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• Cover
• Half Title
• Title
• Copyright
• Contents
• Preface
• Acknowledgements
• 1 Fundamentals of DNA, Chromosomes, and Cells
• 1.1 The Structure and Function of Nucleic Acids
• General concepts: the genetic material, genomes, and genes
• The underlying chemistry of nucleic acids
• Base pairing and the double helix
• DNA replication and DNA polymerases
• Genes, transcription, and the central dogma of molecular biology
• 1.2 The Structure and Function of Chromosomes
• Why we need highly structured chromosomes, and how they are organized
• Chromosome function: replication origins, centromeres, and telomeres
• 1.3 DNA and Chromosomes in Cell Division and the Cell Cycle
• Differences in DNA copy number between cells
• The cell cycle and segregation of replicated chromosomes and DNA molecules
• Mitosis: the usual form of cell division
• Meiosis: a specialized reductive cell division giving rise to sperm and egg cells
• Why each of our gametes is unique
• Summary
• Questions
• Further Reading
• 2 Fundamentals of Gene Structure, Gene Expression, and Human Genome Organization
• 2.1 Protein-Coding Genes: Structure and Expression
• Gene organization: exons and introns
• RNA splicing: stitching together the genetic information in exons
• Translation: decoding messenger RNA to make a polypeptide
• From newly synthesized polypeptide to mature protein
• 2.2 RNA Genes and Noncoding RNA
• The extraordinary secondary structure and versatility of RNA
• RNAs that act as specific regulators: from quirky exceptions to the mainstream
• 2.3 Working Out the Details of Our Genome and What They Mean
• The Human Genome Project: working out the details of the nuclear genome
• What the sequence didn’t tell us and the goal of identifying all functional human DNA sequences
• 2.4 A Quick Tour of Some Electronic Resources Used to Interrogate the Human Genome Sequence and Gene Products
• Gene nomenclature and the HGNC gateway
• Databases storing nucleotide and protein sequences
• Finding related nucleotide and protein sequences
• Links to clinical databases
• 2.5 The Organization and Evolution of the Human Genome
• A brief overview of the evolutionary mechanisms that shaped our genome
• How much of our genome is functionally significant?
• The mitochondrial genome: economical usage but limited autonomy
• Gene distribution in the human genome
• The extent of repetitive DNA in the human genome
• The organization of gene families
• The significance of gene duplication and repetitive coding DNA
• Highly repetitive noncoding DNA in the human genome
• Summary
• Questions
• Further Reading
• 3 Principles Underlying Core DNA Technologies
• 3.1 Amplifying DNA by DNA Cloning
• Amplifying desired DNA within bacterial cells
• The need for vector DNA molecules
• Physical clone separation
• The need for restriction nucleases
• DNA libraries and the uses and limitations of DNA cloning
• 3.2 Amplifying DNA Using the Polymerase Chain Reaction (PCR)
• Basics of the polymerase chain reaction (PCR)
• Quantitative PCR and real-time PCR
• 3.3 Principles of Nucleic Acid Hybridization
• Formation of artificial heteroduplexes
• Hybridization assays: using known nucleic acids to find related sequences in a test nucleic acid population
• Microarray hybridization: large-scale parallel hybridization to immobilized probes
• 3.4 Principles of DNA Sequencing
• Dideoxy DNA sequencing
• Massively parallel DNA sequencing (next-generation sequencing)
• Summary
• Questions
• Further Reading
• 4 Principles of Genetic Variation
• 4.1 DNA Sequence Variation Origins and DNA Repair
• Genetic variation arising from errors in chromosome and DNA function
