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Cytogenomics
 ­Cytogenomics

Edited by

Thomas Liehr
Jena University Hospital, Friedrich Schiller University,
Institute of Human Genetics, Jena, Germany
http://cs-tl.de/
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Contents

Contributors.............................................................................................................. xv

CHAPTER 1 A definition for cytogenomics - Which also

may be called chromosomics...........................................1
Thomas Liehr
From cytogenetics to cytogenomics................................................ 1
A definition of cytogenomics........................................................... 2
Conclusion....................................................................................... 5
References........................................................................................ 5

SECTION 1 Technical aspects

CHAPTER 2 Overview of currently available approaches
used in cytogenomics......................................................11
Thomas Liehr
What is cytogenomics?.................................................................. 11
Cytogenomic approaches............................................................... 12
Before the word "genetics" was defined.................................... 16
Cytogenetics.............................................................................. 16
Molecular genetics.................................................................... 17
Molecular cytogenetics.............................................................. 17
Epigenetics................................................................................ 19
Conclusion..................................................................................... 20
References...................................................................................... 20
CHAPTER 3 Cytogenetics......................................................................25
Anja Weise and Thomas Liehr
What is cytogenetics?.................................................................... 25
A short history of cytogenetics...................................................... 28
Material and methods in cytogenetics........................................... 29
Material applied in cytogenetics................................................ 29
Cytogenetic methods................................................................. 29
Advantages and restrictions........................................................... 30
Applications................................................................................... 31
Human genetic diagnostics........................................................ 31
Cytogenetic-based research....................................................... 31
Conclusion..................................................................................... 32
References...................................................................................... 33

v
vi Contents

CHAPTER 4 Molecular cytogenetics...................................................35

Thomas Liehr
What is molecular cytogenetics?................................................... 35
Probes applied in molecular cytogenetics and how
to get them.................................................................................. 36
Locus-specific probes................................................................ 36
Repetitive probes....................................................................... 38
Partial chromosome paints........................................................ 38
Whole chromosome paints........................................................ 39
Whole genome........................................................................... 39
Probe-sets applied in molecular cytogenetics................................ 39
Different resolutions in molecular cytogenetics............................ 39
Cytogenomic applications of molecular cytogenetics................... 40
Conclusion..................................................................................... 41
References...................................................................................... 41

CHAPTER 5 Molecular combing solutions to characterize

replication kinetics and genome
rearrangements.................................................................47
Prakhar Bisht and Mario Davide Maria Avarello
Introduction.................................................................................... 47
Basic studies on DNA replication.................................................. 50
Replication kinetic studies by molecular combing........................ 51
Replication kinetic studies in locus-specific manner..................... 55
Genomic Morse Code (GMC) and its applications....................... 58
GMC designs for CNV studies...................................................... 58
GMC designs for sequence-specific alterations............................. 59
GMC design for gene editing studies............................................ 61
Conclusion..................................................................................... 64
References...................................................................................... 65

CHAPTER 6 Molecular karyotyping.....................................................73

Anja Weise and Thomas Liehr
Background.................................................................................... 73
Advantages and restrictions........................................................... 75
Material applied in molecular karyotyping.................................... 78
Microarrays in molecular karyotyping.......................................... 79
Applications................................................................................... 80
Molecular karyotyping approach in human genetic
diagnostics............................................................................. 80
 Contents vii

