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Description

 

Protein expression is an increasingly important tool for research on gene function. What is needed is not just a lab manual providing established methods as well as the latest state-of-the-art protocols, but also clear advice on what expression system to choose when.

Expression Systems: Methods Express uniquely fills this need. It covers expression across a broad range of systems, including the following. *Baculovirus expression vectors

*CHO cells

* E. coli

*HEK293-EBNA1 cells

* Lactococcus lactis and other gram positive bacteria

* S. cerevisiae

*transfected insect cells

 

* Pichia pastoris

*mammalian cells using BacMam viruses

*lentiviral vectors

*wheat germ cell-free system

 

The book takes the reader through how to make an informed choice of appropriate system, taking into account the protein target, the time involved, the ultimate use of the expressed protein, and the laboratory equipment required. It also provides step-by-step methods for each system. In addition, the book describes the optimisation of expression strategies, expression engineering using ribosome display, and how to select protein variants with improved expression.

 

Every chapter discusses the merits and limitations of the approaches available, describes the key techniques in full practical detail, and provides sensible advice for immediate use at the bench.

 

Contents

 

1. Expression strategy (Michael Dyson)

 

2. Protein expression in Escherichia coli (Rosalind Kim)

 

3. Expression engineering of synthetic antibodies using ribosome display (Matthew DeLisa and Lydia M. Contreras Martinez)

 

4. Refolding proteins from inclusion bodies (Renaud Vincentelli)

 

5. Selection of protein variants with improved expression using GFP-derived folding and solubility reporters (Geoffrey Waldo and St 'e9phanie Cabantous)

 

6. Protein expression in the wheat germ cell-free system (Yaeta Endo and Tatsuya Sawasaki)

 

7. Saccharomyces cerevisiae ; A microbial eukaryotic expression system (Christine Lang)

 

8. Expression of proteins in Pichia pastoris (Geoff and Joan Lin-Cereghino and Wilson Leung)

 

9. Improved baculovirus expression vectors (Linda King, Richard Hitchman and Robert Possee)

 

10. Transient transfection of insect cells for rapid expression screening and protein production (Robert Novy et al.)

 

11. Generation of stable CHO cell lines for protein expression (Zhijian Lu et al.)

 

12. Transient expression in HEK293-EBNA1 cells (Yves Durocher, Roseanne Tom and Louis Bisson)

 

13. Nisin- and subtilin-controlled gene expression systems for Gram-positive bacteria (Oscar Kuipers and Jan Kok)

 

14. Protein expression using lentiviral vectors (Bernard Massie, Renald Gilbert and Sophie Broussau)

 

15. Expression in mammalian cells using BacMam viruses (Yu-Chen Hu and Hsiao-Ping Lee)

 

List of suppliers

 

Index