2.3.5 --- Human T Cell Leukemia Cell Line (Jurkat) --- p.49 / Chapter 2.4 --- Total RNA Extraction --- p.49 / Chapter 2.5 --- Reverse Transcription from Cell Line --- p.51 / Chapter 2.6 --- Polymerase Chain Reaction (PCR) ofKCNAl 0 Fragment from Frist Strand cDNA --- p.51 / Chapter 2.7 --- Northern Hybridization --- p.52 / Chapter 2.7.1 --- Probe Preparation --- p.52 / Chapter 2.7.2 --- Separating RNA on an Agarose Gel --- p.52 / Chapter 2.7.3 --- RNA Transfer and Fixation --- p.52 / Chapter 2.7.4 --- Hybridization --- p.54 / Chapter 2.7.5 --- Post-hybridization --- p.54 / Chapter 2.7.6 --- Chemiluminescent Detection --- p.55 / Chapter 2.8 --- Intracellular Free Calcium Ion ([Ca2+］i) Measurement by Confocal Imaging System --- p.56 / Chapter 2.8.1 --- Bathing Solutions --- p.56 / Chapter 2.8.2 --- Preparation of Cells for [Ca2+]i Measurement --- p.56 / Chapter 2.8.3 --- Confocal Imaging System --- p.57 / Chapter 2.8.3.1 --- Fluo-3/AM Dye Loading --- p.57 / Chapter 2.8.3.2 --- [Ca2+]i Measurement --- p.57 / Chapter Chapter3: --- Results --- p.59 / Chapter 3.1 --- Phylogenetic Tree Reconstruction ofKCNAl0 --- p.59 / Chapter 3.2 --- Hydropathy Analysis ofKCNAl0 --- p.60 / Chapter 3.3 --- Molecular Sub-Cloning ofKCNAl0 --- p.61 / Chapter 3.3.1 --- Polymerase Chain Reaction (PCR) ofKCNAl0 Fragment from KCNA10 Clone --- p.61 / Chapter 3.3.2 --- Clones Confirmation --- p.63 / Chapter 3.4 --- In situ Hybridization Analysis ofKCNAl0 mRNAExpression --- p.65 / Chapter 3.4.1 --- Expression ofKCNAl0 in Human Kidney (Nephron) --- p.66 / Chapter 3.4.2 --- Expression ofKCNAl0 in Human Cerebral Artery --- p.69 / Chapter 3.4.3 --- Expression ofKCNAl0 in Human Cerebellum --- p.71 / Chapter 3.4.4 --- Expression ofKCNAl0 in Human Hippocampus --- p.73 / Chapter 3.4.5 --- Expression ofKCNAl0 in Human Occipital Cortex --- p.75 / Chapter 3.4.6 --- Expression ofKCNAl0 in Human Esophagus --- p.77 / Chapter 3.4.7 --- Expression ofKCNAl0 in Human Lung --- p.79 / Chapter 3.4.8 --- Expression ofKCNAl0 in Human Thyroid Glands --- p.81 / Chapter 3.4.9 --- Expression ofKCNAl0 in Human Adrenal Glands --- p.83 / Chapter 3.4.10 --- Expression ofKCNAl0 in Human Spleen --- p.86 / Chapter 3.5 --- RT-PCR ofKCNAl0 Fragment from Different Tissues --- p.88 / Chapter 3.6 --- Northern Blot Analysis of KCNA10 in Different Tissues --- p.90 / Chapter 3.7 --- Effects of Blocking KCNA10 on Ca2+ influx in Human Renal Proximal Tubule Epithelial Cells --- p.91 / Chapter Chapter4: --- Discussion --- p.97 / Chapter 4.1 --- Phylogency ofKCNAlO --- p.97 / Chapter 4.2 --- Hydropathy Plot for KCNA10 --- p.97 / Chapter 4.3 --- Expression ofKCNAl0 --- p.98 / Chapter 4.3.1 --- In situ Hybridization --- p.98 / Chapter 4.3.2 --- RT-PCR & Northern Blot Analysis --- p.99 / Chapter 4.4 --- Functional Implication of KCNA10 Expression in Different Human Tissues --- p.100 / Chapter 4.4.1 --- Unique Functional Properties ofKCNAlO --- p.100 / Chapter 4.4.2 --- Role ofKCNAlO in Renal Proximal Tubule --- p.101 / Chapter 4.4.2.1 --- Functions ofK+ Channels in Kidney --- p.101 / Chapter 4.4.2.2 --- The Function ofKCNAlO --- p.104 / Chapter 4.4.3 --- Role ofKCNAl0 in Blood Vessels --- p.106 / Chapter 4.4.3.1 --- Endothelial Cells --- p.106 / Chapter 4.4.3.2 --- Smooth Muscle Cells --- p.108 / Chapter 4.4.4 --- Role ofKCNA10 in CNS --- p.109 / Chapter 4.4.5 --- Role ofKCNAl0 in Secretory Cells --- p.111 / Chapter 4.4.6 --- Role ofKCNAl0 in Lung --- p.112 / Chapter 4.5 --- Conclusion --- p.114 / Chapter Chapter5： --- Reference --- p.115
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_324505 |
| Date | January 2003 |
| Contributors | Chan, Ho Yu., Chinese University of Hong Kong Graduate School. Division of Physiology. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xi, 122 leaves : ill. (some col.) ; 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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