Molecular karyotyping approach in cytogenomic

research.................................................................................. 80
Conclusion..................................................................................... 81
References...................................................................................... 82
CHAPTER 7 Sequencing approaches..................................................87
Martin Ungelenk
Introduction.................................................................................... 87
The first generation........................................................................ 88
Maxam and Gilbert’s chemical cleavage................................... 88
Sanger sequencing..................................................................... 90
Further advances in Sanger sequencing.................................... 90
The second generation................................................................... 92
Pyrosequencing......................................................................... 95
Polony sequencing..................................................................... 95
The Applied Biosystem SOLiD platform.................................. 96
Ion torrents pH measurements with semiconductors................ 97
Illumina’s sequence-by-synthesis approach.............................. 98
Complete genomics/BGI combinatorial probe anchor
synthesis.............................................................................. 100
Third generation........................................................................... 101
Single-molecule real-time sequencing.................................... 101
Nanopore sequencing.............................................................. 103
Fourth generation......................................................................... 104
Complementary methods............................................................. 105
Single-cell sequencing............................................................. 105
Optical mapping...................................................................... 106
Sequencing by hybridization................................................... 106
Summary and conclusion............................................................. 107
References.................................................................................... 116
CHAPTER 8 Next-generation cytogenomics: High-resolution
structural variation detection by optical genome
mapping............................................................................ 123
Yannick Delpu, Hayk Barseghyan, Sven Bocklandt,
Alex Hastie, and Alka Chaubey
Introduction.................................................................................. 123
Introduction to Optical Genome Mapping................................... 125
General principle..................................................................... 125
Structural and copy number variation detection...................... 127
Data visualization, filtering, and reviewing............................. 129
viii Contents

Applications in constitutional cytogenetics................................. 132

Prenatal cytogenetics............................................................... 132
Postnatal cytogenetics............................................................. 134
Applications in hematological malignancies............................... 137
Clinical benchmark studies..................................................... 138
Novel SV identification........................................................... 140
Application in solid tumors......................................................... 141
Concluding remarks..................................................................... 142
References.................................................................................... 143

CHAPTER 9 Application of CRISPR/Cas9 to visualize defined

genomic sequences in fixed chromosomes and
nuclei................................................................................ 147
Takayoshi Ishii, Kiyotaka Nagaki, and Andreas Houben
Introduction.................................................................................. 147
Application of CRISPR-FISH..................................................... 148
Funding information.................................................................... 152
References.................................................................................... 152

CHAPTER 10 Approaches for studying epigenetic aspects

of the human genome..................................................... 155
Tigran Harutyunyan and Galina Hovhannisyan
Introduction.................................................................................. 155
Analysis of DNA methylation..................................................... 157
Principles of DNA methylation analysis................................. 158
Global DNA methylation........................................................ 160
Genome-wide DNA methylation............................................. 168
Gene-specific methylation....................................................... 171
Summary for methylated DNA detection................................ 174
Analysis of histone modifications................................................ 177
Western blot............................................................................. 178
Immunofluorescence assay...................................................... 179
Chromatin immunoprecipitation assay.................................... 180
Mass spectrometry assay......................................................... 182
Summary for histone modifications........................................ 183
Higher-order chromatin structure analysis.................................. 183
Chromatin remodeling............................................................. 183
3D nucleus organization and epigenetic regulation................. 185
Analysis of noncoding RNAs...................................................... 188
Real-time quantitative reverse transcription PCR................... 189
Messenger RNA sequencing........................................................ 191
 Contents ix

Concluding remarks..................................................................... 192

Acknowledgment......................................................................... 193
References.................................................................................... 193

SECTION 2 Current cytogenomic research

CHAPTER 11 Chromoanagenesis phenomena and their
formation mechanisms................................................... 213
Franck Pellestor, Jean-Baptiste Gaillard, Anouck Schneider,
Jacques Puechberty, and Vincent Gatinois
Introduction.................................................................................. 213
An overview of chromoanagenesis phenomena: Definitions
and hallmarks........................................................................... 214
Chromothripsis........................................................................ 214
Chromoplexy........................................................................... 216
Chromoanasynthesis................................................................ 217
Mechanisms for chromoanagenesis occurrence.......................... 218
The micronucleus-mediated model......................................... 219
The chromatin bridge model................................................... 224
Factors promoting the emergence of
chromoanagenesis.................................................................... 227
Conclusion................................................................................... 231
References.................................................................................... 231

