Abstract
Publisher Summary This chapter discusses the site-directed mutagenesis and expression of coagulation factors VIII and V in mammalian cells. It also reviews different strategies and provides protocols that have proved successful for the expression and characterization of these proteins in mammalian cells. The two host–vector systems that have proved most successful for obtaining high-level expression of factor V and factor VIII in mammalian cells are discussed and the methods for oligonucleotide site-directed mutagenesis and analysis of factor V and factor VIII transfected cells are described in the chapter. The expression of foreign genes in mammalian cells has become an increasingly important technology in biochemistry and molecular and cellular biology; the choice of a particular expression strategy is dependent on the particular objectives of the study. The criteria in evaluating what expression scheme to employ include (1) the method desired for introducing the foreign gene into the cell, (2) the particular requirement for a specific cell type in which to obtain expression, (3) the amount of protein expression required to achieve the goals of the study, and (4) the particular need for an inducible vector to obtain expression of proteins that are potentially toxic.
Published Version
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