An Introduction to Genetic Engineering

by Desmond S. T. Nicholl

• ISBN13: 9780521004718
• ISBN10: 0521004713
• Edition: 2nd
• Format: Paperback
• Copyright: 2002-02-11
• Publisher: Cambridge University Press
• More Book Details
• Purchase Benefits

Summary

Des Nicholl presents here a new, fully revised, and expanded edition of his popular undergraduate-level textbook. Many of the features of the original edition have been retained; the book still offers a concise technical introduction to the subject of genetic engineering. However, the book is now divided into three main sections: the first introduces students to basic molecular biology, the second section explains the methods used to manipulate genes, and the third deals with modern applications of genetic engineering. A whole chapter is now devoted to the polymerase chain reaction. Applications covered in the book include genomics, protein engineering, gene therapy, cloning, and transgenic animals and plants. A final chapter discusses the ethical questions surrounding genetic engineering in general. An Introduction to Genetic Engineering is essential reading for undergraduate students of biotechnology, genetics, molecular biology and biochemistry.

Author Biography

Des Nicholl is a Senior Lecturer in Biological Sciences, The University of Paisley, Scotland, UK

Table of Contents

Preface to the second edition	xi
Introduction	1	(8)
What is genetic engineering?	1	(2)
Laying the foundations	3	(1)
First steps	4	(2)
What is in store?	6	(3)
Part I: The basis of genetic engineering	9	(46)
Introducting molecular biology	11	(16)
The flow of genetic information	11	(2)
The structure of DNA and RNA	13	(3)
Gene organisation	16	(5)
Gene structure in prokaryotes	18	(1)
Gene structure in eukaryotes	19	(2)
Gene expression	21	(2)
Genes and genomes	23	(4)
Genome size and complexity	23	(1)
Genome organisation	24	(3)
Working with nucleic acids	27	(16)
Isolation of DNA and RNA	27	(2)
Handling and quantification of nucleic acids	29	(1)
Radiolabelling of nucleic acids	30	(3)
End labelling	30	(1)
Nick translation	31	(1)
Labelling by primer extension	31	(2)
Nucleic acid hybridisation	33	(1)
Gel electrophoresis	33	(2)
DNA sequencing	35	(8)
Maxam-Gilbert (chemical) sequencing	37	(1)
Sanger-Coulson (dideoxy or enzymatic) sequencing	37	(3)
Electrophoresis and reading of sequences	40	(3)
The tools of the trade	43	(12)
Restriction enzymes - cutting DNA	43	(5)
Type II restriction endonucleases	44	(1)
Use of restriction endonucleases	45	(2)
Restriction mapping	47	(1)
DNA modifying enzymes	48	(4)
Nucleases	48	(1)
Polymerases	49	(2)
Enzymes that modify the ends of DNA molecules	51	(1)
DNA ligase - joining DNA molecules	52	(3)
Part II: The methodology of gene manipulation	55	(96)
Host cells and vectors	57	(30)
Host cell types	58	(2)
Prokaryotic hosts	58	(1)
Eukaryotic hosts	59	(1)
Plasmid vectors for use in E. coli	60	(6)
What are plasmids?	61	(1)
Basic cloning plasmids	61	(2)
Slightly more exotic plasmid vectors	63	(3)
Bacteriophage vectors for use in E. coli	66	(9)
What are bacteriophages?	66	(4)
Vectors based on bacteriophage λ	70	(4)
Vectors based on bacteriophage M13	74	(1)
Other vectors	75	(5)
Hybrid plasmid/phage vectors	76	(1)
Vectors for use in eukaryotic cells	77	(2)
Artificial chromosomes	79	(1)
Getting DNA into cells	80	(7)
Transformation and transfection	80	(1)
Packaging phage DNA in vitro	81	(2)
Alternative DNA delivery methods	83	(4)
Cloning strategies	87	(28)
Which approach is best?	87	(2)
Cloning from mRNA	89	(9)
Synthesis of cDNA	90	(3)
Cloning cDNA in plasmid vectors	93	(3)
Cloning cDNA in bacteriophage vectors	96	(2)
Cloning from genomic DNA	98	(8)
Genomic libraries	99	(2)
Preparation of DNA fragments for cloning	101	(2)
Ligation, packaging and amplification of libraries	103	(3)
