Bibliothèque de L'IMIST/CNRST

Includes bibliographical references (pages 375-377) and index.

From Genes to Genomes; Contents; Preface; 1 From Genes to Genomes; 1.1 Introduction; 1.2 Basic molecular biology; 1.2.1 The DNA backbone; 1.2.2 The base pairs; 1.2.3 RNA structure; 1.2.4 Nucleic acid synthesis; 1.2.5 Coiling and supercoilin; 1.3 What is a gene?; 1.4 Information flow: gene expression; 1.4.1 Transcription; 1.4.2 Translation; 1.5 Gene structure and organisation; 1.5.1 Operons; 1.5.2 Exons and introns; 1.6 Refinements of the model; 2 How to Clone a Gene; 2.1 What is cloning?; 2.2 Overview of the procedures; 2.3 Extraction and purification of nucleic acids.

2.3.1 Breaking up cells and tissues2.3.2 Alkaline denaturation; 2.3.3 Column purification; 2.4 Detection and quantitation of nucleic acids; 2.5 Gel electrophoresis; 2.5.1 Analytical gel electrophoresis; 2.5.2 Preparative gel electrophoresis; 2.6 Restriction endonucleases; 2.6.1 Specificity; 2.6.2 Sticky and blunt ends; 2.7 Ligation; 2.7.1 Optimising ligation conditions; 2.7.2 Preventing unwanted ligation: alkaline phosphatase and double digests; 2.7.3 Other ways of joining DNA fragments; 2.8 Modification of restriction fragment ends; 2.8.1 Linkers and adaptors; 2.8.2 Homopolymer tailing.

2.9 Plasmid vectors2.9.1 Plasmid replication; 2.9.2 Cloning sites; 2.9.3 Selectable markers; 2.9.4 Insertional inactivation; 2.9.5 Transformation; 2.10 Vectors based on the lambda bacteriophage; 2.10.1 Lambda biology; 2.10.2 In vitro packaging; 2.10.3 Insertion vectors; 2.10.4 Replacement vectors; 2.11 Cosmids; 2.12 Supervectors: YACs and BACs; 2.13 Summary; 3 Genomic and cDNA Libraries; 3.1 Genomic libraries; 3.1.1 Partial digests; 3.1.2 Choice of vectors; 3.1.3 Construction and evaluation of a genomic library; 3.2 Growing and storing libraries; 3.3 cDNA libraries; 3.3.1 Isolation of mRNA.

3.3.2 cDNA synthesis3.3.3 Bacterial cDNA; 3.4 Screening libraries with gene probes; 3.4.1 Hybridization; 3.4.2 Labelling probes; 3.4.3 Steps in a hybridization experiment; 3.4.4 Screening procedure; 3.4.5 Probe selection and generation; 3.5 Screening expression libraries with antibodies; 3.6 Characterization of plasmid clones; 3.6.1 Southern blots; 3.6.2 PCR and sequence analysis; 4 Polymerase Chain Reaction (PCR); 4.1 The PCR reaction; 4.2 PCR in practice; 4.2.1 Optimisation of the PCR reaction; 4.2.2 Primer design; 4.2.3 Analysis of PCR products; 4.2.4 Contamination.

4.3 Cloning PCR products4.4 Long-range PCR; 4.5 Reverse-transcription PCR; 4.6 Quantitative and real-time PCR; 4.6.1 SYBR Green; 4.6.2 TaqMan; 4.6.3 Molecular beacons; 4.7 Applications of PCR; 4.7.1 Probes and other modified products; 4.7.2 PCR cloning strategies; 4.7.3 Analysis of recombinant clones and rare events; 4.7.4 Diagnostic applications; 5 Sequencing a Cloned Gene; 5.1 DNA sequencing; 5.1.1 Principles of DNA sequencing; 5.1.2 Automated sequencing; 5.1.3 Extending the sequence; 5.1.4 Shotgun sequencing; contig assembly; 5.2 Databank entries and annotation; 5.3 Sequence analysis.

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.