A Two-Plasmid System for Transient Expression of cDNAs in Primate Cells

Abstract

This two-plasmid system for transient gene expression can be used in a wide variety of primate cells. It consists of a cDNA expression vector and a helper plasmid. The cDNA cloned in the expression vector is transcribed by the powerful major immediate-early promoter of murine cytomegalovirus. A segment of the rabbit β-globin gene placed downstream of the cDNA provides signals for splicing and polyadenylation. The helper plasmid provides SV4O T antigen and adenovirus VA RNA. The T antigen induces replication of the expression vector, which contains an SV4O origin of replication, and VA RNA enhances translation of the transcribed mRNA. In monkey kidney cells, with human tissue-type plasminogen activator (t-PA) cDNA as reporter gene, the helper plasmid boosted t-PA production 30-fold and up to 500 ng/ml t-PA accumulated in the medium in the 5 days following transfection of the two plasmids. In nonprimate cells the helper plasmid stimulated expression 3- to 5-fold.