• Various endogenous and exogenous sources can cause damage to DNA by altering its chemical structure
• The wide range of DNA repair mechanisms
• Repair of DNA damage or altered sequence on a single DNA strand
• Repair of DNA lesions that affect both DNA strands
• Undetected DNA damage, DNA damage tolerance, and translesion synthesis
• 4.2 Population Genomics and the Scale of Human Genetic Variation
• DNA variants, polymorphisms, and human population genomics
• Small-scale variation: single nucleotide variants and small insertions and deletions
• Microsatellites and other variable number of tandem repeat (VNTR) polymorphisms
• Structural variation and low copy number variation
• Taking stock of human genetic variation
• 4.3 Functional Genetic Variation and Protein Polymorphism
• The vast majority of genetic variation has a neutral effect on the phenotype, but a small fraction is harmful
• Different types of Darwinian natural selection operate in human lineages
• Generating protein diversity by gene duplication: the example of olfactory receptor genes
• 4.4 Extraordinary Genetic Variation in the Immune System
• Pronounced genetic variation in four classes of immune system proteins
• Programmed and random post-zygotic genetic variation
• Somatic mechanisms allow cell-specific production of immunoglobulins and T-cell receptors
• MHC (HLA) proteins: functions and polymorphism
• The medical importance of the HLA system
• Summary
• Questions
• Further Reading
• 5 Single-Gene Disorders: Inheritance Patterns, Phenotype Variability, and Allele Frequencies
• 5.1 Introduction: Terminology, Electronic Resources, and Pedigrees
• Background terminology and electronic resources with information on single-gene disorders
• Investigating family history of disease and recording pedigrees
• 5.2 The Basics of Mendelian and Mitochondrial DNA Inheritance Patterns
• Autosomal dominant inheritance
• Autosomal recessive inheritance
• Sex-linked inheritance
• Matrilineal inheritance for mitochondrial DNA disorders
• 5.3 Uncertainty, Heterogeneity, and Variable Expression of Mendelian Phenotypes
• Difficulties in defining the mode of inheritance in small pedigrees
• Heterogeneity in the correspondence between phenotypes and the underlying genes and mutations
• Nonpenetrance and age-related penetrance
• 5.4 Allele Frequencies in Populations
• Allele frequencies and the Hardy-Weinberg law
• Applications and limitations of the Hardy-Weinberg law
• Ways in which allele frequencies change in populations
• Population bottlenecks and founder effects
• Mutation versus selection in determining allele frequencies
• Heterozygote advantage: when natural selection favors carriers of recessive disease
• Summary
• Questions
• Further Reading
• 6 Principles of Gene Regulation and Epigenetics
• The two fundamental types of gene regulation
• Cis-acting and trans-acting effects in gene regulation
• 6.1 Genetic Regulation of Gene Expression
• Promoters: the major on–off switches in genes
• Modulating transcription and tissue-specific regulation
• Transcription factor binding and specificity
• Genetic regulation during RNA processing: RNA splicing and RNA editing
• Translational regulation by trans-acting regulatory proteins
• Post-transcriptional gene silencing by microRNAs
• Repressing the repressors: competing endogenous RNAs sequester miRNA
• 6.2 Chromatin Modification and Epigenetic Factors in Gene Regulation
• An overview of the molecular basis of epigenetic mechanisms
• How changes in chromatin structure produce altered gene expression
• Histone modification and histone substitution in nucleosomes
• Modified histones and histone variants affect chromatin structure
• The function of DNA methylation in mammalian cells
