Recombinant Gene Expression Protocols

1. Front Matter

   Pages i-xx

   PDF

2. Introduction

   1. Recombinant Gene Expression Protocols

      • Rocky S. Tuan

      Pages 3-8

3. Expression in E. Coli

   1. Designing Expression Plasmid Vectors in E. coli

      • Paulina Balbás

      Pages 11-16

   2. Design of Bacterial Hosts for lac-Based Expression Vectors

      • Herbert P. Schweizer, RoxAnn R. Karkhoff-Schweizer

      Pages 17-27

   3. Bacteriophage λ-Based Expression Vectors

      • Alan C. Christensen

      Pages 29-36

   4. Strategies for Gene Fusions

      • Stefan Ståhl, Per-Åke Nygren, Mathias Uhlén

      Pages 37-54

   5. Application of the E. coli trp Promoter

      • Daniel G. Yansura, Steven H. Bass

      Pages 55-62

   6. Expression and Secretion of Proteins in E. coli

      • Ophry Pines, Masayori Inouye

      Pages 73-87

   7. Synthetic Two-Cistron Expression System

      • Brigitte E. Schoner

      Pages 89-97

   8. Expression of cDNA-Encoded Proteins by Cell-Free Transcription and Translation

      • Chistopher Coleclough

      Pages 99-110

4. Expression in E. Coli

   1. Expression of Proteins in E. coli Utilizing a Dual Promoter-Based Vector: pLACT7

      • George A. Garcia, Shaorong Chong

      Pages 63-71

5. Expression in Yeast

   1. Yeast Plasmids

      • Keshav K. Singh, Jack A. Heinemann

      Pages 113-130

   2. Inducible Expression Cassettes in Yeast: GAL4

      • Lawrence M. Mylin, James E. Hopper

      Pages 131-148

   3. Inducible Expression Cassettes in Yeast: ADH2

      • Virginia L. Price

      Pages 149-157

   4. Constitutive Expression Vectors: PGK

      • Ian R. Graham, Alistair Chambers

      Pages 159-169

6. Viral Expression Systems

   1. Design and Construction of Recombinant Vaccinia Viruses

      • Christopher C. Broder, Patricia L. Earl

      Pages 173-197

   2. A Simplified Method of Screening for Isolation of Recombinant Vaccinia Virus

      • Haifeng Chen, R. Padmanabhan

      Pages 199-206

   3. Design of Retroviral Expression Vectors

      • Kenine E. Comstock, Nathaniel F. Watson, John C. Olsen

      Pages 207-222

   4. Use of Defective Herpes-Derived Plasmid Vectors

      • Filip Lim, Dean Hartley, Philip Starr, Song Song, Phung Lang, Linda Yu et al.

      Pages 223-232

7. Expression in Nonmammalian Higher Eukaryotes

   1. Use of Baculovirus Expression Vectors

      • David R. O’Reilly

      Pages 235-246

The current explosive progress in molecular biological research can be definitively traced to the development of molecular cloning technology. The ability to insert specific gene sequences into cloning vectors and their subse­ quent expansion is the cornerstone of modem molecular biology. A direct practical outcome of molecular cloning technology is its application to ex­ press specific recombinant genes. Currently, recombinant gene products are used in a wide spectrum of applications, including gene therapy, production of bioactive pharmaceuticals, synthesis of novel biopolymers, in agriculture and animal husbandry, and so on. A fundamental requirement for successful recombinant gene expression is the design of the cloning vector and the choice of the host organism for expression. Recombinant Gene Expression Protocols grows out of the need for a laboratory manual that provides the reader the background and rationale, as well as the practical protocols for the preparation of "expression constructs" and their introduction into appropriate host cells and/or organisms. The chap­ ters in this book are grouped by their expression hosts, including E. coli, yeast, mammalian cells, nonmammalian eukaryotes such as plants, Xenopus, and insects, as well as in transgenic organisms. In-depth information is presented on the important characteristics of expression cloning vectors and the various methods for efficiently introducing expression constructs into target cells and/ or organisms. Throughout Recombinant Gene Expression Protocols, the authors have consistently striven for a balanced presentation of both background informa­ tion and actual laboratory details.

• Thomas Jefferson University, Philadelphia

  Rocky S. Tuan

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