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Active researchers in the field describe advanced mutagenesis techniques, focusing on improvements to conventional site-directed mutagenesis. Methods are provided for chemical site-directed mutagenesis, PCR-based mutagenesis, and mutagenesis based on gene disruption that is both in vitro- and in situ-based. Methods are described for in vitro gene evolution, for gene disruption based on transposon, recombination, and cassette mutagenesis, and for facilitating the introduction of multiple mutations. The editor works in the private sector.Book News, Inc.

Book Info

Features advanced mutagenesis protocols to establish the function of proteome components; methods for accelerated protein evolution using in vitro evolution, gene shuffling, and random mutagenesis; reproducible methods for site-directed and site-specific mutagenesis, and more. Previous edition: c1996

Dedication
Preface
Contributors
1 Rapid and Reliable Site-Directed Mutagenesis Using Kunkel's Approach 1
2 Site-Directed Mutagenesis Using Altered [beta]-Lactamase Specificity 7
3 Site-Directed Mutagenesis Facilitated by Dpnl Selection on Hemimethylated DNA 19
4 Multiple Site-Directed Mutagenesis In Vitro 29
5 Two-Stage Polymerase Chain Reaction Protocol Allowing Introduction of Multiple Mutations, Deletions, and Insertions, Using QuikChange Site-Directed Mutagenesis 37
6 Efficient and Accurate Site-Directed Mutagenesis of Large Plasmids 45
7 Combining Site-Specific Chemical Modification with Site-Directed Mutagenesis: Versatile Strategy to Move Beyond Structural Limitations of 20 Natural Amino Acids Side Chains in Protein Engineering 55
8 Site-Directed Mutagenesis Mediated by a Single Polymerase Chain Reaction Product 67
9 Megaprimer Method for Polymerase Chain Reaction-Mediated Generation of Specific Mutations in DNA 71
10 Generation of Epitope-Tagged Proteins by Inverse Polymerase Chain Reaction Mutagenesis 77
11 Site-Directed Mutagenesis by Polymerase Chain Reaction 85
12 Generation of Multiple Site-Specific Mutations by Polymerase Chain Reaction 95
13 Phenotypic Expression of Polymerase Chain Reaction-Generated Random Mutations in a Foreign Gene After its Introduction into an Acinetobacter Chromosome by Natural Transformation 103
14 Polymerase Chain Reaction-Mediated Mutagenesis in Sequences Resistant to Homogeneous Amplification 117
15 Polymerase Chain Reaction-Based Signature-Tagged Mutagenesis 127
16 High-Throughput Scanning Mutagenesis by Recombination Polymerase Chain Reaction 139
17 In Vitro Scanning-Saturation Mutagenesis 149
18 Random Transposon Mutagenesis of Large DNA Molecules in Escherichia Coli 165
19 Random Chromosomal Gene Disruption Using Cassette Mutagenesis 173
20 Transplacement Mutagenesis: A Recombination-Based In Situ Mutagenesis Protocol 189
21 Preparation of Transposon Insertion Lines and Determination of Insertion Sites in Arabidopsis Genome 209
22 Evolutionary Molecular Engineering by Random Elongation Mutagenesis 221
23 Random Mutagenesis for Protein Breeding 231
24 DNA Shuffling and Family Shuffling for In Vitro Gene Evolution 243
25 Mutagenic Polymerase Chain Reaction of Protein-Coding Genes for In Vitro Evolution 259
Index 269