Library Science

A comprehensive compendium of cutting-edge 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 nonhomologus 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. Copy for Both Volumes Directed Evolution Library Creation: Methods and Protocols and Directed Enzyme Evolution: Screening and Selection Methods constitute an extraordinary collection of all the key methods used today for directed evolution research. Described in step-by-step detail to ensure robust experimental results, these methods will enable both newcomers and more experienced investigators to design and implement directed evolution strategies for the engineering of novel proteins. The first volume describes methods for the creation of mutated DNA molecules, or DNA libraries, encoding variants of desired proteins. The second volume describes methods for screening DNA libraries to isolate mutant proteins that exhibit a specified function.

Ed. Arnold, Frances H.; Georgiou, George

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Table of Contents

1. Generating Mutant Libraries Using Error-Prone PCR (pp. 3-10)
   Cirino, Patrick C.; Mayer, Kimberly M.; Umeno, Daisuke
2. Preparing Libraries in Escherichia coli (pp. 11-16)
   Tobias, Alexander V.
3. Preparing Libraries in Saccharomyces cerevisiae (pp. 17-22)
   Bulter, Thomas; Alcalde, Miguel
4. Creating Random Mutagenesis Libraries by Megaprimer PCR of Whole Plasmid (MEGAWHOP) (pp. 23-28)
   Miyazaki, Kentaro
5. Construction of Designed Protein Libraries Using Gene Assembly Mutagenesis (pp. 29-38)
   Bessette, Paul H.; Mena, Marco A.; Nguyen, Annalee W.; Daugherty, Patrick S.
6. Production of Randomly Mutated Plasmid Libraries Using Mutator Strains (pp. 39-44)
   Nguyen, Annalee W.; Daugherty, Patrick S.
7. Evolution of Microorganisms Using Mutator Plasmids (pp. 45-52)
   Selifonova, Olga; Schellenberger, Volker
8. Random Insertion and Deletion Mutagenesis (pp. 53-64)
   Murakami, Hiroshi; Hohsaka, Takahiro; Sisido, Masahiko
9. Random Oligonucleotide Mutagenesis (pp. 65-74)
   Sneeden, Jessica L.; Loeb, Lawrence A.
10. Saturation Mutagenesis (pp. 75-84)
    Georgescu, Radu; Bandara, Geethani; Sun, Lianhong
11. DNA Shuffling (pp. 85-90)
    Joern, John M.
12. Family Shuffling with Single-Stranded DNA (pp. 91-98)
    Zha, Wenjuan; Zhu, Tongbo; Zhao, Huimin
13. In Vitro DNA Recombination by Random Priming (pp. 99-104)
    Esteban, Olga; Woodyer, Ryan D.; Zhao, Huimin
14. Staggered Extension Process (StEP) In Vitro Recombination (pp. 105-110)
    Aguinaldo, Anna Marie; Arnold, Frances H.
15. RACHITT Gene Family Shuffling by Random Chimeragenesis on Transient Templates (pp. 111-128)
    Coco, Wayne M.
16. The Creation of ITCHY Hybrid Protein Libraries (pp. 129-142)
    Ostermeier, Marc; Lutz, Stefan
17. Preparation of SCRATCHY Hybrid Protein Libraries Size- and In-Frame Selection of Nucleic Acid Sequences (pp. 143-152)
    Lutz, Stefan; Ostermeier, Marc
18. Sequence Homology-Independent Protein Recombination (SHIPREC) (pp. 153-164)
    Udit, Andrew K.; Silberg, Jonathan J.; Sieber, Volker
19. Producing Chimeric Genes by CLERY In Vitro and In Vivo Recombination (pp. 165-174)
    Abecassis, Valerie; Pompon, Denis; Truan, Gilles
20. Analysis of Shuffled Libraries by Oligonucleotide Probe Hybridization (pp. 177-188)
    Meinhold, Peter; Joern, John M.; Silberg, Jonathan J.
21. Sequence Mapping of Combinatorial Libraries on Macro- or Microarrays Experimental Design of DNA Arrays (pp. 189-198)
    Abecassis, Valerie; Truan, Gilles; Jaffrelo, Loic; Pompon, Denis
22. Sequence Mapping of Combinatorial Libraries on Macro- and Microarrays Bioinformatic Treatment of Data (pp. 199-211)
    Pompon, Denis; Truan, Gilles; Abecassis, Valerie

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