Inducible high-level expression vector for mammalian cells, pEF-LAC carrying human elongation factor 1alpha promoter and lac operator.
We have constructed an inducible high-level expression vector, pEF-LAC. pEF-LAC has a modified human polypeptide chain elongation factor 1alpha (EF-1alpha) promoter containing three lactose operator sequences. Using the cat reporter gene, we characterized the transcriptional activity of pEF-LAC. In the transient transfection of NIH3T3 and BaF3 cells, the transcriptional activity of pEF-LAC was higher than that of the original human elongation factor 1alpha promoter, simian virus 40 (SV40) promoter, and Rous sarcoma virus (RSV) long terminal repeat (LTR). Cotransfection of the lactose repressor expression plasmid effectively suppressed the promoter activity of pEF-LAC, and the activity was fully recovered by addition of isopropyl beta-D-thiogalactopyranoside (IPTG). Even in the stable transfection of Rat-1 cells, the promoter activity of the integrated pEF-LAC was much higher than that of the RSV-LTR and regulated in an IPTG-dependent manner. These results suggest that pEF-LAC is a useful vector for the inducible high-level expression of the cloned gene in a variety of mammalian cells.
Edamatsu
H
H
Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Midori-ku, Yokohama, Japan.
Kaziro
Y
Y
Itoh
H
H
D016475:3T3 Cells; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002460:Cell Line; D003001:Cloning, Molecular; D029968:Escherichia coli Proteins; D015870:Gene Expression; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D007763:Lac Operon; D056968:Lac Repressors; D051379:Mice; D009875:Operator Regions, Genetic; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D051381:Rats; D012097:Repressor Proteins / Q000235:genetics; D014162:Transfection
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D056968:Lac Repressors; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors; D012097:Repressor Proteins