In Vitro Mutagenesis Protocols: Third Edition

In the post-genomic era, in vitro mutagenesis has emerged as a critically important tool for establishing the functions of components of the proteome. The third edition of In Vitro Mutagenesis Protocols represents a practical toolbox containing protocols vital to advancing our understanding of the connection between nucleotide sequence and sequence function. Fully updated from the previous editions, this volume contains a variety of specialty tools successfully employed to unravel the intricacies of protein-protein interaction, protein structure-function, protein regulation of biological processes, and protein activity, as well as a novel section on mutagenesis methods for unique microbes as a guide to the generalization of mutagenesis strategies for a host of microbial systems. Written in the highly successful Methods in Molecular Biology™ series format, chapters include brief introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and expert tips on troubleshooting and avoiding known pitfalls. Authoritative and up-to-date, In Vitro Mutagenesis Protocols, Third Edition offers today's researchers a valuable compendium of reliable and powerful techniques with which to illuminate the proteome and its rich web of biological implications.

Table of Contents:
Mutagenesis in Various Microbial Backgrounds.- Mutagenesis Protocols in Saccharomyces cerevisiae by In Vivo Overlap Extension.- In Vitro Mutagenesis of Brucella Species.- Random Mutagenesis Strategies for Campylobacter and Helicobacter Species.- Mutagenesis of the Repeat Regions of Herpesviruses Cloned as Bacterial Artificial Chromosomes.- An Efficient Protocol for VZV BAC-Based Mutagenesis.- A Method for Rapid Genetic Integration into Plasmodium falciparum Utilizing Mycobacteriophage Bxb1 Integrase.- PCR Mutagenesis.- Random Mutagenesis by Error-Prone PCR.- A Rapid and Versatile PCR-Based Site-Directed Mutagenesis Protocol for Generation of Mutations Along the Entire Length of a Cloned cDNA.- Rapid Sequence Scanning Mutagenesis Using In Silico Oligo Design and the Megaprimer PCR of Whole Plasmid Method (MegaWHOP).- Insertion and Deletion Mutagenesis by Overlap Extension PCR.- Targeted Amplification of Mutant Strands for Efficient Site-Directed Mutagenesis and Mutant Screening.- A Modified Inverse PCR Procedure for Insertion, Deletion, or Replacement of a DNA Fragment in a Target Sequence and Its Application in the Ligand Interaction Scan Method for Generation of Ligand-Regulated Proteins.- Amplification of Orthologous Genes Using Degenerate Primers.- Reviews.- Computational Evaluation of Protein Stability Change upon Mutations.- Approaches for Using Animal Models to Identify Loci That Genetically Interact with Human Disease-Causing Point Mutations.- Protein Evolution Mutagenesis.- Using Peptide Loop Insertion Mutagenesis for the Evolution of Proteins.- Massive Mutagenesis®: High-Throughput Combinatorial Site-Directed Mutagenesis.- Directed In Vitro Evolution of Reporter Genes Based on Semi-Rational Design and High-Throughput Screening.- Ribosome Display for RapidProtein Evolution by Consecutive Rounds of Mutation and Selection.- Protein Structure and Function Mutagenesis.- Fine-Tuning Enzyme Activity Through Saturation Mutagenesis.- Characterization of Structural Determinants of Type 1 Corticotropin Releasing Hormone (CRH) Receptor Signalling Properties.- Site-Directed Mutagenesis for Improving Biophysical Properties of VH Domains.- Phenotype Based Functional Gene Screening Using Retrovirus-Mediated Gene Trapping in Quasi-Haploid RAW 264.7 Cells.- Site-Directed Disulfide Cross-Linking to Probe Conformational Changes of a Transporter During Its Functional Cycle: Escherichia coli AcrB Multidrug Exporter as an Example.- Site-Specific Incorporation of Extra Components into RNA by Transcription Using Unnatural Base Pair Systems.- Random Mutagenesis.- Mutagen™: A Random Mutagenesis Method Providing a Complementary Diversity Generated by Human Error-Prone DNA Polymerases.- Random-Scanning Mutagenesis.- Easy Two-Step Method for Randomizing and Cloning Gene Fragments.- Mutator Bacterial Strain Mutagenesis.- Random Mutagenesis Using a Mutator Strain.- En Passant Mutagenesis: A Two Step Markerless Red Recombination System.