From Genes to Genomes Concepts and Applications of DNA Technology

ISBN-10: 0470683856

ISBN-13: 9780470683859

Edition: 3rd 2012

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The latest edition of this highly successful textbook introduces the key techniques and concepts involved in cloning genes and in studying their expression and variation. The new edition features: Increased coverage of whole-genome sequencing technologies and enhanced treatment of bioinformatics. Clear, two-colour diagrams throughout. A dedicated website including all figures. Noted for its outstanding balance between clarity of coverage and level of detail, this book provides an excellent introduction to the fast moving world of molecular genetics.
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Book details

Edition: 3rd
Copyright year: 2012
Publisher: John Wiley & Sons, Limited
Publication date: 12/9/2011
Binding: Paperback
Pages: 400
Size: 6.50" wide x 9.25" long x 0.75" tall
Weight: 1.408
Language: English

From Genes to Genomes
Basic molecular biology
The DNA backbone
The base pairs
RNA structure
Nucleic acid synthesis
Coiling and supercoilin
What is a gene?
Information flow: gene expression
Gene structure and organisation
Exons and introns
Refinements of the model
How to Clone a Gene
What is cloning?
Overview of the procedures
Extraction and purification of nucleic acids
Breaking up cells and tissues
Alkaline denaturation
Column purification
Detection and quantitation of nucleic acids
Gel electrophoresis
Analytical gel electrophoresis
Preparative gel electrophoresis
Restriction endonucleases
Sticky and blunt ends
Optimising ligation conditions
Preventing unwanted ligation: alkaline phosphatase and double digests
Other ways of joining DNA fragments
Modification of restriction fragment ends
Linkers and adaptors
Homopolymer tailing
Plasmid vectors
Plasmid replication
Cloning sites
Selectable markers
Insertional inactivation
Vectors based on the lambda bacteriophage
Lambda biology
In vitro packaging
Insertion vectors
Replacement vectors
Supervectors: YACs and BACs
Genomic and cDNA Libraries
Genomic libraries
Partial digests
Choice of vectors
Construction and evaluation of a genomic library
Growing and storing libraries
cDNA libraries
Isolation of mRNA
cDNA synthesis
Bacterial cDNA
Screening libraries with gene probes
Labelling probes
Steps in a hybridization experiment
Screening procedure
Probe selection and generation
Screening expression libraries with antibodies
Characterization of plasmid clones
Southern blots
PCR and sequence analysis
Polymerase Chain Reaction (PCR)
The PCR reaction
PCR in practice
Optimisation of the PCR reaction
Primer design
Analysis of PCR products
Cloning PCR products
Long-range PCR
Reverse-transcription PCR
Quantitative and real-time PCR
SYBR Green
Molecular beacons
Applications of PCR
Probes and other modified products
PCR cloning strategies
Analysis of recombinant clones and rare events
Diagnostic applications
Sequencing a Cloned Gene
DNA sequencing
Principles of DNA sequencing
Automated sequencing
Extending the sequence
Shotgun sequencing; contig assembly
Databank entries and annotation
Sequence analysis
Identification of coding region
Expression signals
Sequence comparisons
DNA sequences
Protein sequence comparisons
Sequence alignments: Clustal
Protein structure
Structure predictions
Protein motifs and domains
Confirming gene function
Allelic replacement and gene knockouts
Analysis of Gene Expression
Analysing transcription
Northern blots
Reverse transcription-PCR
In situ hybridization
Methods for studying the promoter
Locating the promoter
Reporter genes
Regulatory elements and DNA-binding proteins
Yeast one-hybrid assays
DNase I footprinting
Gel retardation assays
Chromatin immunoprecipitation (ChIP)
Translational analysis
Western blots
Immunocytochemistry and immunohistochemistry
Products from Native and Manipulated Cloned Genes
Factors affecting expression of cloned genes
Translation initiation
Codon usage
Nature of the protein product
Expression of cloned genes in bacteria
Transcriptional fusions
Stability: conditional expression
Expression of lethal genes
Translational fusions
Yeast systems
Cloning vectors for yeasts
Yeast expression systems
Expression in insect cells: baculovirus systems
Mammalian cells
Cloning vectors for mammalian cells
Expression in mammalian cells
Adding tags and signals
Tagged proteins
Secretion signals
In vitro mutagenesis
Site-directed mutagenesis
Synthetic genes
Assembly PCR
Synthetic genomes
Protein engineering
Subunit vaccines
DNA vaccines
Genomic Analysis
Overview of genome sequencing
Next generation sequencing (NGS)
Pyrosequencing (454)
SOLiD sequencing (Applied Biosystems)
Bridge amplification sequencing (Solexa/Ilumina)
Other technologies
De novo sequence assembly
Repetitive elements and gaps
Analysis and annotation
Identification of ORFs
Identification of the function of genes and their products
Other features of nucleic acid sequences
Comparing genomes
Genome browsers
Relating genes and functions: genetic and physical maps
Linkage analysis
Ordered libraries and chromosome walking
Transposon mutagenesis and other screening techniques
Transposition in bacteria
Transposition in Drosophila
Transposition in other organisms
Signature-tagged mutagenesis
Gene knockouts, gene knockdowns and gene silencing
Analysis of Genetic Variation
Single nucleotide polymorphisms
Direct sequencing
SNP arrays
Larger scale variations
Microarrays and indels
Other methods for studying variation
Genomic Southern blot analysis: restriction fragment length polymorphisms (RFLPs)
VNTR and microsatellites
Pulsed-field gel electrophoresis
Human genetic variation: relating phenotype to genotype
Linkage analysis
Genome-wide association studies (GWAS)
Database resources
Genetic diagnosis
Molecular phylogeny
Methods for constructing trees
Post-Genomic Analysis
Analysing transcription: transcriptomes
Differential screening
Other methods: transposons and reporters
Array-based methods
Expressed sequence tag (EST) arrays
PCR product arrays
Synthetic oligonucleotide arrays
Important factors in array hybridization
Transcriptome sequencing
Translational analysis: proteomics
Two-dimensional electrophoresis
Mass spectrometry
Post-translational analysis: protein interactions
Two-hybrid screening
Phage display libraries
Integrative studies: systems biology
Metabolomic analysis
Pathway analysis and systems biology
Modifying Organisms: Transgenics
Transgenesis and cloning
Common species used for transgenesis
Control of transgene expression
Animal transgenesis
Basic methods
Direct injection
Retroviral vectors
Embryonic stem cell technology
Gene knockouts
Gene knock-down technology: RNA interference
Gene knock-in technology
Applications of transgenic animals
Disease prevention and treatment
Live vaccine production: modification of bacteria and viruses
Gene therapy
Viral vectors for gene therapy
Transgenic plants and their applications
Introducing foreign genes
Gene subtraction
Transgenics: a coda 353 Glossary
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