Common fragile site genes CNTLN and LINGO2 are associated with increased genome instability in different tumors [Elektronische Ressource] / presented by Sarah Zahedi Hamedani

Sarah Zahedi

Sujets

• Biologie

Informations

• Publié par: ruprecht-karls-universitat_heidelberg
• Publié le: 01 janvier 2010
• Nombre de lectures: 43
• Langue: English
• Poids de l'ouvrage: 1 Mo

Extrait

INAUGURAL-DISSERTATION



Submitted to the
Combined Faculties for the Natural Sciences and for Mathemathics
Of the Ruprecht-Karls University of Heidelberg
For the degree of
Doctor of Natural Sciences



Presented by
Sarah Zahedi Hamedani
born in Tehran


Date of oral examination:
15.12. 2010







Common fragile site genes, CNTLN and LINGO2, are
associated with increased genome instability in
different tumors








Referees: Prof. Dr. Gert Fricker
Prof. Dr. Manfred Schwab









To my parents, for their love
and to my brother, Sahab
















Since I've been…
Table of Contents



Table of Contents


ITable of contents
IVAcknowledgement
VAbstract
VIZusammenfassung
VIIList of abbreviations


1. Introduction ______________________________ _______________1

1.1 Genomic instability in cancer 1
1.2 Fragile sites 2
1.2.1 Rare fragile sites 3
1.2.2 Common fragile sites 3
1.2.2.1 Genes at common fragile sites 4
1.2.2.2 Instability at common fragile sites 6
1.2.2.2.1 Common fragile site instability in vitro 6
1.2.2.2.2 Comm in cancer 6
1.2.2.3 Mechanism of instability at common fragile site 11
1.2.2.3.1 Unstable sequences and late replication at CFSs 11
1.2.2.3.2 Cell cycle checkpoints and repair pathways in common fragile site
instability 13
1.2.2.4 Evolutionary conservation of common fragile sites 16

1.3 Genome instability on the short arm of chromosome 9 17
1.3.1 FRA9G 18
1.3.2 FRA9C 19
1.4 Aims of this study 22



2. Materials and Methods ____________________________ __23
232.1 Materials
23 2.1.1 Chemicals
I
Table of Contents



24 2.1.2 Other Materials
24 2.1.3 Laboratory Equipments
26 2.1.4 Antibiotics
26 2.1.5 Enzymes
26 2.1.6 Fluorescent dyes
26 2.1.7 Kits, Media and Solution
27 2.1.8 Nucleic Acids
29 2.1.9 Buffers and Solution
30 2.1.10 Oligonucleotides
31 2.1.11 Plasmids
31 2.1.12 Software
31 2.1.13 Online Databases and Programs
32 2.1.14 Human tumor cell lines
33 2.1.15 Lymohpblastoid cell lines from healthy individuals
342.2 Methods 34
34 2.2.1 Array CGH
36 2.2.2 Cytogenetic techniques
41 2.2.3 Nucleic Acid manipulations
41 2.2.3.1 Total RNA extraction
41 2.2.3.2 Reserve transcription of total RNA and RT-PCR
42 2.2.3.3 Quantitative RT-PCR
43 2.2.3.4 Genomic DNA extraction
442.2.4 Culture of cells
442.2.5 Freezing and thawing of cells
442.2.6 5-Azactytidine treatment



3. Results _______________________________________________45
3.1 Chromosomal rearrangement on 9p in different tumor types 45
3.1.1 Copy number changing profiles 45
3.1.2 FRA9G, CDKN2A and FRA9C are breakpoint-clustering loci on 9p 48
3.1.3 FRA9G, and FRA9C demonstrate multiple rearrangements within their 50
sequence in tumor cells
II
Table of Contents



3.1.3.1 FRA9G 51
3.1.3.2 CDKN2A 52
3.1.3.3 FRA9C 53
3.1.3.3.1 Fluorescence in situ hybridization (FISH) confirms aCGH data 55
3.1.4 Differentiated pattern of 9p rearrangements in four tumor types 58