The Influence of Oxidative Stress, Carcinogens and Cloning on DNA ...

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Index 1 Index 1 Introduction ........................................................................................................................ 1 1.1 Epigenetics ................................................................................................................. 1 1.1.1 Histone modification .............................................................................................. 1 1.1.2 DNA methylation ................................................................................................... 2 1.2 <strong>The</strong> role <strong>of</strong> DNA methylation .................................................................................... 4 1.2.1 Gene expression ..................................................................................................... 4 1.2.2 Chromatin structure <strong>and</strong> transcription repression .................................................. 5 1.2.3 Genomic imprinting ............................................................................................... 6 1.2.4 X-chromosome inactivation ................................................................................... 7 1.2.5 Cancer ..................................................................................................................... 7 1.2.5.1 DNA methylation: Effective biomarker for tumourigenesis .......................... 9 1.2.6 In-vitro fertilization (IVF) .................................................................................... 11 1.2.7 <strong>Cloning</strong> ................................................................................................................. 11 1.2.7.1 <strong>Cloning</strong> efficiency ........................................................................................ 12 1.2.7.2 Effect <strong>of</strong> cloning on epigenetics ................................................................... 13 1.2.8 Twins <strong>and</strong> epigenetics overview .......................................................................... 14 1.3 DNA adducts ............................................................................................................ 15 1.3.1 DNA adducts <strong>and</strong> cancer ...................................................................................... 15 1.3.2 Endogenous adducts ............................................................................................. 16 1.3.2.1 <strong>Oxidative</strong> stress ............................................................................................ 20 1.3.2.2 8-oxoguanosine ............................................................................................ 23 1.3.2.3 Methods <strong>of</strong> analysis <strong>of</strong> 8-oxoguanosine ....................................................... 23 1.3.3 Exogenous DNA adducts ..................................................................................... 24 1.3.3.1 Environmental chemicals ............................................................................. 24 1.3.3.2 Aristolochic acid (AA) ................................................................................. 29 1.3.3.3 Methods for the determination <strong>of</strong> AA .......................................................... 32 1.4 Methods for determination <strong>of</strong> the DNA-methylation level ...................................... 33 1.4.1 Gene-specific methylation analysis methods ....................................................... 33 1.4.2 Genomic-wide methylation analysis methods ...................................................... 37 1.4.3 Capillary Electrophoresis (CE) ............................................................................ 39 2 Aim <strong>and</strong> objectives <strong>of</strong> the study ....................................................................................... 40 3 Results <strong>and</strong> Discussion ..................................................................................................... 41
Page 1 and 2: The Influe
Page 3 and 4: The work at h<stro
Page 5 and 6: First, I feel always grateful to Al
Page 7: I would like to extend my thanks to
Page 11 and 12: The following acro
Page 13 and 14: FDA Food and drug
Page 15: RIA Radioimmunoassay ROS Reactive o
Page 19 and 20: 3 Index 3.5.4.1 Quantification <str