CHAPTER 12 3D cytogenomics: Structural variation in the

three-dimensional genome............................................ 247
Veronica Yumiceba, Uirá Souto Melo, and
Malte Spielmann
Introduction.................................................................................. 247
3D genome: Hierarchical organization of genetic information
in the nucleus...........................................................................248
Topologically associating domains.............................................. 249
Techniques to detect chromatin interactions in the
nuclear space............................................................................ 250
Features of a Hi-C map................................................................ 251
Position effects and TADs............................................................ 253
Intra-TAD structural variants..................................................254
Inter-TAD structural variants: Enhancer hijacking.................255
SVs detection in the 3D genome................................................. 259
Clinical application and future perspectives................................ 260
References.................................................................................... 261
x Contents

CHAPTER 13 Multilayer organization of chromosomes.................... 267

Joan-Ramon Daban
Introduction.................................................................................. 268
Experimental approaches to study the internal structure
of mitotic chromosomes........................................................... 268
Chromatin plates in aqueous solution.......................................... 271
Self-organization of multilayer chromatin................................... 274
Stacked thin layers explain the morphology of bands and
chromosome rearrangements................................................... 276
Cytogenetic map and genome sequence.................................. 276
Chromosome bands................................................................. 278
Thin bands............................................................................... 279
Band splitting.......................................................................... 279
Replication bands.................................................................... 280
Sister chromatid exchanges..................................................... 280
Chromosome translocations.................................................... 280
The topological organization of DNA is preserved in
fixed chromosomes.............................................................. 282
Functional implications of multilayered chromosomes............... 282
Planar chromatin in interphase................................................ 282
Chromosome bands in interphase...........................................284
Topologically associating domains......................................... 284
Gene expression...................................................................... 285
DNA replication...................................................................... 286
DNA repair.............................................................................. 287
Concluding remarks..................................................................... 287
References.................................................................................... 288

CHAPTER 14 Nuclear architecture...................................................... 297

Thomas Liehr
Background.................................................................................. 297
Short history of nucleomics......................................................... 298
1888 to 1950s.......................................................................... 298
1950s to 1980s......................................................................... 298
1980s to present....................................................................... 299
Present insights into nuclear architecture.................................... 301
Conclusion................................................................................... 302
References.................................................................................... 303
 Contents xi

CHAPTER 15 Nuclear stability in early embryo. Chromosomal

aberrations....................................................................... 307
Vladislav S. Baranov and Tatiana V. Kuznetzova
Introduction.................................................................................. 307
The nucleus of the zygote............................................................ 308
Zygotes and blastomeres......................................................... 308
Primary differentiation and blastocyst formation.................... 310
Chromosomal abnormalities........................................................ 311
Methods of karyotype analysis................................................ 311
Chromosomal abnormalities in early embryogenesis............. 312
Genomic imprinting................................................................ 314
Chromosomal mosaicism........................................................ 315
Chromothripsis........................................................................ 318
Conclusion................................................................................... 319
Acknowledgments....................................................................... 320
References.................................................................................... 320
CHAPTER 16 Cytogenomic landscape of the human brain.............. 327
Ivan Y. Iourov, Svetlana G. Vorsanova, and Yuri B. Yurov
Introduction to molecular neurocytogenetics/
neurocytogenomics.................................................................. 327
Cytogenomic variation in the human brain: Neuronal
diversity versus disease............................................................ 328
Cyto(onto)genomic variations in the unaffected brain............ 329
Neurocytogenomic variations and brain diseases.................... 330
Chromosomal organization in the human brain: Is there
a postgenomic perspective?...................................................... 335
Pathway-based analysis of neurocytogenomic landscape...........337
Concluding remarks: What is and what should be...................... 338
What is..................................................................................... 338
What should be........................................................................ 338
Acknowledgments....................................................................... 340
References.................................................................................... 340
CHAPTER 17 Interchromosomal interactions with meaning
for disease....................................................................... 349
Anja Weise and Thomas Liehr
Background.................................................................................. 349
Methods used in interchromosomal nucleomics.......................... 350
xii Contents

Insights from two own studies..................................................... 351

