Directed evolution library creation : methods and protocols / edited by Frances H. Arnold and George Georgiou.

Contributor(s): Arnold, Frances Hamilton | Georgiou, George
Material type: TextTextSeries: Methods in molecular biology (Clifton, N.J.): v. 231.Publisher: Totowa, N.J. : Humana Press, ©2003Description: 1 online resource (x, 224 pages) : illustrationsContent type: text Media type: computer Carrier type: online resourceISBN: 9781592593958; 1588292851; 9781588292858; 159259395XSubject(s): Gene libraries -- Laboratory manuals | Genetic engineering -- Laboratory manuals | Molecular evolution -- Laboratory manuals | Microbial mutation -- Laboratory manuals | Combinatorial chemistry -- Laboratory manuals | Genetic recombination -- Laboratory manuals | Gene Library | Combinatorial Chemistry Techniques | Banques de gènes -- Manuels de laboratoire | Génie génétique -- Manuels de laboratoire | Évolution moléculaire -- Manuels de laboratoire | Micro-organismes -- Mutation -- Manuels de laboratoire | Chimie combinatoire -- Manuels de laboratoire | Recombinaison génétique -- Manuels de laboratoire | SCIENCE -- Life Sciences -- Molecular Biology | Combinatorial chemistry | Gene libraries | Genetic engineering | Genetic recombination | Microbial mutation | Molecular evolution | Genetica | Moleculaire biologie | Genetische manipulatie | Wetenschappelijke technieken | Engenharia genética | Biologia molecular | Mutagênese | Recombinação genéticaGenre/Form: Fulltext. | Illustration. | Internet Resource. | Laboratory Manual. | Laboratory manuals. Additional physical formats: Print version:: Directed evolution library creation.DDC classification: 572.8 LOC classification: QH442.4 | .D57 2003Online resources: Click here to access online
Contents:
Cover -- Preface -- Contents -- Contributors -- MUTAGENESIS AND RECOMBINATION METHODS -- 1 Generating Mutant Libraries Using Error-Prone PCR -- 2 Preparing Libraries in Escherichia coli -- 3 Preparing Libraries in Saccharomyces cerevisiae -- 4 Creating Random Mutagenesis Libraries by Megaprimer PCR of Whole Plasmid (MEGAWHOP) -- 5 Construction of Designed Protein Libraries Using Gene Assembly Mutagenesis -- 6 Production of Randomly Mutated Plasmid Libraries Using Mutator Strains -- 7 Evolution of Microorganisms Using Mutator Plasmids -- 8 Random Insertion and Deletion Mutagenesis -- 9 Random Oligonucleotide Mutagenesis -- 10 Saturation Mutagenesis -- 11 DNA Shuffling -- 12 Family Shuffling with Single-Stranded DNA -- 13 In Vitro DNA Recombination by Random Priming -- 14 Staggered Extension Process (StEP) In Vitro Recombination -- 15 RACHITT -- 16 The Creation of ITCHY Hybrid Protein Libraries -- 17 Preparation of SCRATCHY Hybrid Protein Libraries -- 18 Sequence Homology-Independent Protein Recombination (SHIPREC) -- 19 Producing Chimeric Genes by CLERY -- ANALYSIS OF LIBRARY DIVERSITY -- 20 Analysis of Shuffled Libraries by Oligonucleotide Probe Hybridization -- 21 Sequence Mapping of Combinatorial Libraries on Macro- or Microarrays -- 22 Sequence Mapping of Combinatorial Libraries on Macro- and Microarrays -- Index -- Last Page.
Summary: Directed protein evolution-a powerful technique for the discovery of new enzymes and therapeutic proteins-has matured to the point that it is now the centerpiece of enzyme engineering, where it has catalyzed the development of numerous experimental methods. In Directed Evolution Library Creation: Methods and Protocols, seasoned practitioners from many leading laboratories share their best experimental protocols for the generation of molecular diversity. Described in step-by-step detail to ensure experimental success, these protocols include readily reproducible methods for random mutagenesis of entire genes or segments of genes, for homologous and nonhomologous recombination, and for constructing in vivo libraries in bacteria and yeast. In addition to the various protocols for creating libraries, this volume also describes ways to analyze libraries, particularly those made by recombination. An accompanying volume, Directed Enzyme Evolution: Screening and Selection Methods (ISBN: 1-58829-286-X), is devoted entirely to selection and screening methods that can be applied to the directed evolution of enzymes. Taken together, Directed Evolution Library Creation: Methods and Protocols and Directed Enzyme Evolution: Screening and Selection Methods capture for newcomers and more experienced investigators alike all the key methods for using directed protein evolution to better understand protein structure-function relationships, to discover new enzymes and therapeutic proteins, and to design new assays suitable for specific applications.
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Includes bibliographical references and index.

Cover -- Preface -- Contents -- Contributors -- MUTAGENESIS AND RECOMBINATION METHODS -- 1 Generating Mutant Libraries Using Error-Prone PCR -- 2 Preparing Libraries in Escherichia coli -- 3 Preparing Libraries in Saccharomyces cerevisiae -- 4 Creating Random Mutagenesis Libraries by Megaprimer PCR of Whole Plasmid (MEGAWHOP) -- 5 Construction of Designed Protein Libraries Using Gene Assembly Mutagenesis -- 6 Production of Randomly Mutated Plasmid Libraries Using Mutator Strains -- 7 Evolution of Microorganisms Using Mutator Plasmids -- 8 Random Insertion and Deletion Mutagenesis -- 9 Random Oligonucleotide Mutagenesis -- 10 Saturation Mutagenesis -- 11 DNA Shuffling -- 12 Family Shuffling with Single-Stranded DNA -- 13 In Vitro DNA Recombination by Random Priming -- 14 Staggered Extension Process (StEP) In Vitro Recombination -- 15 RACHITT -- 16 The Creation of ITCHY Hybrid Protein Libraries -- 17 Preparation of SCRATCHY Hybrid Protein Libraries -- 18 Sequence Homology-Independent Protein Recombination (SHIPREC) -- 19 Producing Chimeric Genes by CLERY -- ANALYSIS OF LIBRARY DIVERSITY -- 20 Analysis of Shuffled Libraries by Oligonucleotide Probe Hybridization -- 21 Sequence Mapping of Combinatorial Libraries on Macro- or Microarrays -- 22 Sequence Mapping of Combinatorial Libraries on Macro- and Microarrays -- Index -- Last Page.

Print version record.

Directed protein evolution-a powerful technique for the discovery of new enzymes and therapeutic proteins-has matured to the point that it is now the centerpiece of enzyme engineering, where it has catalyzed the development of numerous experimental methods. In Directed Evolution Library Creation: Methods and Protocols, seasoned practitioners from many leading laboratories share their best experimental protocols for the generation of molecular diversity. Described in step-by-step detail to ensure experimental success, these protocols include readily reproducible methods for random mutagenesis of entire genes or segments of genes, for homologous and nonhomologous recombination, and for constructing in vivo libraries in bacteria and yeast. In addition to the various protocols for creating libraries, this volume also describes ways to analyze libraries, particularly those made by recombination. An accompanying volume, Directed Enzyme Evolution: Screening and Selection Methods (ISBN: 1-58829-286-X), is devoted entirely to selection and screening methods that can be applied to the directed evolution of enzymes. Taken together, Directed Evolution Library Creation: Methods and Protocols and Directed Enzyme Evolution: Screening and Selection Methods capture for newcomers and more experienced investigators alike all the key methods for using directed protein evolution to better understand protein structure-function relationships, to discover new enzymes and therapeutic proteins, and to design new assays suitable for specific applications.

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