Page 21 and 22: 1 Introduction 1.1 Epigenetics 1 In
Page 23 and 24: 3 Introduction Most CpG isl
Page 25 and 26: 5 Introduction regions can suppress
Page 27 and 28: 7 Introduction This is due to the n
Page 29 and 30: 9 Introduction Tumour suppressor ge
Page 31 and 32: 1.2.6 In-vitro fertilization (IVF)
Page 33 and 34: 13 Introduction in normal embryos d
Page 35 and 36: 1.3 DNA adducts 1.3.1 DNA adducts <
Page 37 and 38: 17 Introduction Oxidation, methylat
Page 39 and 40: CH 3 N N H O N NH 7-methylguanine H
Page 41 and 42: 21 Introduction take (221, 222) <st
Page 43 and 44: 1.3.2.2 8-oxoguanosine 23 Introduct
Page 45 and 46: HO O OH N N DNA B[a]P B[a]P-7,8-dih
Page 47 and 48: O O N-OH-3-ABA O O NH NO 2 3-NBA Ni
Page 49 and 50: 1.3.3.2 Aristolochic acid (AA) 29 I
Page 51 and 52: 31 Introduction phologically, both
Page 53 and 54: 33 Introduction method had been use
Page 55 and 56: 35 Introduction cating a methylated
Page 57 and 58: 37 Introduction cleotide primer ext
Page 59 and 60: 39 Introduction depurination is an
Page 61 and 62: 3 Results and Disc
Page 63 and 64: 43 Results and Dis
Page 65 and 66: 45 Results and Dis
Page 67 and 68:
47 Results and Dis
Page 69 and 70:
Rel. fluorescence [RFU] Rel. fluore
Page 71 and 72:
Results and Discus
Page 73 and 74:
53 Results and Dis
Page 75 and 76:
O 3P HO O 3P O O 3P N N Sugar N N S
Page 77 and 78:
Results and Discus
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59 Results and Dis
Page 81 and 82:
Rel. fluorescence [RFU] 350 300 250
Page 83 and 84:
63 Results and Dis
Page 85 and 86:
Rel. fluorescence [RFU] 8-oxo-dGMP
Page 87 and 88:
67 67 Results and
Page 89 and 90:
3.4.6 Study of the
Page 91 and 92:
71 Results and Dis
Page 93 and 94:
73 Results and Dis
Page 95 and 96:
75 Results and Dis
Page 97 and 98:
77 Results and Dis
Page 99 and 100:
S within 79 Results and</st
Page 101 and 102:
81 Results and Dis
Page 103 and 104:
3.5.2 Influence <s
Page 105 and 106:
Liver Lung Kidney 3 Weeks Control 3
Page 107 and 108:
Results and Discus
Page 109 and 110:
3.5.2.4 Correlation between the add
Page 111 and 112:
Kidney (2 mg/kg BW) Bladder (2 mg/k
Page 113 and 114:
Small intestine Liver Lungs 2 3.65
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95 Results and Dis
Page 117 and 118:
97 Results and Dis
Page 119 and 120:
3.5.4.3 Computation and</st
Page 121 and 122:
Description Mean values comparison
Page 123 and 124:
Methylation level [%] 5.2 5.1 5 4.9
Page 125 and 126:
Sample Code K1 K2 K3 K4 K5 K6 K7 K8
Page 127 and 128:
107 107 Results and</strong
Page 129 and 130:
Page 131 and 132:
Page 133 and 134:
Sample Code Type State Methylation
Page 135 and 136:
Page 137 and 138:
117 Summary and le
Page 139 and 140:
5 Materials and Me
Page 141 and 142:
5.2 Experimental preparations 5.2.1
Page 143 and 144:
� Preparation of
Page 145 and 146:
) MN/SPD mixture with 15 times conc
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127 Experimental part umn. After ce
Page 149 and 150:
5.3.1.4 CE-LIF measurement 129 Expe
Page 151 and 152:
5.3.4 Preparation of</stron
Page 153 and 154:
133 Experimental part Hydrolysis <s
Page 155 and 156:
5.4.5 SPE system specifications are
Page 157 and 158:
6 Appendix Glossary 137 Appendix Ab
Page 159 and 160:
A brief history of
Page 161 and 162:
Hydrolysis buffer (250 mM HEPES, 10
Page 163 and 164:
Which Pipettes for derivatisation w
Page 165 and 166:
Removal of polyimi
Page 167 and 168:
147 Appendix Figure 60: LC chromato
Page 169 and 170:
7 List of figures
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151 List of figure
Page 173 and 174:
153 List of tables
Page 175 and 176:
155 Literature 20. Smiraglia, D.J.;
Page 177 and 178:
157 Literature 58. Kawahara, M.; Wu
Page 179 and 180:
159 Literature squamous cell carcin
Page 181 and 182:
161 Literature 130. Bird, A.; "DNA
Page 183 and 184:
163 Literature 167. Poirier, M.C. <
Page 185 and 186:
165 Literature 202. Thannickal, V.J
Page 187 and 188:
167 Literature 240. Pedersen, B.K.;
Page 189 and 190:
169 Literature Dna Samples by Disso
Page 191 and 192:
171 Literature 306. Kress, S.; Jahn
Page 193 and 194:
173 Literature 342. Mengs, U.; "Acu
Page 195 and 196:
175 Literature 377. Clark, S.J.; Ha
Page 197 and 198:
177 Literature 414. Everaerts, F.M.
Page 199 and 200:
179 Literature 450. Teixeira, H.D.
Page 201 and 202:
181 Literature 484. Grollman, A.P.;
Page 203 and 204:
183 Literature 519. Zakhartchenko,
Page 205 and 206:
Curriculum Vitae Personal informati
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