• DNA methylation: mechanisms, heritability, and global roles during early development and gametogenesis
• Long noncoding RNAs in mammalian epigenetic regulation
• Genomic imprinting: differential expression of maternally and paternally inherited alleles
• X-chromosome inactivation: compensating for sex differences in gene dosage
• 6.3 Abnormal Epigenetic Regulation in Mendelian Disorders and Uniparental Disomy
• Principles of epigenetic dysregulation
• “Chromatin diseases” due to mutations in genes specifying chromatin modifiers
• Disease resulting from dysregulation of heterochromatin
• Uniparental disomy and disorders of imprinting
• Abnormal gene regulation at imprinted loci
• Summary
• Questions
• Further Reading
• 7 How Genetic Variation in DNA and Chromosomes Causes Disease
• 7.1 An Overview of How Genetic Variation Results in Disease
• The importance of repeat sequences in triggering pathogenesis
• 7.2 Pathogenic Nucleotide Substitutions and Tiny Insertions and Deletions
• Pathogenic single nucleotide substitutions within coding sequences
• Mutations that result in premature termination codons
• Genesis and frequency of pathogenic point mutations
• Surveying and curating point mutations that cause disease
• 7.3 Pathogenesis Due to Variation in Short Tandem Repeat Copy Number
• The two main classes of pathogenic variation in short tandem repeat copy-number
• Dynamic disease-causing mutations due to unstable expansion of short tandem repeats
• Unstable expansion of short tandem repeats can cause disease in different ways
• 7.4 Pathogenesis Triggered by Long Tandem Repeats and Interspersed Repeats
• Pathogenic exchanges between repeats occurs in both nuclear DNA and mtDNA
• Nonallelic homologous recombination and transposition
• Pathogenic sequence exchanges between chromatids at mispaired tandem repeats
• Disease arising from sequence exchanges between distantly located repeats in nuclear DNA
• 7.5 Chromosome Abnormalities
• Structural chromosomal abnormalities
• Chromosomal abnormalities involving gain or loss of complete chromosomes
• 7.6 Molecular Pathology of Mitochondrial Disorders
• Mitochondrial disorders due to mtDNA mutation show maternal inheritance and variable proportions of mutant genotypes
• The two major classes of pathogenic DNA variant in mtDNA: large deletions and point mutations
• 7.7 Effects on the Phenotype of Pathogenic Variants in Nuclear DNA
• Mutations affecting how a single gene works: an overview of loss of function and gain of function
• The effect of pathogenic variants depends on how the products of alleles interact: dominance and recessiveness revisited
• Gain-of-function and loss-of-function mutations in the same gene can produce different phenotypes
• Multiple gene dysregulation resulting from aneuploidies and mutations in regulatory genes
• 7.8 A Protein Structure Perspective of Molecular Pathology
• Pathogenesis arising from protein misfolding
• The many different ways in which protein aggregation can result in disease
• 7.9 Genotype–Phenotype Correlations and Why Monogenic Disorders are Often Not Simple
• The difficulty in getting reliable genotype–phenotype correlations
• Modifier genes and environmental factors: common explanations for poor genotype–phenotype correlations
• Summary
• Questions
• Further Reading
• 8 Identifying Disease Genes and Genetic Susceptibility to Complex Disease
• 8.1 Identifying Genes in Monogenic Disorders
• A historical overview of identifying genes in monogenic disorders
• Linkage analysis to map genes for monogenic disorders to defined subchromosomal regions
• Chromosome abnormalities and other large-scale mutations as routes to identifying disease genes
• Exome sequencing: let’s not bother getting a position for disease genes!