Advanced cloning strategies	106	(9)
Synthesis and cloning of cDNA	106	(3)
Expression of cloned cDNA molecules	109	(2)
Cloning large DNA fragments in BAC and YAC vectors	111	(4)
The polymerase chain reaction	115	(17)
The (short) history of the PCR	115	(3)
The methodology of the PCR	118	(5)
The essential features of the PCR	118	(3)
The design of primers for PCR	121	(1)
DNA polymerases for PCR	121	(2)
More exotic PCR techniques	123	(6)
PCR using mRNA templates	123	(1)
Nested PCR	124	(2)
Inverse PCR	126	(1)
RAPD and several other acronyms	127	(2)
Processing of PCR products	129	(1)
Applications of the PCR	130	(2)
Selection, screening and analysis of recombinants	132	(19)
Genetic selection and screening methods	133	(5)
The use of chromogenic substrates	133	(2)
Insertional inactivation	135	(1)
Complementation of defined mutations	136	(1)
Other genetic selection methods	137	(1)
Screening using nucleic acid hybridisation	138	(3)
Nucleic acid probes	138	(1)
Screening clone banks	139	(2)
Immunological screening for expressed genes	141	(2)
Analysis of cloned genes	143	(8)
Characterisation based on mRNA translation in vitro	143	(2)
Restriction mapping	145	(1)
Blotting techniques	145	(3)
DNA sequencing	148	(3)
Part III: Genetic engineering in action	151	(112)
Understanding genes and genomes	153	(25)
Analysis of gene structure and function	153	(6)
A closer look at sequences	154	(1)
Finding important regions of genes	155	(2)
Investigating gene expression	157	(2)
From genes to genomes	159	(6)
Analysing genomes	160	(1)
Mapping genomes	161	(4)
Genome sequencing	165	(2)
Sequencing technology	165	(1)
Genome projects	165	(2)
The human genome project	167	(11)
Whose genome, and how many genes does it contain?	169	(1)
Genetic and physical maps of the human genome	170	(4)
Deriving and assembling the sequence	174	(1)
What next?	175	(3)
Genetic engineering and biotechnology	178	(19)
Making proteins	179	(4)
Native and fusion proteins	179	(2)
Yeast expression systems	181	(1)
The baculovirus expression system	182	(1)
Mammalian cell lines	183	(1)
Protein engineering	183	(2)
Examples of biotechnological applications of rDNA technology	185	(12)
Production of enzymes	185	(2)
The BST story	187	(3)
Therapeutic products for use in human health-care	190	(7)
Medical and forensic applications of gene manipulation	197	(27)
Diagnosis and characterisation of medical conditions	197	(13)
Diagnosis of infection	198	(1)
Patterns of inheritance	198	(3)
Genetically based disease conditions	201	(9)
Treatment using rDNA technology - gene therapy	210	(5)
Getting transgenes into patients	211	(3)
Gene therapy for adenosine deaminase deficiency	214	(1)
Gene therapy for cystic fibrosis	214	(1)
DNA profiling	215	(9)
The history of `genetic fingerprinting'	216	(2)
DNA profiling and the law	218	(1)
Mysteries of the past revealed by genetic detectives	219	(5)
Transgenic plants and animals	224	(23)
Transgenic plants	224	(13)
Why transgenic plants?	225	(1)
Tiplasmids as vectors for plant cells	226	(2)
Making transgenic plants	228	(2)
Putting the technology to work	230	(7)
Transgenic animals	237	(10)
Why transgenic animals?	237	(1)
Producting transgenic animals	238	(3)
Applications of transgenic animal technology	241	(6)
The other sort of cloning	247	(12)
Early thoughts and experiments	247	(3)
First steps towards cloning	249	(1)
Nuclear totipotency	250	(1)
Frogs and toads and carrots	250	(3)
A famous sheep - the breakthrough achieved	253	(3)
Beyond Dolly	256	(3)
Brave new world or genetic nightmare?	259	(4)
Is science ethically and morally neutral?	259	(1)
Elements of the ethics debate	260	(2)
Does Frankenstein's monster live inside Pandora's box?	262	(1)
Suggestings for further reading	263	(3)
Using the World Wide Web	266	(4)
Glossary	270	(17)
Index	287