Rearrangement and mutagenesis of a shuttle vector plasmid after passage in mammalian cells.

Abstract

A shuttle vector plasmid that contains sequences from SV40 plasmid pBR322 and a bacterial marker gene, galactokinase, was constructed. After replication in [African green monkey kidney] cells permissive for virus progeny, plasmid DNA was introduced into a galactokinase-deficient bacterial [Escherichia coli] strain, and the relative frequency of colonies with plasmids but without galactokinase activity was determined. This assay showed that 1% of the plasmids were defective after passage in the mammalian cells. Individual mutant plasmids were examined and found to contain deletions, duplications, point mutations and insertions of cell DNA.