• 8.2 Approaches to Mapping and Identifying Genetic Susceptibility to Complex Disease
• The polygenic and multifactorial nature of common genetic disorders
• Difficulties with lack of penetrance and phenotype classification in complex disease
• Estimating heritability: the contribution made by genetic factors to the variance of complex diseases
• The very limited success of linkage analyses in identifying genes underlying complex genetic diseases
• The fundamentals of allelic association and the importance of HLA-disease associations
• Linkage disequilibrium as the basis of allelic associations
• How genomewide association studies are carried out
• Moving from candidate subchromosomal region to identify causal genetic variants in complex disease can be challenging
• The limitations of GWA studies and the issue of missing heritability
• Alternative genome-wide studies and the role of rare variants and copy number variants in complex disease
• The assessment and prediction of risk for common genetic diseases and the development of polygenic risk scores
• 8.3 Aspects of the Genetic Architecture of Complex Disease and the Contributions of Environmental and Epigenetic Factors
• Common neurodegenerative disease: from monogenic to polygenic disease
• The importance of immune system pathways in common genetic disease
• The importance of protective factors and how a susceptibility factor for one complex disease may be a protective factor for another disease
• Gene–environment interactions in complex disease
• Epigenetics in complex disease and aging: significance and experimental approaches
• Summary
• Questions
• Further Reading
• 9 Genetic Approaches to Treating Disease
• 9.1 An Overview of Treating Genetic Disease and of Genetic Treatment of Disease
• Three different broad approaches to treating genetic disorders
• Very different treatment options for different inborn errors of metabolism
• Genetic treatment of disease may be conducted at many different levels
• 9.2 Genetic Inputs into Treating Disease with Small Molecule Drugs and Therapeutic Proteins
• An overview of how genetic differences affect the metabolism and performance of small molecule drugs
• Phenotype differences arising from genetic variation in drug metabolism
• Genetic variation in enzymes that work in phase II drug metabolism
• Altered drug responses resulting from genetic variation in drug targets
• When genotypes at multiple loci in patients are important in drug treatment: the example of warfarin
• Translating genetic advances: from identifying novel disease genes to therapeutic small molecule drugs
• Translating genomic advances and developing generic drugs as a way of overcoming the problem of too few drug targets
• Developing biological drugs: therapeutic proteins produced by genetic engineering
• Genetically engineered therapeutic antibodies with improved therapeutic potential
• 9.3 Principles of Gene and Cell Therapy
• Two broad strategies in somatic gene therapy
• The delivery problem: designing optimal and safe strategies for getting genetic constructs into the cells of patients
• Different ways of delivering therapeutic genetic constructs, and the advantages of ex vivo gene therapy
• Viral delivery of therapeutic gene constructs: relatively high efficiency but safety concerns
• Virus vectors used in gene therapy
• The importance of disease models for testing potential therapies in humans
• 9.4 Gene Therapy for Inherited Disorders: Practice and Future Directions
• Multiple successes for ex vivo gene supplementation therapy targeted at hematopoietic stem cells
• In vivo gene therapy: approaches, barriers, and recent successes
• An overview of RNA and oligonucleotide therapeutics
• RNA interference therapy
• Future therapeutic prospects using CRISPR-Cas gene editing
• Therapeutic applications of stem cells and cell reprogramming
• Obstacles to overcome in cell therapy
• A special case: preventing transmission of severe mitochondrial DNA disorders by mitochondrial replacement
• Summary
• Questions
• Further Reading
• 10 Cancer Genetics and Genomics
• 10.1 Fundamental Characteristics and Evolution of Cancer
• The defining features of unregulated cell growth and cancer
• Why cancers are different from other diseases: the contest between natural selection operating at the level of the cell and the level of the organism
• Cancer cells acquire several distinguishing biological characteristics during their evolution
• The initiation and multistage nature of cancer evolution and why most human cancers develop over many decades
• Intratumor heterogeneity arises through cell infiltration, clonal evolution, and differentiation of cancer stem cells
• 10.2 Oncogenes and Tumor Suppressor Genes
• Two fundamental classes of cancer gene
• Viral oncogenes and the natural roles of cellular oncogenes
• How normal cellular proto-oncogenes are activated to become cancer genes
