Prokaryotic in vivo expression systems

Abstract

Abstract Bacterial expression systems are commonly used for production of heterologous gene products of both eukaryotic and prokaryotic origin. This chapter will discuss strategies and provide protocols for the expression of heterologous proteins in Escherichia coli, which is the bacterial system that is most widely and routinely used for this purpose. While many specialized vectors are available for gene expression in other Gram-negative and Gram-positive bacteria, such systems are usually employed for the genetic analysis and manipulation of specific organisms, rather than for high level expression of proteins to be purified and used for research. The emphasis of this chapter is on protocols that are suitable for producing proteins in the laboratory for research purposes. There are additional considerations in the selection of systems for large scale production (i.e. hundreds of litres or more of culture) and for production of proteins for clinical investigation or commercial use. In this chapter we briefly review some of the major factors to consider in selecting an E. coli expression strategy and provide protocols that have been successful in our laboratories in a variety of expression applications. Additional details and extensive literature citations are available in several recent reviews (1-6).