Variability of transcriptional regulation after gene transfer with the retroviral tetracycline system.
Inducible transcription and position-independent expression are critical issues after gene transfer. To gain insight into the amount of variability of transcriptional regulation due to random proviral integration, we analyzed a total of 200 C6 glioma and rat-1 fibroblast clones retrovirally infected with the conventional and reverse tet systems where a luciferase reporter gene was placed under control of a tetracycline-responsive promoter. Repressed luciferase activities differed by up to 81000-fold among individual clones. Repressed activities close to baseline levels were observed in eight clones, all of them transduced with the conventional tet system. Regulation factors ranged from less than two-fold (indicating absence of regulation), observed in 17 clones to 90-fold. Regulation was higher with the conventional tet system as compared with the reverse tet system. Our data show that even under these standardized conditions there was a very high variability in absolute expression levels and regulability between individual clones, and they suggest that homogeneous transcriptional regulation in a cellular population remains a challenge for research in biotechnology.
Paulus
W
W
Institute of Neuropathology, University of Münster, Domagkstr. 19, D-48129, Münster, Germany. werner.paulus@uni-muenster.de
Baur
I
I
Keyvani
K
K
Senner
V
V
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D008156:Luciferases / Q000235:genetics; D051381:Rats; D012190:Retroviridae / Q000235:genetics*; D013754:Tetracyclines; D015533:Transcriptional Activation / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D016662:Virus Integration
D000900:Anti-Bacterial Agents; D013754:Tetracyclines; D008156:Luciferases