PuSH - Publikationsserver des Helmholtz Zentrums München

Bornkamm, G.W. ; Berens, Ch.* ; Kuklik-Roos, C. ; Bechet, J.-M.* ; Laux, G. ; Bachl, J. ; Korndörfer, M. ; Schlee, M. ; Hölzel, M. ; Malamoussi, A. ; Chapman, R.

Stringent doxycycline-dependent control of gene activities using an episomal one-vector system.

Nucleic Acids Res. 33, 1-11:e137 (2005)
Verlagsversion Volltext DOI PMC
Open Access Gold
Creative Commons Lizenzvertrag
Conditional expression systems are of pivotal importance for the dissection of complex biological phenomena. Here, we describe a novel EBV-derived episomally replicating plasmid (pRTS-1) that carries all the elements for conditional expression of a gene of interest via Tet regulation. The vector is characterized by (i) low background activity, (ii) high inducibility in the presence of doxycycline (Dox) and (iii) graded response to increasing concentrations of the inducer. The chicken beta actin promoter and an element of the murine immunoglobin heavy chain intron enhancer drive constitutive expression of a bicistronic expression cassette that encodes the highly Dox-sensitive reverse tetracycline controlled transactivator rtTA2(S)-M2 and a Tet repressor-KRAB fusion protein (tTS(KRAB)) (silencer) placed downstream of an internal ribosomal entry site. The gene of interest is expressed from the bidirectional promoter P(tet)bi-1 that allows simultaneous expression of two genes, of which one may be used as surrogate marker for the expression of the gene of interest. Tight down regulation is achieved through binding of the silencer tTS(KRAB) to P(tet)bi-1 in the absence of Dox. Addition of Dox releases repression and via binding of rtTA2(S)-M2 activates P(tet)bi-1.
Altmetric
Weitere Metriken?
[➜Einloggen]
Zusatzinfos bearbeiten [➜Einloggen]
Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter EPSTEIN-BARR-VIRUS; MAMMALIAN-CELLS; C-MYC; TRANSGENE EXPRESSION; RIBOSOME BIOGENESIS; ADENOVIRUS VECTOR; EUKARYOTIC CELLS; TIGHT CONTROL; TETRACYCLINE; ACTIVATION
ISSN (print) / ISBN 0305-1048
e-ISSN 1362-4962
Zeitschrift Nucleic Acids Research
Quellenangaben Band: 33, Heft: 16, Seiten: 1-11, Artikelnummer: e137 Supplement: ,
Verlag Oxford University Press
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Clinical Molecular Biology and Tumor Genetics (K.MOLBI)