• Tumor suppressor genes: normal functions, the two-hit paradigm, and loss of heterozygosity in linked markers
• The key roles of gatekeeper tumor suppressor genes in suppressing G1-S transition in the cell cycle
• The additional role of p53 in activating different apoptosis pathways to ensure that rogue cells are destroyed
• Tumor suppressor involvement in rare familial cancers and non-classical tumor suppressors
• The significance of miRNAs and long noncoding RNAs in cancer
• 10.3 Genomic Instability and Epigenetic Dysregulation in Cancer
• Different types of chromosomal instability in cancer
• Deficiency in mismatch repair results in unrepaired replication errors and global DNA instability
• Different classes of cancer susceptibility gene according to epigenetic function, epigenetic dysregulation, and epigenome–genome interaction
• 10.4 New Insights from Genome-Wide Studies of Cancers
• Genome sequencing has revealed extraordinary mutational diversity in tumors and insights into cancer evolution
• Defining the landscape of driver mutations in cancer and establishing a complete inventory of cancer-susceptibility genes
• Tracing the mutational history of cancers: just one of the diverse applications of single-cell genomics and transcriptomics in cancer
• Genome-wide RNA sequencing enables insights into the link between cancer genomes and cancer biology and aids tumor classification
• 10.5 Genetic Inroads into Cancer Therapy
• Targeted anticancer therapies are directed against key cancer cell proteins involved in oncogenesis or in escaping immunosurveillance
• CAR-T Cell therapy and the use of genetically engineered T cells to treat cancer
• The molecular basis of tumor recurrence and the evolution of drug resistance in cancers
• The promise of combinatorial drug therapies
• Summary
• Questions
• Further Reading
• 11 Genetic and Genomic Testing in Healthcare: Practical and Ethical Aspects
• 11.1 An Overview of Genetic Testing
• The different source materials and different levels of genetic testing
• 11.2 Genetic Testing for Chromosome Abnormalities and Pathogenic Structural Variation
• Screening for aneuploidies using quantitative fluorescence PCR
• Detecting large-scale copy number variants using chromosome SNP microarray analysis
• Detecting and scanning for oncogenic fusion genes using, respectively, chromosome FISH and targeted RNA sequencing
• Detecting pathogenic moderate- to small-scale deletions and duplications at defined loci is often achieved using the MLPA or ddPCR methods
• Two very different routes towards universal genome-wide screens for structural variation: genome-wide sequencing and optical genome mapping
• 11.3 Genetic and Genomic Testing for Pathogenic Point Mutations and DNA Methylation Testing
• Diverse methods permit rapid genotyping of specific point mutations
• The advantages of multiplex genotyping
• Mutation scanning: from genes and gene panels to whole exome and whole genome sequencing
• Interpreting and validating sequence variants can be aided by extensive online resources
• Detecting aberrant DNA methylation profiles associated with disease
• 11.4 Genetic and Genomic Testing: Organization of Services and Practical Applications
• The developing transformation of genetic services into mainstream genomic medicine
• An overview of diagnostic and pre-symptomatic or predictive genetic testing
• The different ways in which diagnosis of genetic conditions is carried out in the prenatal period
• Preimplantation genetic testing is carried out to prevent the transmission of a harmful genetic defect using in vitro fertilization
• Noninvasive prenatal testing (NIPT) and whole genome testing of the fetus
• An overview of the different types of genetic screening
• Pregnancy screening for fetal abnormalities
• Newborn screening allows the possibility of early medical intervention
• Different types of carrier screening can be carried out for autosomal recessive conditions
• New genomic technologies are being exploited in cancer diagnostics
• Bypassing healthcare services: the rise of direct-to-consumer (DTC) genetic testing
• The downsides of improved sensitivity through whole genome sequencing: increased uncertainty about what variants mean
• 11.5 Ethical, Legal, and Societal Issues (ELSI) in Genetic Testing
• Genetic information as family information
• Consent issues in genetic testing
• The generation of genetic data is outstripping the ability to provide clinical interpretation
• New disease gene discovery and changing concepts of diagnosis
• Complications in diagnosing mitochondrial disease
• Complications arising from incidental, additional, secondary, or unexpected information
• Consent issues in testing children
• Ethical and societal issues in prenatal diagnosis and testing
• Ethical and social issues in some emerging treatments for genetic disorders
• The ethics of germline gene modification for gene therapy and genetic enhancement
• Summary
• Questions
• Further Reading
• Glossary
• Index