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Proteomic studies on Cordyceps and characterization of its anti-proliferation effect on kidney cancer cells.

Lai, Sze Tsai. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 94-104). / Abstracts in English and Chinese. / Thesis Committees --- p.i / Statement --- p.ii / Abstract --- p.iii / 摘要 --- p.v / Acknowledgments --- p.vi / List of Abbreviations --- p.vii / Table of Contents --- p.ix / List of Tables --- p.xiii / List of Figures --- p.xiv / Chapter 1 --- Literature review --- p.1 / Chapter 1.1 --- Introduction to Cordyceps --- p.1 / Chapter 1.2 --- Fungal proteomics --- p.2 / Chapter 1.2.1 --- Extraction method --- p.2 / Chapter 1.2.2 --- Proteomic study of Cordyceps --- p.3 / Chapter 1.3 --- Ingredients of Cordyceps and their related biological activities --- p.5 / Chapter 1.3.1 --- Polysaccharides --- p.5 / Chapter 1.3.2 --- Nucleosides --- p.6 / Chapter 1.3.2.1 --- Cordycepin --- p.6 / Chapter 1.3.2.2 --- Adenosine --- p.8 / Chapter 1.4 --- Cordyceps and their related biological activities --- p.9 / Chapter 1.4.1 --- Cordyceps militaris --- p.9 / Chapter 1.4.2 --- Cordyceps sinensis --- p.10 / Chapter 1.5 --- Proteomic analysis of proteome change --- p.12 / Chapter 1.5.1 --- Proteomic tools used to study the change in protein expression --- p.12 / Chapter 1.5.2 --- Two-dimensional gel electrophoresis --- p.13 / Chapter 1.5.3 --- Mass spectrometry --- p.13 / Chapter 1.6 --- Objective --- p.16 / Chapter 2 --- Methodology --- p.17 / Chapter 2.1 --- Cultivation of Cordyceps militaris --- p.17 / Chapter 2.2 --- Proteomic study on Cordyceps militaris --- p.17 / Chapter 2.2.1 --- Extraction of proteins from Cordyceps militaris --- p.17 / Chapter 2.2.2 --- Protein quantification --- p.18 / Chapter 2.2.3 --- 2D Gel electrophoresis --- p.19 / Chapter 2.2.4 --- Image analysis --- p.20 / Chapter 2.2.5 --- In gel digestion and MALDI-ToF MS --- p.20 / Chapter 2.3 --- Preparation of Cordyceps extracts for anti-proliferation assay on cell lines --- p.21 / Chapter 2.3.1 --- Types of the extracts of Cordyceps --- p.21 / Chapter 2.3.2 --- Preparation of the extracts of Cordyceps --- p.21 / Chapter 2.4 --- Anti-proliferation assay on cell lines for extract screening --- p.22 / Chapter 2.4.1 --- Cell lines and culturing condition --- p.22 / Chapter 2.4.2 --- Viable cell count using trypan blue exclusion method --- p.22 / Chapter 2.4.3 --- Anti-proliferation assay on SV7 tert using MTT assay --- p.23 / Chapter 2.4.4 --- Determination of the IC5o values --- p.24 / Chapter 2.4.5 --- Statistical Analysis --- p.24 / Chapter 2.5 --- Anti-proliferation assay on other cell lines using the two screened extracts --- p.24 / Chapter 2.5.1 --- Cell lines and culturing condition --- p.24 / Chapter 2.5.2 --- "Anti-proliferation assay on on HepG2, H292, Neuro2a,WIL2-NS cells using MTT assay" --- p.25 / Chapter 2.6 --- Proteomic studies for SV7tert and Hs68 after the treatment of extracts --- p.25 / Chapter 2.6.1 --- Protein sample preparation of SV7tert and Hs68 --- p.25 / Chapter 2.6.2 --- Protein quantification --- p.26 / Chapter 2.6.3 --- 2D Gel electrophoresis --- p.26 / Chapter 2.6.4 --- Image analysis --- p.26 / Chapter 2.7 --- Western Immunoblotting --- p.26 / Chapter 2.7.1 --- Protein sample preparation of SV7tert and Hs68 --- p.26 / Chapter 2.7.2 --- SDS-PAGE --- p.27 / Chapter 2.7.3 --- Protein Blotting --- p.27 / Chapter 2.7.4 --- Membrane Blocking and Antibody Incubations --- p.28 / Chapter 2.7.5 --- Detection of Proteins --- p.28 / Chapter 3 --- Results --- p.29 / Chapter 3.1 --- Proteins identification in Cordyceps militaris --- p.29 / Chapter 3.1.1 --- 2D gel electrophoresis analysis and resolution --- p.29 / Chapter 3.1.2 --- Identification and categorization of proteins of mycelia and fruiting body of Cordyceps militaris --- p.30 / Chapter 3.2 --- Investigation of anti-proliferating activity of extracts using MTT assays