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Development of a tightly-regulated tetracycline-dependent transcriptional activator and repressor co-expression system for the strong induction of transgene expression.
| Content Provider | Europe PMC |
|---|---|
| Author | Hosoda, Hiroshi Miyao, Takahisa Uchida, Shusaku Sakai, Shinsuke Kida, Satoshi |
| Abstract | The teteracycline (Tc)-dependent and -inducible transcriptional activator (rtTA) system has been used to express regulated transgene expression in vitro and in vivo. However, previous reports have demonstrated that, even in the absence of Tc, the rtTA binds weakly to the tetracycline response element (TRE), leading to a low level of background activity. In order to reduce the leaky gene expression induced by rtTA, we previously established a tightly regulated system (A-IRES-R system) that makes use of both the rtTA (A) and a Tc-dependent repressor (TetR-Kruppel-associated box; KRAB) (R). In addition, others have described a transactivator rtTA2-M2 (M2) that displays higher sensitivity to Dox than rtTA. In this study, to further develop the A-IRES-R system, we generated a derivative Tc system (M2-IRES-R system) that co-expresses both rtTA-M2 and TetR-KRAB from a single vector. We show that compared to the A-IRES-R system, the M2-IRES-R system leads to a greater level of induced TRE-mediated transcription in the presence of doxycycline (Dox) and yet displays a similar level of basal TRE-mediated transcription in the absence of Dox. Furthermore, the M2-IRES-R system also displays less leaky gene expression in the absence of Dox compared to rtTA-M2 and rtTA systems. Taken together, our results suggest that the M2-IRES-R system enables to tightly regulate and highly induce the expression of transgene compared to other systems. Electronic supplementary material The online version of this article (doi:10.1007/s10616-011-9335-z) contains supplementary material, which is available to authorized users. |
| Related Links | https://europepmc.org/backend/ptpmcrender.fcgi?accid=PMC3081044&blobtype=pdf |
| ISSN | 09209069 |
| Volume Number | 63 |
| DOI | 10.1007/s10616-011-9335-z |
| PubMed Central reference number | PMC3081044 |
| Issue Number | 3 |
| PubMed reference number | 21336964 |
| Journal | Cytotechnology |
| e-ISSN | 15730778 |
| Language | English |
| Publisher | Springer Netherlands |
| Publisher Date | 2011-02-20 |
| Publisher Place | Dordrecht |
| Access Restriction | Open |
| Rights License | © Springer Science+Business Media B.V. 2011 |
| Subject Keyword | rtTA-M2 Tight regulation Transgenic technique Basal transcription |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Clinical Biochemistry Bioengineering Biomedical Engineering Biotechnology |