Manvelyan et al. (2009) and Othman et al. (2012).................. 351
Maass et al. (2018).................................................................. 351
Outlook........................................................................................ 352
References.................................................................................... 353
CHAPTER 18 Shaping of genome by long noncoding RNAs............. 357
Yuichi Ichikawa and Noriko Saitoh
Introduction.................................................................................. 357
Technologies to determine genome-wide RNA-chromatin
interactions............................................................................... 360
Chromatin isolation by RNA purification (ChIRP),
followed by high-throughput sequencing (ChIRP-seq)....... 360
Capture hybridization analysis of RNA targets....................... 360
Proximity ligation-based methods like RADICL-seq............. 361
Nuclear lncRNAs that recruit chromatin modifiers..................... 362
Nuclear lncRNAs that promote long-range chromatin
interactions............................................................................... 363
Interchromosomal interactions via subnuclear structures............ 364
Breast cancer-specific lncRNAs regulate a long-range
chromatin interaction............................................................... 366
Conclusion................................................................................... 367
Acknowledgments....................................................................... 368
References.................................................................................... 368
CHAPTER 19 Repetitive elements, heteromorphisms, and copy
number variants............................................................... 373
Thomas Liehr
Background.................................................................................. 373
Types of polymorphic DNA......................................................... 374
Phenotypically visible polymorphisms................................... 374
Single nucleotide polymorphisms........................................... 378
Microsatellites......................................................................... 379
Small-scale insertions/inversions/deletions/duplications........ 380
Minisatellites........................................................................... 381
Small-scale repetitive elements............................................... 381
Submicroscopic copy number variants (CNVs)...................... 382
Chromosomal heteromorphisms............................................. 383
Euchromatic variants............................................................... 384
Chromosomal and whole genomic variants............................ 384
Insights and conclusions.............................................................. 385
References.................................................................................... 386
 Contents xiii

CHAPTER 20 Epigenetics...................................................................... 389

Thomas Eggermann
Introduction.................................................................................. 389
Epigenetic regulation and chromatin organization...................... 390
Epigenetic landscape during development.............................. 390
The maternal and the paternal genomes are functionally
disparate in higher mammals................................................... 393
Disturbances of the subcortical maternal complex...................... 393
The imprinted regions in 11p15.5 as an example of the
interaction between local chromatin regulation and local
chromatin organization........................................................ 395
Conclusion and outlook............................................................... 397
References.................................................................................... 397

Subject Index.......................................................................................................... 403

This page intentionally left blank
Contributors

Mario Davide Maria Avarello

Genomic Vision, Bagneux, France
Vladislav S. Baranov
D.O. Ott Research Institute of Obstetrics, Gynecology and Reproductology, Saint
Petersburg, Russia
Hayk Barseghyan
Bionano Genomics, San Diego, CA; Children’s National Research Institute,
Washington, DC, United States
Prakhar Bisht
Genomic Vision, Bagneux, France
Sven Bocklandt
Bionano Genomics, San Diego, CA, United States
Alka Chaubey
Bionano Genomics, San Diego, CA, United States
Joan-Ramon Daban
Department of Biochemistry and Molecular Biology, Faculty of Biosciences,
Autonomous University of Barcelona, Barcelona, Spain
Yannick Delpu
Bionano Genomics, San Diego, CA, United States
Thomas Eggermann
Uniklinik RWTH Aachen, Institut für Humangenetik, Aachen, Germany
Jean-Baptiste Gaillard
Unit of Chromosomal Genetics and Research Platform Chromostem,
Department of Medical Genetics, Arnaud de Villeneuve Hospital, Montpellier,
France
Vincent Gatinois
Unit of Chromosomal Genetics and Research Platform Chromostem,
Department of Medical Genetics, Arnaud de Villeneuve Hospital; INSERM 1183
Unit “Genome and Stem Cell Plasticity in Development and Aging” Institute of
Regenerative Medicine and Biotherapies, St Eloi Hospital, Montpellier, France
Tigran Harutyunyan
Department of Genetics and Cytology, Yerevan State University, Yerevan,
Armenia
Alex Hastie
Bionano Genomics, San Diego, CA, United States

xv
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