title: Development of single-vector Tet-on inducible systems with high sensitivity to doxycycline.
abstract: Single-vector Tet-on systems were developed to enable the tight regulation of transgenes in mammalian cells with a low dosage of doxycycline. Both the regulatory and the responsive units were integrated in a single vector and separated by a short DNA segment (214 bp). In the developed single-vector Tet-on systems, a high level of expression of the transgene can be induced by doxycycline at a concentration of as low as 1 ng/ml, which is 500-1,000 times lower than that usually utilized in other Tet-on systems. The single-vector Tet-on system developed here exhibited 3.5-10.8 times greater inducibility of the transgene in response to doxycycline than did a dual-vector system from a commercial source. Further studies indicate that the basal activity of Tet-on systems depends greatly on the strength of the promoter that controls the transactivator. The basal activity of Tet-on systems was high when the transactivator that was directed by the human cytomegalovirus promoter, and it was almost undetectable when the transactivator was placed under the control of a moderate strength mouse mammary tumor virus promoter. Moreover, the introduction of selectable markers allows the developed single-vector Tet-on systems to facilitate the generation of conditional transgenic cells and animals with high inducibility, low basal activity and detrimental effects of the long-term administration of doxycycline.
authors: Department of Biological Science and Technology, National Chiao Tung University, 75 Po-Ai Street, Hsinchu 300, Taiwan, ROC.
authors: Jiun-Shuan
authors: JS
authors: Chao
authors: Chun-Chieh
authors: CC
authors: Chao
authors: Chu-Li
authors: CL
authors: Chang
authors: Yi-Rong
authors: YR
authors: Chiu
authors: Chiun-Jye
authors: CJ
authors: Yuan
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D003587:Cytomegalovirus / Q000235:genetics; D004318:Doxycycline / Q000494:pharmacology*; D005822:Genetic Vectors / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D020218:Response Elements / Q000187:drug effects; D013752:Tetracycline / Q000494:pharmacology*; D015534:Trans-Activators / Q000235:genetics; D014162:Transfection; D019076:Transgenes
chemical_list: D000900:Anti-Bacterial Agents; D015534:Trans-Activators; D013752:Tetracycline; D004318:Doxycycline