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

✍ Scribed by F.A.M. Asselbergs; P. Grand

• Publisher: Elsevier Science
• Year: 1993
• Tongue: English
• Weight: 447 KB
• Volume: 209
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1993.1128

✦ Synopsis

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 (\beta)-globin gene placed downstream of the cDNA provides signals for splicing and polyadenylation. The helper plasmid provides SV40 T antigen and adenovirus VA RNA. The T antigen induces replication of the expression vector, which contains an SV40 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 \mathrm{ng} / \mathrm{ml} t-P A) 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. aci 1993 Academic Press, Inc.