Analysis of Genes and Genomes

by Richard J. Reece (University of Manchester, UK)

• ISBN13: 9780470843796
• ISBN10: 0470843799
• Format: Hardcover
• Copyright: 2004-01-01
• Publisher: WILEY

Summary

This beautifully illustrated textbook provides a clear guide to the tools and techniques of genetic engineering, gene cloning and molecular biology. All aspects of genetic engineering in the post-genomic era are covered, beginning with the basics of DNA structure and DNA metabolism. Using an example-driven approach, the fundamentals of creating mutations in DNA, cloning in bacteria, yeast, plants and animals are all clearly presented.Strong emphasis is placed on the latest, post genomic technologies including DNA macro and microarrays, genome-wide two hybrid analysis, proteomics and bioinformatics. A modern post-genome era introduction to key techniques used in genetic engineering. An example driven past-to-present approach to allow the experiments of today to be placed in an historical context The book is beautifully illustrated in full-colour throughout. Associated website including updates, additional content and illusions

Table of Contents

Preface.

Acknowledgements.

Dedication.

1. DNA: structure and function.

1.1 Nucleic acid is the material of heredity.

1.2 Structure of nucleic acids.

1.3 The double helix.

1.4 Reversible denaturing of DNA.

1.5 Structure of DNA in the cell.

1.6 The eukaryotic nucleosome.

1.7 The replication of DNA.

1.8 DNA polymerases.

1.9 The replication process.

1.10 Recombination.

1.11 Genes and genomes.

1.12 Genes within a genome.

1.13 Transcription.

1.14 RNA processing.

1.15 Translation.

2. Basic techniques in gene analysis.

2.1 Restriction enzymes.

2.2 Joining DNA molecules.

2.3 The basics of cloning.

2.4 Bacterial transformation.

2.5 Gel electrophoresis.

2.6 Nucleic acid blotting.

2.7 DNA purification.

3. Vectors.

3.1 Plasmids.

3.2 Selectable markers.

3.3 l Vectors.

3.4 Cosmid vectors.

3.5 M1 3 vectors.

3.6 Phagemids.

3.7 Artificial chromosomes.

4. Polymerase chain reaction.

4.1 PCR reaction conditions.

4.2 Thermostable DNA polymerases.

4.3 Template DNA.

4.4 Oligonucleotide primers.

4.5 Primer mismatches.

4.6 PCR in the diagnosis of genetic disease.

4.7 Cloning PCR products.

4.8 RT-PCR.

4.9 Real-time PCR.

4.10 Applications of PCR.

5. Cloning a gene.

5.1 Genomic libraries.

5.2 cDNA libraries.

5.3 Directional cDNA cloning.

5.4 PCR-based libraries.

5.5 Subtraction libraries.

5.6 Library construction in the post-genome era.

6. Gene identification.

6.1 Screening by nucleic acid hybridization.

6.2 Immunoscreening.

6.3 Screening by function.

6.4 Screening by interaction.

6.5 Phage display.

6.6 Two-hybrid screening.

6.7 Other interaction screens – variations on a theme.

7. Creating mutations.

7.1 Creating specific mutations.

7.2 Primer extension mutagenesis.

7.3 Strand selection methods.

7.4 Cassette mutagenesis.

7.5 PCR-based mutagenesis.

7.6 QuikChange(r) mutagenesis.

7.7 Creating random mutations in specific genes.

7.8 Protein engineering.

8. Protein production and purification.

8.1 Expression in E. coli.

8.2 Expression in yeast.

8.3 Expression in insect cells.

8.4 Expression in higher eukaryotic cells.

8.5 Protein purification.

9. Genome sequencing projects.

9.1 Genomic mapping.

9.2 Genetic mapping.

9.3 Physical mapping.

9.4 Nucleotide sequencing.

9.5 Genome sequencing.

9.6 The Human Genome Project.

9.7 Finding genes.

9.8 Gene assignment.

9.9 Bioinformatics.

10. Post-genome analysis.

10.1 Global changes in gene expression.

10.2 Protein function on a genome-wide scale.

10.3 Knock-out analysis.

10.4 Antisense and RNA interference (RNAi).

10.5 Genome-wide two-hybrid screens.

10.6 Protein-detection arrays.

10.7 Structural genomics.

11. Engineering plants.

11.1 Cloning in plants.

11.2 Commercial exploitation of plant transgenics.

11.3 Ethics of genetically engineered crops.

12. Engineering animal cells.

12.1 Cell culture.

12.2 Transfection of animal cells.

12.3 Viruses as vectors.

12.4 Selectable markers and gene amplification in animal cells.

12.5 Expressing genes in animal cells.

13. Engineering animals.

13.1 Pronuclear injection.

13.2 Embryonic stem cells.

13.3 Nuclear transfer.

13.4 Gene therapy.

13.5 Examples and potential of gene therapy.

Glossary.

Appendices.

Nobel prize winners.

References.