Table of Contents

The Function and Organization of Plasmids Finbarr Hayes Choosing a Cloning Vector Andrew Preston Escherichia coli Host Strains Nicola Casali Chemical Transformation of E. coli W. Edward Swords Electroporation of E. coli Claire A. Woodall DNA Transfer by Bacterial Conjugation Claire A. Woodall Cosmid Packaging and Infection of E. coli Mallory J. A. White and Wade A. Nichols Isolation of Plasmids from E. coli by Alkaline Lysis Sabine Ehrt and Dirk Schnappinger Isolation of Plasmids from E. coli by Boiling Lysis Sabine Ehrt and Dirk Schnappinger High-Purity Plasmid Isolation Using Silica Oxide Stefan Grimm and Frank Voß-Neudecker High-Throughput Plasmid Extraction Using Microtiter Plates Michael A. Quail Isolation of Cosmid and BAC DNA from E. coli Daniel Sinnett and Alexandre Montpetit Preparation of Single-Stranded DNA from Phagemid Vectors W. Edward Swords Using Desktop Cloning Software to Plan, Track, and Evaluate Cloning Projects Robert H. Gross Cloning in Plasmid Vectors Carey Pashley and Sharon Kendall Extraction of DNA from Agarose Gels Nicholas Downey Cloning PCR Products with T-Vectors Wade A. Nichols Construction of Genomic Libraries in l-Vectors Yilun Wang, Zheng Cao, Darryl Hood, and James G. Townsel Rapid Screening of Recombinant Plasmids Sangwei Lu Restriction Analysis of Recombinant Plasmids Joanne Goranson-Siekierke and Jarrod L. Erbe Screening Recombinant DNA Libraries Wade A. Nichols Sequencing Using Fluorescent-Labeled Nucleotides Allison F. Gillaspy Site-Directed Mutagenesis Using the Megaprimer Method Zhidong Xu, Alessia Colosimo, and Dieter C. Gruenert Site-Directed Mutagenesis byInverse PCR Clifford N. Dominy and David W. Andrews Creating Nested DNA Deletions Using Exonuclease III Rosamund Powles and Lafras M. Steyn Transposon and Transposome Mutagenesis of Plasmids, Cosmids, and BACs Alistair McGregor In Vitro Transcription and Translation Farahnaz Movahedzadeh, Susana González Rico, and Robert A. Cox Vectors for the Expression of Recombinant Proteins in E. coli Sally A. Cantrell Expression of Recombinant Proteins from lac Promoters Charles R. Sweet Plasmid-Based Reporter Genes: Assays for b-Galactosidase and Alkaline Phosphatase Activities Minghsun Liu Plasmid-Based Reporter Genes: Assays for Green Fluorescent Protein Sergei R. Doulatov Index