on SV7tert and Hs68 cell lines --- p.44 / Chapter 3.2.1 --- Mycelia extract from Cordyceps militaris --- p.44 / Chapter 3.2.2 --- Fruiting body extract from Cordyceps militaris --- p.44 / Chapter 3.2.3 --- Mycelia extract from Cordyceps sinensis --- p.47 / Chapter 3.2.4 --- Fruiting body extract from Cordyceps sinensis --- p.47 / Chapter 3.2.5 --- Screening of extracts --- p.50 / Chapter 3.3 --- "Investigation of anti-proliferating activity of extracts using MTT assays on HepG2,H292, Neuro2a and WIL2-NS cell lines" --- p.51 / Chapter 3.3.1 --- Mycelia extract from Cordyceps militaris --- p.51 / Chapter 3.3.2 --- Fruiting body extract from Cordyceps militaris --- p.51 / Chapter 3.4 --- Changes in total protein expression profiles in SV7tert and Hs68 cell lines --- p.56 / Chapter 3.4.1 --- Corresponding extract treatment of cell lines --- p.56 / Chapter 3.4.2 --- 2D gel electrophoresis analysis of protein from cells (SV7tert or Hs68) --- p.56 / Chapter 3.4.2.1 --- SV7tert study --- p.57 / Chapter 3.4.2.2 --- Hs68 study --- p.57 / Chapter 3.4.3 --- Protein identification --- p.65 / Chapter 3.4.3.1 --- Changes in protein expressions in SV7tert after mycelia extract treatment --- p.65 / Chapter 3.4.3.2 --- Changes in protein expressions in Hs68 after mycelia extract treatment --- p.65 / Chapter 3.4.3.3 --- Changes in protein expressions in SV7tert after fruiting body extract treatment --- p.66 / Chapter 3.4.3.4 --- Changes in protein expressions in Hs68 after fruiting body extract treatment --- p.66 / Chapter 3.5 --- Western immunoblotting --- p.71 / Chapter 3.5.1 --- Corresponding extract treatment of cell lines --- p.71 / Chapter 3.5.2 --- Normalization of protein loading using anti-actin antibody --- p.73 / Chapter 3.5.3 --- Detection of caspase 3 by use of antibody --- p.74 / Chapter 3.5.4 --- Detection of cleaved caspase 3 by use of antibody --- p.74 / Chapter 4 --- Discussion --- p.77 / Chapter 4.1 --- Identification of proteins in Cordyceps militaris --- p.77 / Chapter 4.2 --- Difficulties in identifying the proteins in Cordyceps militaris --- p.80 / Chapter 4.3 --- Investigation of anti-proliferating activity of extracts --- p.80 / Chapter 4.4 --- Changes in cell total protein expression profiles in SV7tert and Hs68 cell lines --- p.81 / Chapter 4.4.1 --- Protein alterations in SV7tert treated with mycelia extract --- p.82 / Chapter 4.4.1.1 --- Heat shock protein 27 (Hsp27) --- p.82 / Chapter 4.4.1.2 --- Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) --- p.83 / Chapter 4.4.2 --- Protein alterations in Hs68 with mycelia extract treatment --- p.84 / Chapter 4.4.2.1 --- Chain B of triosephosphate isomerase - Triosephophate isomerase 1 --- p.84 / Chapter 4.4.2.2 --- Glutathione transferase --- p.85 / Chapter 4.4.3 --- Protein alterations in SV7tert with fruiting body extract treatment --- p.86 / Chapter 4.4.3.1 --- Calreticulin precusor --- p.86 / Chapter 4.4.3.2 --- Nucleophosmin 1 isoform 2 (B23) --- p.87 / Chapter 4.4.3.3 --- Heat shock 70kDa protein 8 isoform 1 - Heat shock 70kDa protein (Hsp70) --- p.88 / Chapter 4.4.3.4 --- Voltage-dependent anion channel 2 (VDAC2) --- p.89 / Chapter 4.4.3.5 --- "Tumor protein, translationally controlled (TCTP)" --- p.90 / Chapter 4.4.3.6 --- RAN binding protein 1 (RANBP1) --- p.91 / Chapter 4.4.4 --- Protein alteration in Hs68 with mycelia extract treatment --- p.92 / Chapter 4.5 --- Conclusion --- p.93 / References --- p.94

Identiferoai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_326386
Date January 2008
ContributorsLai, Sze Tsai., Chinese University of Hong Kong Graduate School. Division of Molecular Biotechnology.
Source SetsThe Chinese University of Hong Kong
LanguageEnglish, Chinese
Detected LanguageEnglish
TypeText, bibliography
Formatprint, xv, 104 leaves : ill. (some col.) ; 30 cm.
RightsUse 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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